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Effects of log storage on water quality and microbiology in experimental enclosures in Babine Lake, British… Wentzell, Paula Lanette 1987

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EFFECTS OF LOG STORAGE ON WATER QUALITY AND MICROBIOLOGY EXPERIMENTAL ENCLOSURES IN BABINE LAKE, BRITISH COLUMBIA.  by PAULA LANETTE WENTZELL •a •  A THESIS SUBMITTED IN PARTIAL FULFILMENT O F THE REQUIREMENTS FOR THE DECREE O F MASTER O F APPLIED SCIENCE  in THE FACULTY O F GRADUATE STUDIES Department of Civil  We  accept to  Engineering  this thesis as conforming  the required standard  THE UNIVERSITY O F BRITISH  COLUMBIA  April 30, 1987  ©  Paula Lanette Wentzell, 1987  In  presenting this  degree  at  the  thesis in partial  THE  UNIVERSITY  fulfilment  OF  BRITISH  of  the  requirements  COLUMBIA, I agree  for that  an advanced the  Library  shall make it freely available for reference and study. I further agree that permission for extensive  copying of this thesis for scholarly purposes may be  Head  Department  of  my  or  by  his  or  her  representatives.  It  granted by  the  is understood that  copying or publication of this thesis for financial gain shall not be allowed without my written permission.  Department  of Civil Engineering  THE UNIVERSITY OF BRITISH COLUMBIA 2075 Wesbrook Place Vancouver, Canada V6T 1W5  Date: April 30,  1987  ABSTRACT  The environmental  impacts of log storage on water quality and microbiology  in Babine Lake, B.C. were studied in experimental  enclosures. The enclosure work  was a two phase study, with data collected during the summers of 1984 and 1985. The experiments  performed  in the field season of 1984 involved the addition of  mixed lodgepole pine (Pinus contorta) and white spruce (Picea glauca) bark debris, in different amounts, to the enclosures to examine effects on water quality (dissolved oxygen, lignins-tannins, total organic carbon, total inorganic carbon, pH, and alkalinity),  bacterial activity (heterotrophy),  and phytoplanktonic primary production.  The 1985 study attempted to simulate a shallow water log storage facility by adding logs to the enclosures. A comparison was made of differences in water quality (including the above measurements plus chemical oxygen demand, nutrients, volatile fatty acids, and carbohydrates), and sestonic bacterial and phytoplanktonic algal populations (enumeration  and biomass calculations) between  (i) lodgepole pine  and white spruce, and (ii) the number of logs per enclosure. The bark enclosure study resulted in organic enrichment  of the enclosure  ecosystem. More than 50% of the total organic carbon (TOC) was attributed to leached lignins and tannins. The leachate was capable of modifying microorganism production. Phytoplanktonic algal biomass, measured by chlorophyll _a, was completely eliminated  at high concentrations of bark. Short term laboratory studies with bark  leachate inhibited microbial activity of natural aquatic bacteria, however, from the enclosure experiments,  it was apparent that with time a population of microbes  would develop to utilize the chemically complex leachate. The presence of utilizable organic material (i.e. wood sugars) and an active microbial community resulted in a significant decrease in the dissolved oxygen levels.  ii  The results of the log study found significant decreases in the amount of organic extractives leached from logs compared to the TOC concentration in the bark experiment.  For example, the T O C concentrations in the 5 log enclosures were  approximately 20 mg/l by the end of the 25 day test period ( s  10% was lignin  and tannin carbon) ; this value was significantly smaller compared to the bark experiments, where the TOC levels in the heavy bark treatment (20  kg) reached —  400 mg/l after 25 days. A comparison between the bark and log experiments an equivalent bark dry weight  (on  basis) found water quality degradation by bark  leachates more severe than log leachates. The log leachate stimulated bacterial production and did not adversely affect phytoplankton biomass. The increase in bacterial production, a direct result of (i) an available carbon source, determined by TOC measurements and the very low concentrations of volatile fatty acids and reduced carbohydrates (rapidly utilized), and (ii)  an increase in water temperature  caused a decrease in dissolved oxygen levels.  The spruce log enclosures for both June and July had higher heterotrophic uptake rates than the pine log enclosures. This indicated a difference in the chemical composition of the spruce and pine log leachates. This chemical difference was detected in the TOC and C O D measurements and the microbial uptake kinetics experiment.  These measurements found that the pine log leachate was more readily  degraded (high affinity, V  m a x  /  K t  )  by a heterotrophic population than the spruce  leachate, and that this difference could account for the higher dissolved oxygen demand of the pine log enclosures compared to the spruce logs. Detailed chemical analyses of the leachates in the presence of microbial inhibitors may help to delineate this discrepancy between the spruce heterotrophic uptake rates and the chemical and bacterial measurements of the pine log leachates. The enclosure studies indicated that log storage in a shallow, poorly flushed, littoral area of a lake would possibly result in an accumulation of organic  iii  components leached from the log and bark debris. The organic enrichment aquatic ecosystem would contribute thus, negatively  of the  to a potential decrease in dissolved oxygen,  affecting fish habitat, but could increase microbial production.  iv  Table of Contents  ABSTRACT  ii  LIST OF TABLES  ix  LIST OF FIGURES  xi  ACKNOWLEDGEMENTS  xiii  1. INTRODUCTION  1  2. LITERATURE REVIEW  3  2.1 Location, morphology, and limnology of Babine Lake  3  2.2  5  History of bark and log leaching experiments  2.3 Chemical composition of wood and bark extractives 3. EXPERIMENTAL DESIGN, METHODS, AND MATERIALS 3.1 Experimental design of 1984 bark study  9 16 16  3.1.1 Experimental treatments  16  3.1.2  18  Enclosure preparation  3.2 Experimental design of 1985 log study 3.2.1 Enclosure preparation  18 19  4. METHODS OF ANALYSIS  22  4.1 Water quality  22  4.1.1 General water quality measurements  22 Dissolved oxygen and temperature  22 pH and alkalinity  22  4.1.2  Organic pollution indicators  23 Total organic and inorganic carbon  23 Chemical oxygen demand  24 Lignins and tannins  25  4.1.3  Nutrients and organic substrates  25  v  4.2  4.3 Total Kjeldhal nitrogen  25 Total phosphorus  25 Total reducing carbohydrates  26 Volatile fatty acids  26  Bacterial population  27  4.2.1 Biomass and numbers  27  4.2.2  Heterotrophy  28  4.2.3  Heterotrophic uptake kinetics  29 Bark leachate inhibition experiments  29 Bacterial uptake studies with log leachates  31  Phytoplanktonic algal population  31  4.3.1 Biomass and enumeration  31  4.3.2  Phytoplanktonic primary production  34  4.4 Comparison between the bark and log studies  34  4.5 Statistical analysis  35  5. RESULTS  36  5.1 Enclosure experiments with bark - 1984  36  5.1.1 Comparison of the lake and control data  36  5.1.2  37  Water quality General water quality measurements  37 Organic pollution indicators  41  5.1.3  Bacterial population  44 Heterotrophy  44 Bark leachate inhibition experiments  47  5.1.4  Phytoplanktonic algal population  47 Phytoplanktonic biomass  4 Phytoplanktonic primary production  vi  7  47  5.2 Enclosure experiments with logs - 1985  48  5.2.1 Comparison of the lake and control data  48  5.2.2 Water quality  51 General water quality measurements  51 Organic pollution indicators  55  5.2.3 Bacterial population  62 Bacterial numbers, biomass, and heterotrophy  62 Heterotrophic uptake kinetics  68  5.2.4 Phytoplanktonic algal population  71  6. DISCUSSION  73  6.1 Enclosure experiments with bark - 1984  75  6.1.1 Comparison of control and lake results for bark and log studies  75  6.1.2 Water quality  76  6.1.3 Heterotrophy  83  6.1.4 Phytoplanktonic algal population  85  6.2 Enclosure experiments with logs - 1985 6.2.1 Water quality  86 87 Dissolved oxygen (DO)  87 Total organic carbon (TOC) and chemical oxygen demand (COD)  88  6.2.2 Bacterial population  91 Numbers, biomass and heterotrophy  91 Kinetic uptake study  95  6.2.3 Phytoplankton biomass 6.3 Comparison between  98  the enclosure studies and the 3 year  field study by Westwater  Research  98  7. CONCLUSION  102  vii  8. APPENDIX A  1  0  4  9. APPENDIX B  1  0  9  1  1  2  1  1  3  10. ABBREVIATIONS  •  11. REFERENCES  viii  List of Tables  1 Main groups of wood extraneous Fengel and Wegener, 1985).  compounds (Buchanan, 1975 ;  2 Chemical composition of bark extractives Fengel and Wegener, 1985).  1 Summary of the  2 Experimental  1984  enclosure  design of the 1985  (Jensen et a!. 1975  ;  experiments.  enclosure study.  1 Process of preservation for analytical procedures for water quality, bacterial, and phytoplankton studies.  2 Volumes of solutions added to the bark leachate reaction flasks in microbial inhibition studies.  3 Volumes of solutions added to the log leachate reaction flasks in microbial activity studies.  1 Changes in dissolved oxygen, pH, and alkalinity over time in the 1984 bark enclosure studies done at Babine Lake.  2 Temporal changes in the concentration of organic and inorganic carbon compounds in the 1984 bark enclosure experiments at Babine Lake.  3 Temporal changes in heterotrophy and phytoplanktonic biomass and productivity in the 1984 bark enclosure experiments at Babine Lake.  4 A comparison between the pine log control enclosure and Babine Lake of significant changes in several water quality and bacterial measurements.  5 A comparison of significant changes in heterotrophy and bacterial numbers and biomass between the spruce log control enclosure and Babine Lake, 1985.  6 A summary of means and standard deviations of general water quality measurements for the 1985 pine and spruce log study at Babine Lake. ix  5.7  Effects of pine and spruce logs on dissolved oxygen in the June log enclosure experiments.  1985  5.8 Temporal changes in total organic carbon in the pine and spruce log enclosure studies at Babine Lake, 1985.  5.9 Temporal changes in bacterial enclosure studies of 1985,  numbers in the pine and spruce log Babine Lake.  5.10  Heterotrophic uptake kinetic values obtained on Days 0 and 10 from the 5 spruce and pine log enclosures.  5.11  Temporal changes in chlorophyll _a_ concentration in the pine and spruce log enclosure studies done at Babine Lake, 1985.  6.1  A calculation of the dry weight of bark added to the 1984 bark and 1985 log enclosure studies for water quality comparisons.  6.2  A comparison of water quality conditions in enclosures with 1 kg bark (July, 1984) and 1 log (July, 1985) over an 11 day period.  6.3 A comparison of water quality measurements researchers bark leaching experiments.  between several  6.4  Calculations to determine the % of total organic carbon (TOC) comprised of lignins and tannins (L-T) and resultant chemical oxygen demand (COD).  6.5  A comparison of absolute inorganic carbon concentrations between alkalinity and TIC for the 20 kg bark enclosure of 1984.  6.6  Dissolved oxygen saturation for the Babine Lake.  1985  log enclosure studies at  6.7 A comparison of the uptake of carbon per ug of biomass for the 1985 log experiments. 14  bacterial  List of Figures  2.1  Location of log transportation  study areas in Babine Lake.  3.1  Diagram of an experimental enclosure.  5.1  Temporal changes in dissolved oxygen in the 20 kg bark enclosure at Babine Lake ; control = no bark.  5.2 Temporal changes in pH in the 20 kg bark enclosure experiment Babine Lake, 1984 ; control = no bark.  at  5.3 Temporal changes in alkalinity in the 20 kg bark enclosure experiment at Babine Lake, 1984 ; control = no bark.  5.4 Temporal changes in total organic carbon in the 20 kg bark enclosure at Babine Lake, 1984 ; control = no bark.  5.5 Temporal changes in the total inorganic carbon levels in the 20 kg bark enclosure at Babine Lake, 1984 ; control = no bark.  5.6 Temporal changes in lignins and tannins concentration for the 20 kg bark enclosure at Babine Lake, 1984 ; control = no bark.  5.7 Temporal changes in heterotrophic uptake of labelled glucose in the 20 kg bark experiment at Babine Lake, 1984 ; control = no bark.  5.8 Temporal changes in chlorophyll _a_ concentration in the 20 kg bark enclosure at Babine Lake, 1984 ; control = no bark.  5.9  Effects of spruce and pine logs on dissolved oxygen in enclosures for the 1985 July log study period ; number of logs in treatment are given in parentheses.  5.10  Temporal changes in C O D concentration for the pine and spruce log enclosure studies done at Babine Lake, 1985 ; number of logs in treatment are given in parentheses.  5.11  Linear regression relationship between changes in C O D and dissolved oxygen concentrations for the Babine Lake pine log experiments of 1985.  xi  5.12  5.13  5.14  5.15  Linear regression relationship between changes in C O D and dissolved oxygen concentrations for the spruce log experiments Babine Lake, 1985.  at 61  Changes in L-T concentrations in the 1985 spruce and pine log experiments at Babine Lake ; number of logs in treatment are given in parentheses.  62  Changes in bacterial activity and biomass in the 1985 enclosure experiments done at Babine Lake.  64  pine log  Changes in bacterial activity and biomass in the spruce log enclosure experiments of 1985 at Babine Lake.  65  5.16(A) Bacterial biomass on initiation of the 5 pine logs enclosure at Babine Lake, 1985 ; X1562. 5 urn  67  5.16(B) Bacterial biomass on day 25 of the 5 pine logs enclosure experiment at Babine Lake, 1985 ; X1562. 5 urn  67  5.17  Lineweaver-Burk plot of S/V  vs S on Day 10 for the 5 log pine  and spruce enclosures an8 the control.  6.1  70  Aerial photograph of the Morrison dump site in mid-winter, 1984.  6.2 Aerial photograph of the Morrison dump site on May 14,  xii  1985.  100  101  ACKNOWLEDGEMENTS  This project was funded under Section 88.2  of the B.C. Forests Act. The  students were funded by a GREAT Scholarship to E. Power from the B.C. Science Council and both a YEP university student summer grant and NSERC operating  grant  for P. Wentzell. I would like to thank  Doug Reynolds, Brett Hall, and Jim Mclver for helping  obtain logs for the experimental  enclosure studies and Pat Ogawa of Houston  Forest Products for donating lumber for construction of the experimental enclosures. Norm Aason, Ham Brown, and Jim Barker of the Granisle Power Boat Club are acknowledged for providing housing and marina facilities. I also thank Stewart Barnetson and Colin Harrison, managers of the Fulton River spawning channels, for providing laboratory facilities to conduct chemical analysis and store our equipment from 1984  to  1985.  The assistance given by Susan Liptak and Paula Parkinson of the Environmental Engineering Laboratory, U.B.C., and Ralph Daley previously of the National Water Research Institute, West Vancouver, in water quality  measurements  and bacterial analysis is greatly appreciated. I thank Craig Peddie for his help in computer word processing, Tom Northcote  for his valued opinions and comments  editing this manuscript, and Ken Hall for his patience. And finally, I wish to express sincere gratitude  to Beth Power for her  helping hand in gathering samples, running analysis, constructive criticism, patience, and putting up with a stubborn and sometimes cranky associate.  xiii  1.  In 1982 beetle infestation  INTRODUCTION  the British Columbia Ministry of Forests, in response to a pine in the forests of the Morrison Arm region of Babine Lake,  decided to accelerate logging plans in an attempt to salvage the infested timber. A transportation  system was proposed that would involve the dumping, storage, towing,  and dewatering  of logs. The Department  of Fisheries and Oceans reviewed  the  proposal and gave its conditional approval provided that a three year study be undertaken  to determine  the effects of log storage and transportation  on fish  populations and habitat of Babine Lake, with emphasis on sockeye salmon. The Westwater  Research Center received funding under Section 88.2  of the  B.C. Forests Act provided to the University of British Columbia as a grant from Houston Forest Products Ltd. to study the environmental  impacts of log handling  and storage activities on Babine Lake, British Columbia. The main objectives of the student thesis projects were to experimentally  test hypotheses about the chemical  and biotic effects of log storage in a shallow limnetic environment. This research was done in the summers of 1984 Westwater  and 1985  and conducted in conjunction with  Research Centre's three year study- of log transporation  impacts in Babine  Lake. The hypothesis that littoral water quality and microbiology populations are affected  by log storage activities was tested in experimental  enclosures. Experimental  enclosures were used because conditions inside these large bags can be manipulated  and easily monitored, as opposed to in situ work where  experimentation  is difficult.  Wood and bark debris, and log rafts were placed in the enclosures to simulate conditions that could occur when logs are handled and stored in shallow  1  2  water areas. As one of the students in the project,  1 studied changes in (i) water  quality conditions (dissolved oxygen, nutrients, lignins and tannins, organic carbon, chemical oxygen demand, pH, and alkalinity), (iii)  phytoplanktonic  (ii) microbial activity and biomass, and  primary production and biomass. Beth Power, a graduate  in zoology, (i) studied the effects of wood and bark debris on the  student  zooplankton  population, (ii) performed short term bioassay tests to examine the toxicity of log and bark leachate on Daphnia pulex, sockeye fry (Oncorhynchus nerka), and rainbow trout (Salmo gairdneri), and (iii)  examined the effect of bark and log  leachate on the ability of sockeye fry to avoid predators. The results of this project should provide some of the first direct and manipulative  experimental  information  regarding effects of log storage on lake water  quality and related effects on bacterial and algal processes important  to salmonid  survival and reproduction. This research should help greatly in predicting log handling impacts on poorly circulated littoral areas of lakes. The following section is a literature review on the limnology and associated lake dynamics of Babine Lake, previous research concerned with assessing the environmental  impacts from bark, wood debris, and logs, and the chemical  composition of wood and bark.  2. LITERATURE REVIEW  2.1  LOCATION, MORPHOLOGY, AND LIMNOLOGY OF BABINE LAKE  Babine Lake is the largest natural lake in the province of British Columbia. The lake is one of the major sockeye salmon producers in the province, and presently 90% of the Skeena River sockeye originate in tributaries lake (Levy and Hall, 1985). Babine Lake is a long (150 elevation  surrounding the  km), narrow lake with an  of 708 m, located on the Central Interior Plateau of British Columbia, 580  km North of Vancouver at 55° N latitude and 123° W longitude (Figure 2.1).  It  forms one of the major drainage basins in the Skeena River system (Levy and Hall, 1985). It has an area of 490  km , mean depth of 55 m, and a maximum 2  depth  of 186 m (Johnson, 1965). Based on a literature review of the limnology of Babine Lake (Levy and Hall, 1985), and considering trophic state indicators, i.e.  primary  production, chlorophyll _a, and nutrients, the trophic status of the lake best fits the oligotrophic classification. There are several morphometric factors to consider when evaluating possible effects of log storage and transportation  the  activities on Babine Lake. The  small littoral zone combined with a deep mean depth, large volume of water (approximately  27 km  3  ; Narver and Andersen, 1968), and persistant strong winds  (long fetches) do not allow for the formation of a stable thermocline. Therefore, I would postulate that the concentration of materials bark debris dislodged from the transportation  leached from the logs and the  of booms, would not build up to  toxic levels in the epilimnetic zone of the main basin of Babine Lake, but would be dispersed and diluted without significant effect on the aquatic system.  3  4  / Historical Dump Site •  4 FORT  BABINE  BABINE  LAKE./  /  1984  / /  1s  Morrison Arm Dump Site  >. VANCOUVER  1983 1984  2 0 0 miles  1985  (from West, 1978.  p.2)  Existing Northwood Dewatering  Site  1984 TOPLEY 3 LANDING Future Houston Dewatering Site  Existing Northwood Dump Site 984  1983 1984 1985  1983 - pre-impact data 1984 - "extensive post treatment analysis" Historical Dewatering Site  1985 - post-impact data  1984  10  15 miles  Figure 2.1 Location of log transportation study areas in Babine Lake.  5  However,  because the Morrison Arm area of Babine Lake has a sheltered  littoral zone and a shallow mean depth (compared with the main basin of Babine Lake) there exists the possibility of aquatic habitat degradation. Therefore, limnology of Babine Lake is an important  the  factor to consider when assessing the  enclosure results and predicting logging impacts in the lake.  2.2  HISTORY OF BARK AND LOG LEACHING EXPERIMENTS  There are two areas of research concerned with the leaching of  pollutants  from wood and bark. Most of the literature is concerned with industrial processing ; this includes (i) woodwaste  timber  from debarking operations with the bark  and wood debris dumped in landfills or left in piles at the mill site (Asano ef al. 1974  ; Atwater, 1980  et al. 1973  ; Benedict et al. 1974a and 1974b ; Evans, 1973  ; Henricksen and Samdal, 1966  Schermer and Phipps, 1976  ; Slagle, 1976  ; Phipps, 1974  ; Harger  ; Raabe, 1968  ; Sproul and Sharpe, 1970  ;  ; Thomas,  1977), and (ii) the discharge of pulp and paper mill effluent on receiving water quality (Kroner and Moore, 1953  ; Pearson, 1980  ; Poole ef al. 1978). The  remaining literature includes work pertaining to the quantity  and quality of leachates  derived from various hardwood and softwood species, and their effect and benthic communities (Buchanan ef al. 1976 Schaumburg, 1969 ef al. 1976  ; Hansen et al. 1971  ; Ponce, 1974  ; Duval, 1980  ; Pease, 1974  ; Schaumburg, 1973  upon limnetic  ; Graham and  ; Peters, 1974  ; Servici ef al. 1971  ; Peters ; Gellman,  1971). Most of the woodwaste similar in experimental generated  landfill and pulp and paper effluent studies are  design and parameters  measured. The leachate (pollution) ,  by laboratory and field studies is characterized using chemical oxygen  demand (COD), biochemical oxygen demand (BOD), total organic carbon (TOC), pH,  6  odour, alkalinity,  colour, and toxicity tests. Total Kjeldahl nitrogen (TKN), total  phosphorus (TP), and metals are measured in municipal landfills where hogfuel is used as a cover. Most of the laboratory experiments are concerned with analyzing water soluble leachates (extractives) generated by fresh and aged softwood bark in static or constant (continuous) flow lysimeters or both (Benedict et al. 1974a and 1974b ; Henricksen and Samdal, 1966). The softwood species commonly studied includes Douglas fir (Pseudotsuga menziesii), western hemlock (Tsuga heterophylla), lodgepole pine (Pinus contorta), and western red cedar (Thuja plicata). The duration of the experiments (Asano ef al. 1974) 1976)  changed between  the researchers, for example,  10-40  hours  and long term experiments of 22 days (Schermer and Phipps,  to 4 years (Atwater, 1980). The high concentrations of wood and woodwaste  used in the studies  resulted in high concentrations of organic pollutants leached during the  experiment.  For example, Schermer and Phipps (1976) have a C O D value of 8282 mg/l at the terminaton  of their 23 day static leaching study with Douglas fir. Their results were  similar to the data from the short term continuous flow experiments with softwood by Benedict ef al. (1974b), i.e. C O D >  4500 mg/l after 23 days.  Several authors initiated long term studies to determine responsible for leaching and the biodegradability  both the mechanism  of water soluble extractives  (Benedict ef al. 1974a and 19746 ; Henricksen and Samdal, 1966 Schermer and Phipps, 1976 ; Sproul and Sharpe, 1970  ;  ; Thomas, 1977). The long  term static and dynamic (constant flow) studies (minimum involving different species of woodwaste  ; Phipps, 1974  of 90 days), although  and bark, give the following leaching  characteristics : 1. biodegradability of the leachate is a function of the chemical nature of the water soluble compounds (extractives) found in each different species of tree, 2. initial high concentrations of C O D , BOD, total solids, T O C , and colour  7  decrease with time (after =  4 to 10 days) because of (i) a decrease in  readily available near surface water soluble compounds and (ii) a decrease in the concentration gradient or driving force in static experiments, 3. biphasic BOD where the initial stage is characterized by a high BOD (simple wood sugars) mediated, by heterotrophic bacteria and the second stage is represented by a lower BOD (compared to the rapid first stage) due to the slow decomposition by bacteria and fungi of lignin and other chemically complex structures (Benner ef al.  1986). For example, Raabe (1968) and  Kroner and Moore (1953) found high concentrations (>  60%)  of lignin  present in receiving water 20 weeks after initial discharge, 4. the leaching mechanism is a function of the volume of contact water and contact time, i.e. potential BOD and C O D are depleted more rapidly by a large volume of contact water than by a small volume of water.  Existing literature on the effects of log storage on the aquatic environment can be classified into three categories : physical, effects, chemical effects, and biotic effects (Power and Wentzell,  1985). The most noticeable impacts of log handling on  the aquatic environment are the physical changes that occur in the habitat. Chemical changes to water quality induced by log storage have been quantified, but data are generally obtained from laboratory studies. There is information on the effects of log storage on aquatic vegetation, benthic invertebrates, zooplankton, and fish, however, with the exception of the fry feeding experiments conducted by Levy ef al. (1982), most of the research is descriptive. Duval (1980) and Hansen ef al. (1971) provide good literature reviews on log handling and dumping in coastal public waters. Graham and Schaumburg (1969), Pease (1974), Schaumburg (1973), Peters (1974), and Gellman (1971) conducted laboratory and field studies to, firstly, characterize log leachates and, secondly,  8  determine the rate and mechanism of leaching soluble organics from logs and bark. Schaumburg (1973) investigates the magnitude of the pollution problem associated with log handling practices. The results indicate that the open ends and logs without bark are a major source of organic matter (i.e. wood sugars) and a possible pollution problem. For example, after a 7 day leaching period, a 150 year old Douglas fir log section, with bark, had a C O D of 53 mg/l, compared to 189 mg/l from a similar log section without bark. The results of field studies by Schaumburg (1973) and Gellman (1971) indicates leachates from logs held in storage exert BOD and C O D on the holding water, but not in sufficient concentrations to adversely affect water quality.  However,  the distribution and accumulation of logging debris on the sediment can affect epibenthic and benthic communities. Benthal deposits of bark can result in a decrease in abundance of infauna, low dissolved oxygen concentrations within the bark deposits exerted by the BOD of the bark, and, possibly, bark leachates toxic to the infauna (Pease, 1974) Pease (1974) conducted log leachate studies with unaltered sections of western hemlock, Sitka spruce {Picea sitchensis), red cedar, and yellow cedar (Chamaecyparis nootkatensis). The results of Pease's (1974) unaltered log sections are similar to Schaumburg (1973) and Gellman (1971), i.e. significant increases in C O D , BOD, and T O C , and a decrease in pH. Buchanan ef al. (1976) did toxicity bioassays on larval and adult pink shrimp (Pandalus borealis), larval Dungeness crab (Cancer magister) and pink salmon fry (Oncorhynchus gorbuscha) with Sitka spruce and hemlock bark extractives. The bioassay tests have a dissolved oxygen concentration above 6 mg/l. Ponce (1974) also conducted toxicity bioassays with leachate from Douglas fir using guppies (Poecilia reticulata) and steelhead trout (Salmo gairdneri) as the test animals. The study results from Buchanan ef al. (1976) and Ponce (1974) indicates that leachate  9  toxicity is a function of the chemical composition of the water soluble fraction. For example, laboratory bioassays by Peters (1974) with the water soluble fraction of western red cedar heartwood leachate found the extraneous components responsible for respiratory distress leading to death in coho salmon (Oncorhynchus kisutch) fry. Increases in opercular movements in the salmon fry without an increase in oxygen consumption, may be the result of charged compounds (i.e. tropolone) complexing with the iron component of hemoglobin molecules and interfering with respiration (Peters, 1974). Servizi ef al. (1971) studied the effects of Douglas fir, lodgepole pine, Engelmann spruce (.Picea engelmannii), and Alpine fir (Abies lasiocarpa) bark deposits on the survival of sockeye salmon eggs (Oncorhynchus nerka) and alevins in flow through gravel boxes (dissolved oxygen s  4 mg/l). The results indicate that  mortality is a function of Sphaerotilus (filamentous  bacteria) growth (promoted by  leachates from fresh bark) on the gills of the alevins, therefore,  suggesting that the  chemical nature of the leachates was not directly toxic to the test fish at that concentration of bark. The literature to date reveals limited studies on (i) the microorganisms responsible for degrading wood and bark extractives, (ii) microbial activity and growth kinetics, and (iii) poor laboratory simulations of the natural log storage environment. This thesis attempts to simulate an actual log storage facility while maintaining experimental  control of the ecosystem.  2.3 CHEMICAL COMPOSITION OF W O O D AND BARK EXTRACTIVES  The biotic water environment of the enclosures is chemically governed by the materials leached from the wood and bark debris, and the logs. The extent of water quality degradation (i.e. organic enrichment and dissolved oxygen demand) is  10  a function of the quality and quantity of the leachates extracted from the bark and logs. Therefore, knowledge of what is being leached from the logs and bark will help in understanding and interpreting the experimental results. There is a variety of water soluble chemicals contained in wood and bark. It is important to know the chemical nature of the extraneous components leached from the spruce and pine logs because the quantity and quality of aqueous extracts from the wood and bark would determine the extent of water quality degradation. Wise (1959) defines the term extraneous components as " ...a wide variety of organic (and some inorganic) substances that can be extracted with cold water and that are soluble in neutral solvents such as ether, chloroform, the alcohols, benzene, ligroine, and methylene chloride." The cell wall components consist of the structural compounds cellulose, lignin, and polyose (hemicellulose), which form systems insoluble in cold water (20  interpenetrating  °C) and neutral solvents (Wise, 1959). The  structural compounds of coniferous wood, mainly lignin and cellulose, are chemically similar irrespective of tree species ; however, the wood can vary in appearance, specific gravity, and strength (Barton, 1975). All woods contain a wide variety of organic and some inorganic substances that can be extracted with cold water and neutral solvents (Wise, 1959 ; Buchanan, 1975 ; Jensen ef al. 1975). These extraneous compounds are known as extractives and comprise 2% to 15% of coniferous wood (Barton, 1975). More recently, Fengel and Wegener (1984) have classed the organic matter as extractives and the inorganic matter as ash. Extractives are low molecular weight compounds compared to the structural components of mature wood. Extractives impart a distinct set of properties to each tree species and are known as personality chemicals (Fengel and Wegener, 1984 ; Barton, 1975). Wise (1959) states that extraneous compounds serve to  fingerprint  the wood. Some factors that affect the types and amounts of extractives present in  11  a tree are species, season, type of wood (heartwood vs. sapwood) and the distance from the butt of the tree. Some properties affected by extractives are odour, resistance to decay and insect attack, colour, and pulpability (Barton, 1975). Available literature illustrates an obvious difference between chemical components of bark and wood. Fengel and Wegener (1984) state that bark possesses polyphenols and suberin, a lower percentage of polysaccharides, and a higher percentage of extractives than wood. Chemical differences also exist between the inner bark, phloem, and the outer bark, rhytidome. Generally the content of extractives and polysaccharides decreases, and the content of lignin and polyphenolic substances increases from the inner bark to the outer bark (Fengel and Wegener, 1984). The function of the inner bark is to transport assimilates and to serve as a storage organ for food reserves, whereas the outer bark consists of dead cells and forms a protective layer against mechanical and chemical injury (Kurth, 1944). Tables 2.1  and 2.2 lists the major extraneous components found in bark and wood  extractives. The carbohydrates, vitamins, and proteins would provide a utilizable source of carbon which would contribute to a dissolved oxygen demand. Several of the aromatic compounds are chemically complex (i.e. tannins and lignins) and would not cause an immediate dissolved oxygen demand, but would increase water turbidity and colour ; this would cause light attenuation  in the water column. The terpenes  are volatile, oily compounds which leave an oily residue on the surface of water. Resin acids and unsaturated fatty acids in hardboard plant effluent have been shown to be toxic to sockeye salmon (Oncorhynchus nerka) fingerlings if present in the water in concentrations S1  mg/l (Rogers ef al. 1979)  Therefore, it is apparent that  the water soluble compounds of wood and bark could directly or indirectly affect water quality and aquatic microorganisms in the enclosures.  Table  2.1  Main  groups  of  wood  Extractives  extraneous  compounds  (Buchanan,  Description  1975  ;  Location in  1.  Aromatic a.  (phenolic)  lignans  several  dimeric  linked  between  phenylpropane B-C  compounds  atoms  Wegener,  found  1985).  Extraction  tree  predominant  in  roots,  Solvent  heartwood,  foliage,  pine  species  stilbenes  acetone, methanol  exudates  heartwood,  inner  ether  bark  c.  flavonoids  d.  tannins i) h y d r o l z y a b l e  and  derivatives  polyhydroxylic hydrolysis  tannins  acids, ii)  and  compounds  pinosylvin b.  Fengel  condensed  alcohol  leaves  partially  phenols  produces  and  heartwood  gallic  and  ellagic  in  sugars  tannins  bark  soluble  water  alcohol  (phlobaphenes)  2.  natural  Terpenes  from  substances isoprene  butadiene a.  monoterpenes  ;  a-pinene,  chemically  units,  acyclic  gum  derived  alcohol,  and  benzene,  ether  2-methylcyclic  turpentine  of  most  pines  inner  bark,  alcohol,  ether  sapwood b.  oxygenated  monoterpenes  oxygen secreted  c.  diterpene  (resin  acids)  as not  d.  triterpene  e.  sesqui-  f.  tropolones  by  monoterpenes  epithelical  cells  and  occur  acids  heartwood  benzene,  ether  heartwood,  benzene,  ether  sapwood  abundant  leaves,  bark,  exudates 15  and  free  containing  polyterprenes and  thujic  acid  plus  carbon  compounds and  species  exhibit  atoms  found  acidic  of  in the  and  western  red  Cupressaceae  aromatic  cedar ;  properties  heartwood  steam  heartwood  t  volatile  Table  2.1  cont'd  3. Aliphatic acids a. saturated acids b. unsaturated acids  palmitic acid fatty acids are found as triglycerides in the sapwood t  c. fats and oils  ether t  benzene, ether ether  d. waxes 4. Carbohydrates  major components are not extractable (i.e. cell wall compounds)  a. mono-, di-, and polysaccharids i) simple  sucrose  sapwood, inner bark  water  -  fructose  water  -  glucose  sapwood, inner bark sapwood, inner bark, heartwood  -  arabinose  sugars-  heartwood t  b. glycosides  water  water t  c. starch  reserve  supply  t  d. pectic material  cell wall component  t  t  e. cellulose  high molecular  t  t  food  weight polymer of  p-D-glucose f. polyoses  (hemicellulose)  polymers of sugars - hot water  5. Alcohols a. sterols b. aliphatic alcohols  =  no available information  aliphatic rings generally found as esters and glycosides occur as ester components  free and  Table 2.2  Chemical composition of  bark  Extractives  extractives  (Jensen et  al. 1975  ; Fengel and Wegener,  Description  1985).  Extraction  1. Aromatic (phenolic) compounds  Solvent  benzene, ether  a. stilbenes b. polyphenols  ether, hot water classed according to molecular weight and solubility colouring materials ; occur in free state and as glycerides outer bark contains the low molecular weight tannins high molecular weight tannins  i) flavane derivatives ii) hydrolzable tannins iii) condensed tannins (phlobaphenes) iv) polyphenols acids 2. Terpenes and resin acids  triterpenes are the predominant bark acids are secreted in response  alcohol alcohol, water ethanol, hot water 1% NaOH/100 *C  terpenes  to damage  3. Aliphatic acids  benzene, ether  a. saturated acids b. unsaturated acids c. fats d. waxes 4.  Carbohydrates a. cellulose b. hemicellulose i) pectins ii) gums c. mono-, di- and polysaccharides  - not extractable - easily hydrolyzed, however not extracted with indifferent solvents t  t  t -  1.87% spruce  mild acid  t bark and  1.35% pine bark  some are water soluble  Table  2.2 cont'd  5. Alcohols a. sterols  - predominant sterol in pine is a-sitosterol  benzene  6. Vitamins  - vitamins are stored in the fall and depleted by spring vitamins are present in significant amounts  some are water soluble  7. Proteins  - bark acts as storage place for proteins and other assimilates ; amounts vary seasonally  some are water soluble, but most are not  =  no available information  3. EXPERIMENTAL DESIGN, METHODS AND MATERIALS  The enclosures consisted of two parts ; a plastic bag and the float from which the bag was suspended. The bags were cylindrical in shape, with an enclosed bottom, and made of woven polyolefin fabric (1.5  m diameter and 2.1 m  deep). The enclosures contained a volume of 3700 I and were not in contact with the lake sediment, thus there was no exchange of water (Figure 3.1).  The collar  was constructed of black plastic to slow fabric degradation by ultraviolet  light ; the  bag was made of clear, white plastic to allow maximum natural radiation inside the bag. The float was of plywood construction and flotation  was provided by styrofoam  blocks. The bag was suspended from the inside of the float and laced into place (Figure  3.1). Eight identical enclosures were built and moored inside Granisle Power Boat  Club marina between June and August of 1984,  and May and August of 1985. The  marina site was approximately 3 m deep and provided protection from storms and vandalism.  3.1  EXPERIMENTAL DESIGN OF 1984  The experimental  BARK STUDY  design of 1984  attempted  to define a range of lethal and  sublethal leachate concentrations, i.e. 20 kg and 1 kg bark experiments, respectively.  3.1.1  EXPERIMENTAL TREATMENTS  The 1984  field season was concerned primarily with determining the effects  of bark on the enclosure ecosystem. Table 3.1  16  lists the treatments  (amounts of bark  17  plywood  Figure 3.1 Diagram of an experimental enclosure.  18  added to the enclosures) and their duration for the 1984  study period. None of  the treatments were replicated.  3.1.2  ENCLOSURE PREPARATION  The enclosures were initially filled with a Par electric bilge pump. This was later replaced with a 3 horse power Honda water pump. The enclosures were always filled the day before the start of an experiment to allow the system to equilibrate with the surrounding waters. The bark consisted of a mixture of white spruce and lodgepole pine ; the ratio was 40% spruce to 60% pine. The mixture was representative of the ratio of trees being logged and stored at the Morrison Arm storage facility. The bark was fresh and obtained from nearby construction sites where spruce and pine logs cut in April were  being peeled (debarked)  for log homes. The bark included the  rhizome and a thin strip of inner bark. Enclosures were sampled initially (before  treatment) and then every 5 to 7  days for the duration of the experiment (2 to 4 weeks). The adjacent lake was also sampled to allow a comparison between the dynamics of the enclosures and the lake. All samples and in situ measurements were taken at a depth of 1 m.  3.2 EXPERIMENTAL DESIGN OF 1985  LOG STUDY  The experiments of the 1985 field season closely examined the effect of log booms, simulated by log rafts (logs approximately 1.3 diameter)  m in length and 14 cm in  on enclosure water quality, bacterial activity and biomass, and  phytoplanktonic algal biomass. The original eight enclosures of 1984 were used in the log study. The log enclosure studies of 1985  attempted  to qualify the  19  Table 3.1 Summary of the 1984 enclosure experiments.  Duration  Experiment  7 June to 8 July  20 kg bark 5 kg bark  12 July to 26 July  1 kg bark  29 July to 9 August  log raft*  17 July to 7 August  = raft contained 5 pine logs and 3 spruce logs, freshly cut 1 week prior to the start of the experiment ; each log was approximately 1.3 m in length and 14 cm in diameter.  environmental effects of pine and spruce leachates generated from a simulated log storage facility. This was done with a more realistic loading of logs to define a graduated range of effects from severe water quality degradation to no significant change in the enclosure environment. The  effects of pine logs and spruce logs on the enclosure ecosystem were  studied independent of one another, i.e. four enclosures for spruce logs and four enclosures for pine logs. The experimental design of 1985  3.2.1  is outlined in Table  3.2.  ENCLOSURE PREPARATION  The  enclosures were filled with filtered lake water (100  Mm) by a 3 horse  power Honda water pump. The logs used for the study were harvested one week prior to the commencement of the experimental period. It is important to emphasize that the bark and logs used in the 1984  and 1985  experiments were  fresh, i.e. had not been exposed to time and weather conditions. This enabled us  20  to simulate as closely as possible the actual logging and storage operations practised by Houston Forest Products at Morrison Arm where the felled logs are immediately bundled and dumped into the holding bay. All enclosures and the adjacent lake were sampled at 0, 2, 5, 10, 15, 20, and 25 day intervals at a depth of 1 m. The dissolved oxygen measurements were performed approximately every other day. Initial sampling, day 0, was conducted before logs were added to the enclosures. The first set of spruce and pine experiments was in June, where the average enclosure temperature was 9 °C. The experiments were repeated in July to examine the effect of water temperature on enclosure dynamics.  (average enclosure water temperature  was 15.6  °C)  21  Table 3.2 Experimental design of the 1985 enclosure  Species of tree  Experimental interval  Pine  Number of logs/enclosure  June 26/05/85 to 20/06/85  C 1 3 5  July 02/07/85 to 27/07/85  Spruce  C 1 3 5  June 02/06/85 to 27/06/85  C 1 3 5  July 09/07/85 to 03/08/85  C  =  control (no logs added)  study.  C 1 3 5  4. METHODS OF ANALYSIS  4.1 WATER QUALITY  The method of preservation, storage container used, and quantity of water sample preserved are outlined in Table 4.1.  4.1.1  GENERAL WATER QUALITY MEASUREMENTS Dissolved oxygen and  temperature  The D O and temperature were determined in situ with a YSI Model 54 dissolved oxygen/temperature  meter. The oxygen probe was calibrated at the  beginning of each sampling day by wrapping the probe in a wet towel (simulates air saturation), taking the air temperature,  and then correcting for both  temperature  and elevation. pH and  alkalinity  The pH was determined with a portable Fisher pH Meter was determined by titrating a 100 ml water sample with 0.02  119. The alkalinity  N ^ S O ^ to a pH  of 4.5 (Standard Methods, 1985). The alkalinity was calculated with Equation 4.1 :  alkalinity as C a C 0  3  (mg/l) =  A x N x 50,000 ml of sample  22  (4.1)  23  Table 4.1  Process of preservation for analytical procedures for water bacterial, and phytoplankton studies.  Analysis  Volume of sample  T O C , TIC, and VFA L-T  C O D , TKN, TP, and TRC  where : A = N  4.1.2  =  Storage container  quality,  Method of preservation  5 - 7 ml  glass vials  frozen (-10  °C)  250  ml  high density polyethylene sample bottles  frozen (-10  °C)  500  ml  high density polyethylene sample bottles  H S0 2  to  4  pH<2, cold room  ml of standard acid normality  of the standard acid.  ORGANIC POLLUTION INDICATORS Total organic and inorganic carbon  Total carbon (TC) and total inorganic carbon (TIC) Beckman Model 915A Total Carbon Analyzer (detection  were analyzed with a  limit =  0.1  mg/l) by the  combustion infrared method (Standard Methods, 1985). Two standard curves were made, one for Total Carbon (10,  20, 30, 50, and 100 mg carbon/I from 1.0625 g  of crushed and dried KHCgH O /500 ml distilled water) and the other for Total 4  Inorganic  4  Carbon (5, 10, 20, 30, and 50 mg carbon/I from 0.4404 g N a C 0 2  3  and  24  0.3497 g NaHCOg/100 ml distilled water). The water samples were thawed and brought to room temperature. injected into a heated (950  To determine Total Carbon, 50 n\ of sample was  °C), packed ceramic combustion tube in a stream of '  oxygen (Total Carbon Channel). The water sample was vapourized and the carbon oxidized to CC>2 and measured by a nondispersive type of infrared analyzer.  The TIC was measured on the TIC Channel where the furnace was 150  temperature  °C. The volume of water sample injected was 50 n\ and the carrier gas  was oxygen. T O C was found by subtracting the TIC results from the TC data. Chemical oxygen demand  The chemical oxygen demand (COD) was performed according to the dichromate reflux method (detection  limit =  0.01  mg/l) detailed in Standard  Methods (1985), except the sulfuric acid reagent was added directly to the flasks and not through the top of the condenser. Equation 4.2 was used to calculate COD:  C O D (mg/l) =  (blank - sample)  x N  FAS  x 8000  ml of sample where  N  ml dichromate x 0.25  FAS  ml ferrous ammonium sulfate standard blank  blank titre of distilled water, dichromate, sulfuric acid reagent  sample =  sample titre  FAS standard =  consists of distilled water and dichromate  (4.2)  25 Lignins and tannins  Lignins and tannins (L-T) were determined by a spectrophotometric technique where the aromatic hydroxy! groups of these plant constitutents were reacted with tungstophosphoric and molybdophosphoric acids to form a coloured (blue) complex (Standard Methods, 1985). The water samples were thawed, brought to room temperature,  the reagents added to the samples and standards (0.5,  10 mg/l as tannic acid), and left for 1/2  1, 2, 3, 5, and  hour to allow for colour development. The  absorption of the coloured complex was determined with a 1 cm cell on a Pye Unicam SP8-100 Ultraviolet  Spectrophotometer (detection  limit =  0.1  mg/l) at  700  T?m and the concentration determined from the standard curve.  4.1.3  NUTRIENTS AND ORGANIC SUBSTRATES Total  Kjeldhal  nitrogen  Total Kjeldhal nitrogen (TKN) was prepared by rigorous acid digestion and determined spectrophotometrically by formation of the ammonium salicylate complex method as outlined in Technicons' Industrial  Methods 329-74W and 376-75W. The  samples were analyzed with a Technicon Autoanalyzer II (detection at a wavelength of 660  77m.  limit =  0.2  mg/l)  The final TKN concentration was determined from a  standard curve developed for the automated analysis. Total phosphorus  Total phosphorus (TP) was measured spectrophotometrically by the stannous  26  chloride reduction method where ammonium molybdate and stannous chloride reacted with the water sample to form a phosphomolybdenum complex  (Standard  Methods, 1985). The coloured complex was read within 10 minutes of colour development with a 1 cm cell on a Bausch-Lomb Spectronic 88 (detection 0.1  mg/l) at 690  p .  A standard curve was generated from 0.5,  limit  =  1, 2, 3, and 5  mg/l of Fisher Phosphate Standard Solution (1 mg phosphorus/1 ml). Total reducing carbohydrates  A colourimetric method was used for quantitative estimation of carbohydrates (Keleti and Lederer, 1974). The standards consisted of 0.4,  1.2,  2.4,  and 3.4  mg/l  of dextrose. Absorption of the coloured complex was made on a Bausch-Lomb Spectronic 88 at 690 Volatile  rjm  using a 1 cm cell (detection  limit =  0.1  mg/l).  fatty acids  Preliminary measurements of the volatile fatty acids (VFA) acetic, propionic, and butryic were measured on a Hewlett Packard 5750 Gas Chromatograph. However,  the concentration of VFA's was below the detection  chromatographic technique, ^  limit of the gas  2 mg/l. Therefore, an alternate method for  low VFA concentrations (Barcelona et al. 1980) Mg/l). The water samples were derivatized 5880A Series Gas Chromatograph.  was used (detection  determining  limit =  0.1  and measured with a Hewlett Packard  27  4.2  4.2.1  BACTERIAL POPULATION  BIOMASS AND NUMBERS  The total number of bacteria were counted by the acridine orange direct count (AODC) epifluorescent technique as modified by Daley and Hobbie (1975). Nuclepore filters were stained with an irgalan black solution (2 g/l in 2% acetic acid). On sampling days, the prestained filters were rinsed with wash water (lake water filtered with Type HA 0.2  um Millipore filters). The stained filters were placed  on a small filter assembly, 1 ml of sample water was added and 4 ml of wash water. The final volume was filtered and washed down with an additional 5 ml of wash water. The filter was placed in a gas-tight petri dish and allowed to air dry for 8 hours. The petri dishes were then labelled with the volume of water sample filtered, and taken to the National Water  Research Institute in North Vancouver for  counting. Counts were made with a Leitz Ortholux microscope (with an internal magnification of 1.25X) equipped with 12.5X oculars, an NPL 100X oil immersion objective, and a lamp and filter system detailed in Daley and Hobbie (1975). The air dried filters were placed on a filter assembly, stained for 5 minutes with 0.2 um filtered acridine orange, filtered, and washed with 5 ml of 0.2  um (Type HA  Millipore) wash water. The filter was placed on a glass slide with a drop of Cargille A Immersion  Oil, a cover slide added, and the bacteria counted at 1562X  magnification. Random fields were counted until over 400  bacteria were enumerated.  These counts were converted to counts/ml (Appendix A). Counts were performed on the volume of wash water and acridine orange used for sample preparation, and subtracted from the sample counts.  28  Bacterial biomass was determined by using a micrometer and measuring the length and width of rods and the diameter of the spheres. The measurements were converted to cell volumes and ultimately to grams of carbon/ml. The calculations for this procedure are detailed in Appendix A. The biovolume for the filamentous bacteria present during the latter stages of the study were found by counting the number of times a filament determination  crossed the counting grid (Brock, 1978). The  of cell biovolume is listed in Appendix A.  4.2.2 HETEROTROPHY  Heterotrophy, isotope, i.e.  14  a measure of microbial activity, or the uptake of radioactive  C-glucose for the bark study and  14  C-acetate  for the pine and  spruce log enclosure experiments, by microorganisms, was an in situ  experiment  performed in each enclosure and the lake. Isotope preparation involved the dilution (with 0.2 um filtered lake water) of 4 microCurrie's (MQ) activity 14  =  C-acetate  of ^ C-gIucose (specific 4  346 uG/Mmole) to a volume of 30 ml in 1984 and 4 uCi of (specific activity  =  57.6 uCi/umole) to a final volume of 30 ml in  1985. Four 10 ml syringes were rinsed with the sample water and 9 ml of sample drawn into each syringe. The control was injected with 0.2 ml of concentrated formaldehyde. After a 5 minute period to kill the control, 1 ml of isotope solution (1 ml =  0.0537 Mg glucose or 1 ml =  0.06985 Mg acetate) was injected into each  syringe. The 4 syringes were then suspended in an enclosure at a depth of 1 m and incubated for 3 hours. This procedure was repeated for each enclosure and the lake. At the end of the incubation period the samples were filtered (0.2 um, mm, cellulose nitrate membrane filters), and the filters transferred to scintillation vials, where they were inactivated with 10 ml of PCS scintillation solution  25  29  (Amersham Searie).  The scintillation vials were transported to the University of British Columbia (U.B.C.) and counted on a Nuclear Chicago Isocap 300 scintillation counter operated in an external standard mode to correct for quenching. The raw count data was corrected to yield microbial uptake rates in ug of glucose or acetate/l/h. An example calculation for determining the uptake rates is presented in Appendix A.  4.2.3  HETEROTROPHIC UPTAKE KINETICS Bark leachate inhibition experiments  The inhibition of white spruce bark leachate to microbial activity was determined in a laboratory experiment with a bark concentration equivalent to 10 kg bark/3700 I water. After a contact period of 12 days, the bark leachate was filtered through CFC filters and then refiltered with 0.2 jum, cellulose nitrate membrane filters.  The bacterial population used for the inhibition test consisted of 50 ml of settled primary sewage (ensured a population of bacteria) and 400 ml of pond water (dilution of sewage while maintaining an isotonic solution). The radiolabelled glucose (specific activity  =  333 uCi/umole) was prepared to a final volume of 30  ml with sterile, distilled water. Table 4.2 lists the volumes of isotope, sample (sewage  +  pond water), formalin, sterile distilled water, and leachate added to each  25 ml reaction flask. There were 2 leachate concentrations used for the experiments ; 2 ml and 5 ml additions.  30  Table 4.2 Volumes of solutions adde d to the bark leachate reaction flasks in microbial inhibition studies.  Solutions  Volumes added to flasks (ml)  Blank-C  Control  Blank-L  1. Sample  4.0  4.0  4.0  4.0  2. Formalin  0.2  0  0.2  0  3. Make-up water^  5.0  5.0  0  (3.0)  0  0  5.0  (2.0)  1.0  1.0  4. Leachate 5. Isotope  Leachate  +  0  (3.0)  5.0  (2.0) 1.0  1.0  (1) = sterile distilled water * = numbers in brackets represent volume differences for the 2 ml addition of bark leachate  After adding the isotope, the incubation flasks were fitted with tight sealing rubber caps with small plastic cups containing folded GFC 2.4 cm glass fibre filters saturated with 0.1  ml of 1 M hyamine hydroxide to absorb  14  CC>2. At the end of  the 2 hour incubation period, the samples were killed with 0.2 ml 5 N F^SO^. The flasks were swirled and allowed to sit for 20 minutes to collect the  ^^CO^  To determine particulate uptake, the water samples were filtered through 0.2  /im  cellulose nitrate filters, the filters transferred to scintillation vials, and dissolved in 10 ml PCS scintillation solution. The GFC filters were removed from the plastic cups, placed in scintillation vials and 10 ml of PCS scintillation fluid added ; this provided information on the amount of CC>2 respired by the microorganisms. Samples were counted as outlined in Section 4.2.2.  31 Bacterial uptake studies with log leachates  A comparison of bacterial dynamics involved in the utilization  of organic  solutes was done on samples from the lake, control, spruce (5 logs), and pine (5 logs) enclosures taken at the beginning of the experiment The kinetic uptake  (day 0) and at day 10.  procedure used was adopted from Hall ef al. (1972) and the  organic solute used was  14  C-acetate  (specific activity  =  57.6  The isotope was diluted to 0.1333 uCi/ml with 0.2  uCi/umole).  um filtered  Uptake was measured at 5 duplicate solute concentrations, 20, 40, ug/l of unlabelled solute. Table 4.3  60,  lake water. 80, and  100  lists the volumes of isotope, water sample, and  unlabelled acetate added to the 25 ml reaction flasks. A blank for each solute concentration was fixed with 5 N H2SO4 prior to isotope addition. After adding the isotope, 0.1365 ug  14  C - A c . / m l , the incubation flasks were  prepared in the same method outline above in Section The final solute concentration (labelled 85.1,  and 103.3  +  unlabelled acetate) of the flasks was 30.6,  48.8,  67.0,  ug Ac./I. At the end of the 3 hour incubation period, the samples  were killed with 0.2  ml 5 N H S 0 , 2  4  prepared and preserved as outlined in the  previous section.  4.3  4.3.1  PHYTOPLANKTONIC ALGAL POPULATION  BIOMASS AND ENUMERATION  The algal biomass indicator used was chlorophyll _a_ (chl _a). Chi _a_ was determined  by filtering  known volumes of water samples through 47 mm  GFC filters. Approximately  1 ml of MgCO..  diameter  (slurry) was added to the remaining  few  32  Table 4.3 Volumes of solutions added to the log leachate reaction flasks in microbial activity studies.  Solutions  Volumes added (ml)  to flasks  1. Sample  9.0  9.0  9.0  9.0  9.0  2. Unlabelled acetate  0.2  0.4  0.6  0.8  1.0  3. Make-up water  0.8  0.6  0.4  0.2  0  4. Isotope  1.0  1.0  1.0  1.0  1.0  ml of each filtrate. The filters (replicates) were then placed in petri dishes and frozen. Upon return to the Environmental Engineering Laboratory at U.B.C., the chl _a samples from the bark studies were extracted with 11 ml of 90% acetone in graduated centrifuge tubes over a 24 hour period in a dark, cold room (4  °C ;  Strickland and Parsons, 1972). The samples (including replicates) were centrifuged (1500  rpm), the supernatant drawn off, the volume recorded, and the fluorescence  measured on a Turner Designs Fluorometer. Equation 4.3 describes the chl _a_ standard curve ; Equation 4.4 was used for volume corrections :  *  =  Fo (0.968 - 0.00035Fo)  (4.3)  33  chl _a_ (ug/I) =  $'  x  O.Q1(litres)  (4.4)  +  volume of water filtered  where  : Fo = $  = =  *  =  fluorometer  reading  uncorrected chl _a_ concentration averaged uncorrected chl _a_ cone. volume of extraction liquid  To ensure the complete extraction of chl _a_ from the samples, extraction solvents and technique were changed for the log study samples. The frozen filters were placed in graduated, screw top centrifuge tubes and 10 ml of 2:1 (volume : volume) chloroform : methanol extractant  added (Wood, 1985). The extraction  process was allowed to proceed for 5 hours in a dark, cold room (4 tubes were centrifuged (1500  °C), the  rpm), the supernatant removed, volume recorded, and  fluorescence promptly measured on the Turner Designs Fluorometer. A new standard curve was generated for the chloroform : methanol extraction solvent, and the final chl _a_ concentrations were determined from the values obtained from the curve and Equation 4.4. For phytoplanktonic algal identification and enumeration a 150 ml sample was collected from the lake and the enclosures, preserved with 1 ml of Lugol solution, and returned to U.B.C. A volume of the sample water was placed in a 25 ml settling chamber, covered with a microscope slide, and left undisturbed for 24 hours (Standard Methods, 1985). The preserved phytoplankton were enumerated with a Nikon Phase/Contrast Inverted Microscope.  34  4.3.2  PHYTOPLANKTONIC PRIMARY PRODUCTION  Primary production was determind by the in situ uptake of radioactive 14  C - H C O - j " isotope (Strickland and Parsons, 1972). Preparation involved the dilution  of 40 MCI" Four 300  of  1 4  C - H C 0 " with filtered 3  0.2  /zm lake water to a volume of 22 ml.  ml BOD bottles were rinsed and filled with sample water. One BOD  bottle was injected with 1 ml of concentrated formaldehyde (control)  and a second  was injected with 5 ml of D C M U , a photosynthetic inhibitor (Legendre et al. 1983). After a 5 minute period to kill the control and inactivate the DCMU sample, 1 ml of isotope solution (1 ml =  1.818  MCI)  was injected into each bottle.  Four bottles  were suspended in each enclosure and the lake for a 6 hour incubation period. At the end of the incubation period, the BOD bottles were filtered (0.45  Mm  membrane filters), transferred to scintillation vials, acidified to remove any unincorporated  1 4  C-HC0 ", 3  and then inactivated with 10 ml of PCS scintillation  solution. The scintillation vials were counted by the same method outlined in Section 4.2.2. The data were corrected and the uptake rates calculated in rjg C/l/hr (Strickland and Parsons, 1972).  4.4 COMPARSION BETWEEN THE BARK AND LOG STUDIES  Experiments with similar bark weights were used to quantitatively  compare the  bark and log study results. The bark for the 1984  study was dried in an oven at  110  of bark added to the enclosures  °C for 24 hours to determine the dry weight  (Table 6.1).  For the 1985  hours at 110  study, the logs were peeled, the bark oven dried for 24  °C, and the results reported in Table 6.1.  35  4.5 STATISTICAL ANALYSIS  The test results of the bark and log studies were statistically analyzed with U.B.C. ANOVAR, a statistics package providing analysis of variance and covariance adapted by M. Creig and D. Osterlin from B.Y.U. Documentation (1977). A two-way ANOVA (a  =  0.05)  was performed on the 1984  test results to determine  the  significance between the 20 kg bark treatment and control. The two-way ANOVA was used on the 1985  log study results to test significance between  treatments  (number of logs) and control, between species (pine and spruce), over time (25 days), and at different water temperatures statistical analysis for 1985  (i.e.  month to month variations). The  included the significance of the above listed interactions  as well as relationships between any combination of the above, i.e. species x time x number of logs, species x time x month, etc.  5. RESULTS  5.1  ENCLOSURE EXPERIMENTS WITH BARK - 1984  The 20 kg bark experiment meet the criteria for statistical analysis, i.e. homogeneity of variance. However, the 5 kg, 1 kg, and 8 logs treatments violated Bartlett's homogeneity of variance assumption, therefore, statistical interpretation of the data was limited. For example, there was a significant decrease in dissolved oxygen in the 20 kg bark treatment (Figure 5.1).  Although a similar decrease in  oxygen in the 5 kg and 1 kg bark experiments could not be statistically validated, there was a trend similar to that observed in the heavy bark enclosure. Considering that the 20 kg bark enclosure results could be statistically tested and the difficultly in graphing all the bark treatments  because of differences in (i)  sampling intervals and experimental duration, and (ii) temporal changes, the results of the heavy bark treatment and its control were graphed, and the data from the remaining treatments were tabulated.  5.1.1  COMPARISON OF THE LAKE AND CONTROL DATA  A statistical comparison (ANOVA a  =  0.05)  was made between the lake and  control enclosures to investigate impoundment effects on water quality and biological measurements. There was no significant difference between control and lake for alkalinity, dissolved oxygen, lignins and tannins, primary production, and chlorophyll ja_ (Tables 5.1,  5.2, and 5.3).  However, pH and heterotrophy values were statistically  different between the control and the lake (Tables 5.1  36  and 5.3)  ; this may be  37  attributed  to the contained or static environment of the sealed enclosures. Since the  changes in pH and heterotrophy were small, the control and lake values were considered similar.  5.1.2  WATER QUALITY General water quality  (i) Dissolved oxygen  Figure 5.1  measurements  (DO)  demonstrates the severe and significant DO demand created by 20  kg of bark. The sharp decrease in DO concentration observed in the 20 kg bark enclosure was also observed in the lower bark treatments,  however, the severity of  the trend was proportional to the bark concentration. For example, after 5 days, the DO in the 20 kg bark treatment had dropped to less than 1 mg/l while in the 5 kg bark enclosure it took 14 days to reach a similar DO concentration (Table 5.1).  For the 1 kg bark enclosure, the D O was still at 6 mg/l after  (Table 5.1).  The enclosure experiment which contained 8 logs showed an oxygen  consumption approximately equal to that of the 5 kg enclosure (Table  (ii)  pH and  11-days  5.1).  alkalinity «  There was a steady and significant decrease in pH in the 20 kg bark enclosure, compared to the control, where the pH values dropped from 7.30 5.55  in 31 days (Figure 5.2).  to  There was a pH decrease in the 5 kg, 1 kg, and 8  logs experiments, however, the changes were small (<  1 pH unit, see Table  5.1).  T a b l e 5.1  Changes  i n d i s s o l v e d oxygen,  pH, and a l k a l i n i t y  over time  i n the 1984  bark e n c l o s u r e  studies  done at  Babine Lake.  DO  (mg/l)  pH  L  C  7 .02  3 8 .0  3 7 .5  3 8 .5  6 .91  3 5 .0  3 8 .0  3 7 ..5  7 .61  3 6 .0  3 9 .5  3 8 ..5  7 .01  7 . 13  37 . 5  3 8 .0  39. O  7 . 13  7 .26  3 7 .0  3 9 .0  3 9 .,5  Days  T  5 kg bark  0 7 14  10..6  10..4  10,.2  7.13  7 . 17  3..3  9 .9  10. 1  6.55  7 .46  0..4  9 .4  10..9  6.76  7 .74  0  9 .4  9.. 8  9. 8  7 . 30 6.75  1 kg bark  8  logs  T  =  C  = control  L  =  C  L  L  4 11  7..9  8 .6  9..0  6. 0  8,.9  8. 6  6.93  7..29  7,. 12  3 7 .5  38 . 5  3 9 .,5  0  9..4  9..7  9. 5  7 . 10  7 .31  7 .00  3 7 .0  3 9 .0  3 9 ..0  7 14 21  4 .7  9 .8  10. 1  6.43  7 . 20  6 .96  3 6 .9  3 7 .5  3 8 ..0  0. 8  9 .6  9. 2  6.26  7 . 29  7.. 14  3 5 .0  3 8 .0  3 7 ..5  1 .1  8 . .6  8 .7  6 . 27  7 . 13  7 .0 4 .  3 5 .O  3 8 .O  3 8 .,5  treatment 1 ake  T  (mg/l)  T  Treatment  C  Alkalinity  enclosure  (no bark)  39 12  v  I  I  1  1  1  1  1  1  0  5  10  15  20  25  30  35  '  TIME (days) Figure 5.1  Temporal changes in dissolved oxygen in the 20 kg bark enclosure at Babine Lake ; control = no bark.  8  Figure 5.2 Temporal changes in pH in the 20 kg bark enclosure experiment Babine Lake, 1984 ; control = no bark.  at  40  —  60  Control O  O  20 kg Bark I  10-  0  5  10  15  TIME  20  25  30  35  (days)  Figure 5.3 Temporal changes in alkalinity in the 20 kg bark enclosure experiment at Babine Lake, 1984 ; control = no bark. ^  IOOOJ  2  o  0  5  10  15  TIME  20  25  30  35  (days)  Figure 5.4 Temporal changes in total organic carbon in the 20 kg bark enclosure at Babine Lake, 1984 ; control = no bark.  41  In the 20 kg bark enclosure experiment, the alkalinity dropped from 44 mg C a C C y i to 20 mg C a C C y i by the 10th day of the experiment  (Figure  5.3).  However, the alkalinity gradually increased to 31 mg C a C O y i by the end of the experiment. There were no significant decreases in alkalinity values exhibited by the other treatment enclosures (Table  5.1). Organic pollution indicators  (i) Total organic and inorganic carbon (TOC and TIC)  The T O C concentration of the 20 kg bark enclosure increased from 7.8 to 390 mg/l during the 31 day experimental  period (Figure 5.4).  The increase in  organic enrichment with the addition of 20 kg bark, was exhibited in the bark treatments  in lesser degrees of severity (Table 5.2).  mg/l  other  The high levels of TOC  indicated that a substantial amount of organic material was being leached from the bark. For the  1985  log study, the organic matter was characterized to  determine  the general chemical composition of the leachate. The increase in TIC concentration in the 20 kg bark enclosure was significant compared to the control enclosure (Figure 5.5).  However, there was no significant  change in inorganic carbon levels in the 5 kg, 1 kg, and 8 logs enclosure (Table 5.2).  The increase in inorganic carbon in the 20 kg enclosure from 6.7 mg/l to  28.6  mg/l in 5 days should have been accompanied by an increase in alkalinity  since the alkalinity in Babine Lake is mainly attributed  to the bicarbonate and  carbonate anions. However, the alkalinity decreased within the experimental time span. The discrepancy between Section 6.1.2.  inorganic carbon measurements is discussed in  T a b l e 5.2  Temporal  changes  In the  c o n c e n t r a t i o n of o r g a n i c  and Inorganic  e n c l o s u r e experiments  TOC  Treatment  5 kg bark  1 kg bark  8 logs  T  =  C  = control  L  =  Days  T  TIC (mg/1)  C  T  the 1984 bark  C  L-T  T  (mg/l)  C  L  0.5  0  9.0  8.1  5.3  6.7  0.4  0.4  7 14  82.0  11.5  5.5  4.7  22.5  0.6  0.4  114.0  8.5  6.1  6.0  35.0  0.5  0.5  0  9.0  7.7  4.2  6.5  0.5  0.4  0.5  4 11  10.5  8.5  4.0  6.5  1.3  0.5  0.4  10.9  8.8  6.5  6.0  11.2  0.5  0.4  0  8.0  6.1  4.0  9.2  0.4  0.5  0.4  7 14 21  16.0  9.8  4.2  4.2  2.9  0.5  0.4  3.8  0.4  0.4  4.8  0.4  0.9  treatment lake  (mg/l)  carbon compounds i n  at Babine Lake.  enclosure  (no b a r k )  43  40  E o  30-  CD CC <  2 0 kg Bark  o  y  /  20  z < o rr  5  Control  «H  < Io 0  10  15  20  25  30  35  TIME (days) Figure 5.5 Temporal changes in the total inorganic carbon levels in the 20 kg bark enclosure at Babine Lake, 1984 ; control = no bark.  1000-  CD  E  20 kg BarkT  100-  CO  10-  Control  i CO  —O—-  0.1-  0  10  15  TIME  20  25  30  35  (days)  Figure 5.6 Temporal changes in lignins and tannins concentration for the 20 kg bark enclosure at Babine Lake, 1984 ; control = no bark.  44  (ii) Lignins and tannins (L-T)  L-T were leached rapidly from the 20 kg bark enclosure i.e. days (Figure 5.6). =  0.05)  338 mg/l in 10  This increase in the concentration of L-T was very significant (a  compared to the control enclosure and paralleled the increase in the TOC  levels. In the 5 kg bark enclosure, the L-T concentration after a two week leaching period was 10 times lower compared to the 20 kg experiment, enclosure only contained 11.2  mg/l after 11 days (Table 5.2).  had a L-T concentration of 4.8 mg/l (Table 5.2)  while the 1 kg  The 8 log experiment  after three weeks, significantly less  than L-T levels in the 20 kg and 5 kg bark experiments.  5.1.3  BACTERIAL POPULATION  Heterotrophy  The addition of 20 kg bark and 8 logs to the experimental enclosures caused an apparent enhancement of heterotrophy. There was a significant increase (a  =  0.05)  in glucose uptake rates in the 20 kg bark' study after materials had  leached into the water for 7 days or longer (Figure 5.7) of glucose increased from 0.1 7.3  ; for example, the  uptake  jug Glu./l/h at the beginning of the experiment  to  Mg Glu./l/h in 24 days. There was an increase in microbial activity in the 8 log enclosure over a  two week period, i.e. 0.2  Mg Glu./l/h to 16.4  uptake of glucose dropped to 1.0  Mg Glu./l/h (Table 5.3).  However,  the  Mg Glu./l/h during the third week. The observed  trends in microbial activity are discussed in Section 6.1.3.  T a b l e 5.3  Temporal  changes In heterotrophy  and p h y t o p l a n k t o n i c biomass experiments  Heterotrophy  (,,g  and p r o d u c t i v i t y  i n the 1984  bark e n c l o s u r e  at Babine Lake.  Primary p r o d u c t i o n  Glu./1/h)  C h l o r o p h y l l a (j>g/1)  ( g c/l/h) n  Treatment  Days  5 kg bark  0 7 14  0.5  0.7  0.2  0.9  1 .2  0.3  0. 1  0.2  0. 1  1 .5  0.5  0.6  0.2  1 .2  0.7  0  0.2  0. 1  1 .6  0.7  0.6  0.3  1 .4  1. 1  0  0.3  0. 1  0 4 11  0.4  0.3  0.2  0.8  0.2  0.2  0. 1  0. 1  0. 1  2.5  1 .0  0.7  0.6  0  0.4  0.2  0. 1  0. 1  0.9  0.9  0  0. 1  0. 1  0 7 14 21  0.2  1 kg bark  8  T =  logs  8.5 16.4  1 .0  treatment  C = control L = lake  enclosure  (no l o g s )  0.2  0. 1  0.7  0.7  0.6  0. 1  0. 1  O. 1  0.2  0.6  0  0. 1  0. 1  0  O. 1  0. 1  0.3  0.2  1. 1  1. 1  0. 1  0.4  1 .0  1.5  1 .4  0.3  0.4  0.7  0.2  46  10  ^  8-  20 kg Bark  UJ  <  z> LU V>  O o  Control  —I  o O  0-  10  0  15  TIME  20  25  30  35  (days)  Figure 5.7 Temporal changes in heterotrophic uptake of labelled glucose in the 20 kg bark experiment at Babine Lake, 1984 ; control = no bark.  0.20-  ^  =3. 0.15  CD  > X  0.10-  o  §  0  \  05  \ I 20 kg Bark 0.00-  0  5  10  15  TIME  20  25  30  35  (days)  Figure 5.8 Temporal changes in chlorophyll _a_ concentration in the 20 kg bark enclosure at Babine Lake, 1984 ; control = no bark.  47 Bark leachate inhibition  experiments  There was a significant decrease in the net uptake of labelled glucose in the white spruce bark inhibition experiments. The control (no leachate) uptake rate of 14.1  Mg Glu./l/h (n  =  had an average  3), while the uptake rates of both the 2 ml  and 5 ml leachate additions had reduced the heterotrophic activity to zero. Thus, there was complete inhibition of natural sewage microbial populations after a 2 hour incubation period with bark leachates.  5.1.4  PHYTOPLANKTONIC ALGAL POPULATION Phytoplanktonic biomass  There was a rapid and significant decrease in chlorophyll _a_ in the 20 kg bark enclosure from 0.2  Mg/l to 0 in 10 days (Figure 5.8).  This decreasing trend  was also apparent in the 5 kg and 1 kg bark and 8 logs enclosures where chlorophyll _a_ was below detection level by the end of the experimental (Table  period  5.3). Phytoplanktonic primary  production  The changes in phytoplanktonic primary production in the 5 kg bark, 1 kg bark, and 8 logs enclosure experiments are presented in Table 5.3. bark enclosure, an initial productivity rate of 0.8 the fifth day of the experiment  In the 20 kg  Tjg C/l/h was reduced to zero by  ; this decrease in productivity paralleled the drop in  the phytoplanktonic biomass. The 8 log and 5 kg bark experiments also showed some inhibition of phytoplanktonic algal production with reductions of 50% - 70%  48  after 7 days. No changes in primary production were observed in the 1 kg treatment.  5.2  5.2.1  ENCLOSURE EXPERIMENTS WITH LOGS -  1985  COMPARISON OF THE LAKE AND CONTROL DATA  A statistical comparison (ANOVA a  — 0.05)  was made between  the control  enclosures and Babine Lake for reasons outlined in Section 5.1.1. There was a significant increase in organic carbon and chemical oxygen demand in the lake during the first 5 days of the experiments with pine logs in June when compared to the control enclosure. These increases were possibly caused by the resuspension of organic material from the lake sediment, the result of a severe wind storm that occurred during the first two  days of the  experiment.  Generally, there was a significant increase over the June and July 25 day test period in heterotrophy,  bacterial numbers and biomass, and chl _a_ in the control  enclosures (no logs) during studies with both tree species compared to the lake values (Tables 5.4 and 5.5)  ; this was probably caused by the sealed (static)  enclosure design. The bacterial biomass in the June control enclosures and lake fluctuated  over the test interval, however,  by the end of the experiments  spruce and pine) biomass in the control was greater values.  (both  (3 to 4 times) than the lake  Table  5.4  A comparison  between the pine  log control  water  e n c l o s u r e and Babine  q u a l i t y and b a c t e r i a l  T1me  0 Test int.  June  Measurement  COD  5 C  (mg/l)  TOC (mg/l) Bact. no. (cts/ml x 10 ) Bact. bio.  L  Lake of s i g n i f i c a n t  L  several  (days)  10 C  changes In  measurements.  15  C  L  20  C  L  C  25 L  C  L  18.15  15.72  19.00  36.00  19.OO  30.00  15.72  11.79  13.76  17.10  24.70  27.50  6.5 4.1  4.8 3.7  6.0 5.7  13.3 4.4  6.5 -  6.5 3.9  10.5 7.2  9.5 4.3  7.5 10.3  7.0 4.0  10.0 18.2  6.5 9.5  22.2  10.1  -  -  -  -  -  43.0  11.2  1.4  1.2  1.7  1.1  1.4  1.4  1.8  1.4  1.3  1.1  1.4  1.5  9.9  10.9  8.0  6.0  9.5  6.9  11.0  4.8  6.5  4.8  7.2  6.5  72.8  . 72.0  -  -  105.0  32.0  -  24.2  18.3  s  („g Chl.  duly  C/l) a („g/l)  Bact. no. (cts/ml x 10 ) Bact. biO.  -  -  -  5  Test  („g C/l) Heter. („g Ac./l/h)  0.3  0.1  0.2  0  1.0  0.4  0.1  0.1  0.3  0.3  0.7  0.2  Chl.  1.4  1.2  1.6  1.0  1.2  0.9  1.0  1.0  1.5  1.1  1.4  1.1  a ( g/l) v  1nt. = t e s t  C  = control  L  =  lake  Interval  enclosure  (no l o g s )  Bact.  no.  Bact.  bio.  = bacterial = bacterial  Heter.  =  Chl  = chlorophyll  a  numbers biomass  heterotrophic activity a  Table  5.5  A comparison  of s i g n i f i c a n t  changes  spruce log  In heterotrophy  control  and b a c t e r i a l  e n c l o s u r e and  Babine Lake,  Time  0 Test int.  Measurement  June  Bact.  C  no.  (cts/ml Bact.  x  C L  bio.  C  L  20  C  47.8  L  5.1  C  9.6  25 L  6.2  C  L  7.5  6.4  6.6  6.6  8.3  6.0  6.8  4.3  51.4  42.0  -  -  121.4  89.6  -  88.7  27.8  1.0  0.1  0.1  0.2  0.3  0  0.4  0  0.7  0.3  12.0  7.9  13.6  9.0  11.7  9.1  11.7  10.7  10.4  11.9  10.5  12.5  0.1  0  0.3  0.2  0.3  0.2  0.4  0.3  0.3  0.3  0.3  0.1  Ac./l/h)  Bact. no. (cts/ml x  Int. = test  = control = lake  15  5  10 ) 5  Heter. („g Ac./l/h)  Test  L  1985.  (days)  10 C  the  10 )  Heter.  July  L  and biomass between  c/i)  („g („g  5  numbers  Interval  enclosure  (no logs)  B a c t . no. = b a c t e r i a l numbers Bact. b i o . = b a c t e r i a l biomass Heter. = heterotrophic a c t i v i t y  51  5.2.2  WATER QUALITY  There was no significant change (a experimental  =  0.05)  in any of the spruce or pine  enclosures in alkalinity, pH, total Kjeldhal nitrogen, total inorganic  carbon, or phytoplanktonic algal numbers compared to the control enclosures. Total phosphorus, volatile fatty acids, and carbohydrate values were below the  detection  limits of the analytical methods. A summary of the means and standard deviations for dissolved oxygen, temperature,  chemical oxygen demand, total organic carbon,  lignins and tannins, and chlorophyll _a_ are listed in Table General water quality  (i) Dissolved oxygen  5.6.  measurements  (DO)  There was a significant decrease in June D O concentration after 20 days for the 5 and 3 log treatments  for both tree species (Table 5.7).  From Table 5.6,  average June D O concentrations of spruce 5 and 3 log treatments  the  were higher  compared to pine (surface areas of the pine and spruce logs were the same, i.e. uniform length and diameter).  For example, in the 5 log treatment, the average  oxygen concentrations were 9.7 mg/l and 9.0 mg/l for the spruce and pine logs, respectively. Therefore, pine may exert a slightly higher oxygen demand than spruce. In July, a significant decrease in the DO levels was apparent after 5 days for both the 5 and 3 log experiments for both tree species (Figure 5.9). example, the D O in pine 5 and 3 log treatments  For  decreased from 10.3 to 2.5 mg'/l  and 10.3 to 3.0 mg/l, respectively, over the 25 day experimental  period. The  oxygen demand decreased with a reduction in the number of logs. Although a similar DO deficit trend was exhibited by the spruce study in July, pine exerted a significantly greater oxygen demand at all 3 log loadings. The D O concentrations for  Table  5.6 A summary  of means and  standard d e v i a t i o n s of general water q u a l i t y measurements f o r spruce l o g study at Babine Lake.  the 1985 p i n e  and  Parameter  DO (mg/l) Tree  Test  No.  sp.  Int.  of logs  Pine  June  C 1 3  July  Spruce  June  10..9 10..5 9 .5 .  0.. 6 1 .0 .  9..9  1 .7  16 . .58  4 .85  7 .9 .  1 .8 .  1 .6  0..8  9,,9 9..5  1 .6  .07 . 27. 34..54  7 .72 1 1.35  11 . .0 13.. 1  1 ,9 . 3..4  5  9.,0  1 .. 1  9..5  1 .5  40.,63  17 . 18  16.. 1  5.7  C 1  9..4 8 .9  0,,4 0..9  14 , .9 14. 9  1 .9 1 .9  7 .00 10 .79  10.. 1 12..2  3 5  6 ,, 7 6 ., 1  2 .0 2..3  14..9 14..9  1 .9 1 .9  21 . .97 27 ,,60 39..30 41 . 17  25 .50 19 .87  18 . 1 13 .8  C 1  sp. = tree  Av.  S.D.  TOC (mg/l)  S.D.  3 5 Tree  COD (mg/l)  Av.  5 July  CO  S.D.  C 1 3  Av.  Temp.  Av.  S.D.  L-T (mg/l) Av.  Chl a  („g/l)  S.D.  Av.  S.D.  0. 1  1 .5 . 1 ,8  0.,9  0. 1 0.4  1 .5 ,  0. 2 0. 1 0. 2  1 ,0 ,  0.4  1 .5 .  0. 3  2 .6 . 4 .7 .  0.,4 0,,6  1 .4 . 1 .4 ,  0..2 0..3  4 .7 . 5 .. 1  1 ,0 , 0,.9  0 0. 1 0.4 0.3  2.. 3 1 .8  1 .0 . 0..6  1 .9 ,  0,. 5 0..6 0,.9  0. 1  1 .4  0. 1 0.2  1 .4 1. 5  0, 2 0..2 0..2  1 ,2  0.5  1 .7  0..4  0. 3 0.,4  1 1.0. 10..5 9 .9 9 .7  0 .4 0.. 7 0 .7  9 .0  2 .2  23..32  2 . 28  9..0 8 .6  2 .3 2 .2  22 ..27 35 .86  4 . 24 12 .99  10 .5 9 .4 12 . 2  0..9  8 .6  2 .2  35..01  1 1.76  12 .9  9 .4 9..0 7 .8 . 6 .8  0 .4 0..6  15 .7 15 .7  1. 1 1. 1  19..88 28 .86  0 8 .54  7.. 1 4 .0  1 .8 , 1 ., 1  0,.4 O,. 5  0 0  0 .8 0 .9  0..2 0. 2  1 .3 2 .2  15 .7 25 .7  1. 1 1. 1  31 . .69 33 . 33  16 .01 18 .80  7 .6 9. 3  2,.8 2,.7  0,.9 1,.2  0.3 0.5  1 .0 1 .0  0. 3 0..2  species  Test i n t . = test interval C = control enclosure (no l o g s )  Av.  =  S.D. =  average standard d e v i a t i o n  0 .5 0 .6 2 .8  T a b l e 5.7  Effects  of p i n e and  spruce logs  on d i s s o l v e d oxygen  i n the  1985  June l o g e n c l o s u r e  experiments.  Time (days)  Tree sp.  No. of  0  2  5  10  15  18  20  22  25  logs Dissolved  Pine  Spruce  Oxygen (mg/l)  C 1  11.6 12.4  11.5 11.4  11.1 10.8  11.4 10.2  10.7 10.1  10.9 10.0  10.4 9.5  10.8 9.8  9.8 9.6  3 5  11.1 11.5  10.0 9.7  10.0 9.7  9.3 8.4  8.8 8.2  8.7 8.1  8.6 8.0  9.2 8.6  9.5 9.1  C  10.8  11.3  11.4  11.3  11.4  10.4  11.4  11.0  10.6  1 3  10.6 11.1  11.6 10.9  11.2 10.7  10.8 9.9  11.2 10.1  10.0 9.6  10.5 9.8  10.0 9.6  9.6 9.2  5  11.2  10.8  10.6  9.4  9.8  9.2  9.6  9.4  9.0  Tree sp. = t r e e s p e c i e s C = c o n t r o l e n c l o s u r e (no logs)  54  0  5  10  15  20  25  30  TIME (days) Figure 5.9  Effects of spruce and pine logs on dissolved oxygen in enclosures for the 1985 July log study period ; number of logs in treatment are given in parentheses.  55  the pine studies were consistently lower than the spruce values for the same time span and study period. Organic pollution  indicators  (i) Total organic carbon (TOC)  The general trend exhibited  by pine and spruce was an increase in TOC  values with an increase in the number of logs placed in the enclosures (Table  5.8).  There was a significant increase in T O C concentrations leached from the 3 and 5 log enclosures for pine and spruce compared to the control enclosure. There was no difference  between  water temperature  the pine June and July TOC values which indicated that  did not influence the amount of organic matter leached from  pine logs. However, there was a significant decrease in the amount of TOC leached from all spruce log experiments in July than June. For example, the mean TOC concentration for 5 spruce logs over the June 25 day experimental  period was  mg/l compared to 9.3 mg/l for the same time span in July (Table  5.6).  (ii)  Chemical oxygen demand  The general trends exhibited  12.9  (COD)  by both tree species was a significant increase  in C O D values, over the 25 day test interval, for all 3 log loadings compared to the control results (Figure 5.10). For example, the C O D values for the June 5 pine logs experiment  increased from 19.6  mg/l to 70.7  mg/l between  Days 0 and 25.  The C O D concentrations for July displayed a similar trend. There was no significant difference therefore,  water temperature  in C O D values between  the months,  did not influence the amount of oxidizable organic  Table  5.8  Temporal changes  1n t o t a l o r g a n i c  carbon i n the  p i n e and  spruce l o g e n c l o s u r e  s t u d i e s at Babine  Lake,  1985.  Time  Tree species  Study period  No. of logs  0  2  5 T o t a l Organic  Pine  Spruce  June  = control  10  15  6.5 8.0 7.3 6.0  8.5 9.5 10. 3 11.0  6.0 11.5 13.0 20.5  6.5 11.5 14.0 16.8  10.5 13.5 14.8 17.5  July  C 1 3 5  10.0 11.5 11.2 6. 1  14.5 19.0 14.5  9.0 12.5 17.0 12.8  11.0 13.3 20.0 12.3  June  C 1 3 5  8.0 9.0 11.5 8.7  10.5  10.0 10.0 12.5 12.5  C 1 3 5  9.3 2.3 8.0 6.5  enclosure  (no l o g s )  10.0 1 1.5 14.0 5.2 3.0 7.2 6.8  5.8 4.9 13.5 8.0  20  25  Carbon (mg/l)  C 1 3 5  July  C  (days)  9.5 9. 5 12.0 11.0 6.2 4.7 5.8 7.8  19.0  10.0 12.7 17.5 22.0  7.7 15 .O 23 .8 13.5  11.5 12.5 16.8 16.8  6.8 11.5 24.5 21.5  11.5 9.3 13.0 15.8  13.5  9.0 5.5 4.5 12.5  5.5 3.5 7.3 10.5  7.5 10.0 15.0  8.8 12.8 15.5 9.0 4.2 7.2 12.9  SPRUCE  PINE 80  JUNE I.  (5) :A  .//.  0 80  i  i  i  ~  0  i  i  i  i  i  i  5  10  15  20  25  30  0  changes in COD  concentration for the pine and spruce of  logs in treatment  5  10  15  (5)  20  25  30  TIME (days)  TIME (days) Figure 5.10 Temporal  i  lULY]  | JULY  0  i  (3)  log enclosure studies done at Babine  Lake, 1985 ; number  are given in parentheses. on  58  matter leached from the spruce and pine logs. The DO data was plotted against the C O D results using a computer program for the best fit linear regression to determine existed between  D O and C O D (Figures 5.11  tested by covariance analysis to determine  if a temporal relationship  and 5.12). The linear regressions were  differences between the slopes of the  lines and, if the slopes were the same, between levels. The June experiments for both species of trees (except for the spruce control enclosure) found no significant difference existed between the slopes of the control, and 3 and 5 log treatments ; the slope of the 1 log experiment was slightly higher compared to the other enclosures. Therefore, the number of logs in the enclosures did not significantly affect C O D or DO compared to the control enclosure. There was no significant difference between  levels for (i) the June 1 spruce  log and control enclosures, and (ii) the June and July pine control and .1 log experiments. This indicated that the spruce and pine 1 log treatments of June and July had no significant affect on C O D and D O compared with the control enclosures, i.e. the decrease in DO and the increase in C O D in the 1 log experiments were the result of the static, non-flushing environment of the enclosures. In July, there were significant changes in the slopes of the spruce and pine 3 and 5 log enclosures compared to the June slopes, which indicated a temporal relationship between  (iii)  C O D and DO measurements.  Lignins and tannins  The L-T results for 1985  (L-T)  are shown in Figure 5.13. There was a significant  increase in L-T concentration over the test duration in the 5 and 3 pine and  6S  09  62  spruce log enclosures, although there was no significant difference of L-T leached between  in the amounts  June and July for both tree species.  5.2.3 BACTERIAL POPULATION Bacterial numbers, biomass, and heterotrophy  The bacterial numbers and biomass for all the June spruce and pine log treatments increased significantly (a =  0.05) over the duration of the experiments.  Bacterial numbers (Table 5.9) indicated a change in the physical size of a population (counts/ml), microbial growth,  however, enumeration did not depict an accurate account of  i.e. change in morphological forms from coccoids to filaments.  Therefore, biomass measurements (Mg C/l) were made to best represent changes in the microbial communities of the enclosure experiments (Figures 5.14 and 5.15). Calculations for the conversion of bacterial  numbers to biomass are in Appendix A.  Because of the extensive amount of time required to perform biomass measurements, calculations were done for only three sampling intervals (0, 10, 25 days). The bacterial biomass for the June 5 pine and spruce log enclosures increased several orders of magnitude  from day 0 to the end of the test period.  For example, the initial microbial biomass concentration in the 5 pine and spruce log enclosures was 30.1 Mg C/l and 59.8 Mg CJ, respectively ; this increased to 1370  Mg C/I and 1890 Mg C/l (respectively) However,  after 25 days (Figures 5.14 and 5.15).  in July, a significantly smaller bacterial biomass had developed in  the pine and spruce enclosures, compared to the June concentrations. In the 5 spruce logs enclosure, the June biomass increased from 59.8 Mg C/l to 1890 Mg C/l over the 25 day test period (Figure 5.15). This increase in biomass was four  Table  5.9  Temporal changes  in bacterial  numbers  i n the  p i n e and spruce  log enclosure studies  of  1985,  Babine  Lake.  Time (days)  Tree spec i es  Study per i od  No. of 1ogs  0  2  5 Bacterial  Pine  dune  duly  Spruce  dune  duly  C  = control  enclosure  10  Numbers (counts/ml  C 1  4.1 4.2  7.1 8.7  5.7 7.4  8.6  15 x  20  25  7.2 13.2  10.3 11.8  18.2 13.9  10 ) 5  3  2.0  -  9.3  21.0  18.8  14.9  22.4  5  3.8  -  -  27.7  26.2  15.4  18.6  C 1  9.9 9.3  6.4 8.3  8.0 10.6  9.5 -  11.0 3.5  6.5 15.6  7.2 19.1  3 5  12.3 8.2  5.9 7.7  11.5 7.7  14.8 7.4  5.5 3.4  18.9 21.6  12.6 22.2  C  7.6  4.5  6.6  8.3  47.8  9.6  6.8  1 3 5  8.6 8.2 7.8  2.8 5.3 6.2  7.0 10.8 9.9  12.6 12.0 13.9  17.0 11.8 11.5  9.8 8.8 3.9  8.1 24.8 33.1  C 1  12.0 6.7  16.4 15.6  13.6 10.2  11.7 14.5  11.7 15.7  10.4 9.9  10.5 12.5  3 5  9.8 11.6  15.6 19.1  13.1 13.4  8.6 8.5  13.4 12.6  116.7 201.0  1.9 21.3  (no logs)  64  —  EJ  Yl CO  <  o  3  Legend  1000  • S [Z2 •  800  Control  1 Log 3 Logs 5 Logs  600-  CD  400 CC £ U < CO  / /  200-  mLii  0  0 10 25 TIME (days) Figure 5.14  0 10 25 TIME (days)  Changes in bacterial activity and biomass in the 1985 pine log enclosure experiments done at Babine Lake.  65  —  ™  6  1600-  JUNE  JULY  1400CO GO  <  Legend  1200-  I  1000-i  YZ2 | •  CJ  3 Logs 5 Logs  200-  <  CD  Control 1 Log  ^7  0 10 25 TIME (days)  0 10 25 TIME (days)  Figure 5.15 Changes in bacterial activity and biomass in the spruce log enclosure experiments of 1985 at Babine Lake.  times larger than the July 5 spruce logs experiment 68.2  where the initial biomass of  Mg C/l increased to 430 Mg C/l by the end of the experiment  (Figure 5.15).  The increase in bacterial biomass reflected a succession of bacterial species in the log enclosures, i.e. initially small cocci ( 0.5 - 1 Mm length) to filamentous  microorganisms ( 100  Mm diameter) and rods ( +  Mm length) and larger  cocci and rods by the conclusion of the June experiments 5.16(B). The predominant were coccoids of 0.5  0.75  (Figures 5.16(A) and  bacterial morphological forms in the July log experiments  - 0.75  Mm diameter  and small rods of 1.0  (length). It was assumed that the presence of the aforementioned  - 1.5  Mm  bacterial forms  were introduced into the enclosures from a resident population found on the logs. The rapid growth  of the various microbial species was accompanied by an  increase in microbial activity (biomass was able to assimilate the simple radiolabelled solute). The June heterotrophic pine log treatments,  uptake  however, acetate  rates increased with time for all spruce and uptake values for the spruce enclosures were  significantly higher compared to the enclosures containing pine logs. For example, the June acetate  uptake rates by heterotrophic organisms in the 5 spruce logs  enclosure increased from 0.1 pine logs experiment,  Mg Ac./l/h to 3.2  Mg Ac./l/h over 25 days ; for the 5  microbial activity increased from 0.2 to 2.1  same study interval (Figures 5.14  Mg Ac./l/h for the  and 5.15).  Microbial activity for all spruce log treatments was similar between July, i.e. for the 5 spruce log study acetate uptake increased from 0.1 for both June and July to 3.2 and 5.2  June and  Mg Ac./l/h  Mg Ac./l/h after 25 days, respectively. The  uptake for the pine logs, especially the 1 and 3 log enclosures, were significantly higher in July compared to the June values.  ure 5.16(A)  Bacterial  Figure 5.16(B)  biomass o n i n i t i a t i o n of the 5 pine logs e n c l o s u r e Babine Lake, 1985 ; X1562. 5 u m  Bacterial biomass o n day 25 of the 5 p i n e logs e n c l o s u r e experiment at Babine Lake, 1985 ; X1562. 5 um  at  68 H e t e r o t r o p h i c uptake kinetics  Data for the kinetic uptake study done in July, was plotted using a modified Lineweaver-Burk plot of S/V vs S, where S is the solute concentration and V is o ' o r  the solute uptake rate. A computer program plotted the best fit linear regression to the data (Figure 5.17) (ug/i/h),  ; the kinetic values of maximum uptake velocity or  turnover time or  (h),  V  m a x  and active transport constant or affinity constant K  (h) were obtained from the plot and are listed in Table  t  5.10.  It was assumed that the microbial activity in the control enclosure was statistically similar to that in the spruce and pine enclosures at the beginning of the experiment, thus, initial day kinetics for spruce and pine enclosures is represented by the control data. A detailed discussion on assumptions of the  kinetic  study and definition of the kinetic values is presented in Appendix B.  (i)  Net u p t a k e for l o g e x p e r i m e n t a l e n c l o s u r e s  a. C o n t r o l  The increased V  m a x  arid decreased T^ values after 10 days in the control  enclosure indicated that the heterotrophic population had increased in biomass (Table 5.10)  This observation was supported by an increase in the biomass and  1 4  C  uptake data measured over time at a single substrate concentration. The increased V  m a x  values from 6.8 to 10.9  ug/I/h in 10 days, indicated a  larger biomass to be saturated. The T^ decreased from 3.8 hours to 3.1 hours, which implied that the species composition of the bacterial population present after 10 days was more capable of utilizing acetate as an organic substrate than the population present at the beginning of the experiment. This statement was  69 Table 5.10 Heterotrophic uptake kinetic values obtained on Days 0 and the 5 spruce and pine log enclosures.  Enclosure  Time (day)  0 10 10 10  Control Control Spruce (5 logs) Pine (5 logs)  10 from  Kinetic Results  max (Mg/l/h)  t (h)  t (Mg/D  6.8 10.9 142.9 142.9  3.8 3.1 1.5 0.8  25.9 33.6 219.7 109.7  K  V  IK.  max t (h)  0.3 0.3 0.6 1.3  supported by an increase in the ratio of V /K.. ' ' max t 1  It is important  to realize that the changes in the kinetic values in the  control after 10 days were small and not highly significant (a =  0.05). Therefore,  changes in growth and organic solute uptake kinetics were probably the result of the static (i.e. no water-renewal) environment of the enclosure.  b. Pine  There was a significant difference in kinetic measurements after 10 days in the 5 pine logs enclosure. The increased V  m a x  values indicated an increase in the  heterotrophic population and possible eutrophication (Table 5.10). The decrease in T^ from  3.8 h to 0.8 h and the increase in the affinity ratio  V  / , 0.3 h to 1.3 h K  m a x  t  (Table 5.10) implied that the water soluble extracts from the logs (i.e. molecular weight  organic acids) were rapidly utilized.  low  max  240 220 200 180 160 140 120 100 80  60 40  20  S Figure  5.17  Lineweaver-Burk  plot  of  S/V  vs S on Day  10  for  the  0  20  40  60  80  100  120  (jjg/l) 5 log pine  and spruce enclosures and the  control.  o o  71  c. Spruce  The V  for the 5 spruce logs enclosure was the same as for the 5 pine °  max  r  m  1  log enclosure (Table 5.10). However, this did not imply that the population and uptake kinetics were identical. The T^ and  heterotrophic  values for spruce were  significantly different compared to both the pine and control data, i.e. the spruce values for K  t  and T^ were double the pine values of 109.7  respectively (Table  5.10). The affinity ratio of V  the affinity value of pine, therefore  m  ug/l and 0.8 h,  /Kj for spruce was 0.6 h, half  suggesting a chemical difference  between  the  two tree species.  5.2.4  PHYTOPLANKTONIC ALGAL POPULATION  From the data in Table 5.11,  it was apparent that neither the number of  logs nor the tree species had any adverse effect (a  =  0.05)  on the  phytoplanktonic biomass as measured by chlorophyll _a_ (chl _a). In the July pine log enclosures there may have been a stimulation of phytoplanktonic algal standing crop (biomass) because the chl _a_ concentrations increased compared to the control. There was an increase in chl _a_ values in both the 5 and 3 log treatments, chl _a_ values for 3 pine logs, July experiment, over the experimental  increased from 1.4  i.e.  ug/l to 4.0  period compared to fairly constant values (1.0  - 1.6  the  Mg/l  Mg/l)  '  n  the control enclosure over the same period. There was no significant increase with time in chl _a_ concentrations during June in any of the pine or spruce enclosures. However; the July chl _a_ spruce values were significantly lower than the June concentrations, i.e. for the enclosure that contained 5 spruce logs, the average June and July chl _a_ dropped from 1.7 1.0  Mg/l (Table  5.6).  to  T a b l e 5.11  Temporal  changes In c h l o r o p h y l l  a c o n c e n t r a t i o n In Babine Lake,  the p i n e 1985.  and spruce l o g  Time  Tree species  Study Period  No. of logs  0  2  5  dune  duly  Spruce  dune  duly  C  = control  enclosure  (no  at  (days)  10  15  Chlorophyll  P i ne  e n c l o s u r e s t u d i e s done  20  25  a ^^^/^) t  C  1.4  1.5  1.7  1.4  1.8  1.3  1.4  1 3  1.6 1.2  2.0 1.2  2.0 1.4  1.9 1.6  1.8 1.4  1.7 1.4  1.8 1.9  5  1.4  1.6  1.3  1.7  1.4  1.1  1.9  C 1  1.4 1.4  1.4 1.4  1.6 1.9  1.2 1.5  1.0 1.5  1.5 1.3  1.5 0.9  3 5  1.4 1.4  1.4 1.5  2.6 2.4  1.4 1.9  2.1 1.3  3.0 1.5  4.0 3.0  C 1 3  1.6 1.6 1.6  1.5 1.6 1.7  1-7 1.6 1.7  1.6 1.1 1.2  1.1 I.3 1.3  1.4 1.6 1.8  1.3 1.4 1.3  5  1.7  1.9  1.8  1.8  1.0  2.2  1.6  C 1 3 5  1.0 0.9 1.0 1.0  1.0 1.1 1.1 1.0  0.9 1.1 1.0 1.1  0.6 0.8 0.6 0.7  0.9 1.1 1 .4 1.3  0.8 1.0 0.8 0.8  0.6 0.7 0.7 0.9  logs)  6. DISCUSSION  Most research concerned with pollution from log dumping and storage operations is divided into two categories. Firstly, laboratory experiments where pollutants are leached from log sections, and secondly, organic extractives leached from bark in both laboratory and in situ studies. Schaumburg (1973) performed laboratory and field experiments to determine the water pollution potential associated with (i) water storage of logs and (ii) the loss of bark from logs during different phases of logging operations, specifically, the in situ distribution, extent, and rate of benthic deposits and decomposition. There was considerable difference between the results of the 1984 bark enclosure study and the 1985 log enclosure experiments. The qualitative and quantitative differences between the bark and log studies were : 1. chemical differences ; various materials leached from the bark (i.e. inner and outer bark) compared to the logs (i.e. cambium and heartwood exposed by freshly cut ends), and 2. physical differences ; surface area, quantity and size of the bark chips placed in the enclosure.  The Babine Lake enclosure project attempted to make quantitative and qualitative comparisons between the bark and the log experiments to discern if either bark or logs were a major source of pollution to the aquatic environment. To compare the experimental results, it was necessary to consider experiments with similar bark weights. From Table 6.1, the 1 kg bark experiment had a bark dry weight of 0.9 kg, comparable to the enclosure which contained 1 log where 0.7 kg of bark (peeled from the log) was exposed to the water.  73  74  Table 6.1 A c a l c u l a t i o n of the dry weight of bark a d d e d to t h e 1984 bark a n d 1985 l o g e n c l o s u r e studies for water quality c o m p a r i s o n s .  1984  Treatment (Bark)  1 kg 5 kg 20 kg  (1) (2)  = =  1985  Bark< > (kg, dry wt) 1  0.9 4.3 17.0  Treatment (logs)  1 3 5  Pine Spruce (kg, dry wt of bark)< > 2  0.7 1.8 2.6  0.7 1.9 2.4  contained a 60:40 mixture of pine and spruce bark average of June and July logs  The quantity and quality of extractives leached from the bark and logs was a function of surface area and the chemical composition of the tree tissues. For example, the bark used in the 1984  experiments was peeled from fresh cut pine  and spruce logs and placed in the enclosures where both the outer bark and a portion of the inner bark were in contact with the water. The inner bark contains more extractives and polysacchrides than the outer bark (Section  (phloem) 2.3).  Therefore, the contact water leached from a large surface area that included two anatomically and chemically different  structures. The log study of 1985  involved the  addition of logs to the enclosures, thus, the enclosure water contacted and leached extractives from the outer bark and the exposed ends of the logs. A comparison of water quality data from the 1 kg bark and 1 log studies (Table 6.2)  over an 11 day period illustrated that on a comparable dry weight basis,  the water quality conditions in the 1 kg bark enclosure degraded at a faster rate compared to the 1 log studies. Therefore, in active log dumping and storage areas,  75  Table 6.2 A c o m p a r i s o n of water quality c o n d i t i o n s in e n c l o s u r e s w i t h 1 kg bark (July, 1984) a n d 1 l o g (July, 1985) over an 11 day p e r i o d .  Parameter  1 kg ba rk* )  D O (mg/l) T O C (mg/I) L-T (mg/l) C h l _a_ (Mg/l)  9.4 9.0 0.5 0.1  (1) (2)  = =  1  -  6.0 19.3 11.2 0.3  Pine* ) 2  10.4 11.5 0.4 1.4  - 9.0 - 13.6 - 0.7 - 1.5  Spruce^'  9.6 2.3 0.6 0.9  - 9.1 - 4.9 - 0.5 - 0.8  equivalent to 0.9 kg dry wt. (60 : 40 pine and spruce bark mixture) 1 log enclosure experiment in July, equivalent to 0.7 kg of dry wt. bark  loose floating and sinking bark would have a greater negative effect on water column and benthic ecosystems, compared to floating logs.  6.1  6.1.1  ENCLOSURE EXPERIMENTS WITH BARK - 1984  COMPARISON OF CONTROL AND LAKE RESULTS FOR BARK AND LOG EXPERIMENTS  The statistical (a  =  0.05)  increase in heterotrophy, bacterial biomass,  phytoplanktonic algal biomass, and organic enrichment in the control enclosures compared to the lake values were probably the result of the contained (sealed) environment of the enclosure (no exchange of water with the lake) compared to the dynamic, wind mixed lake (i.e. resuspension of nutrients from the sediments). These results are consistent with Parson's et al. (1977) nutrient enrichment study in  76  controlled experimental  ecosystems. In the control enclosure (seawater, no nutrients)  phytoplanktonic algal biomass, measured by chlorophyll _a, increased slightly by the end of the 30 day experiment. activity (determined  by V  There was no significant change in heterotrophic  ) in the control enclosure with time.  Therefore, it was concluded that the data collected from the lake was not statistically different compared to the data generated by the control enclosure and provided a basis for comparisons between  a perturbed  system (bark treatments), i.e.  Morrison Arm logging operation, and an unperturbed environment  (control),  i.e.  Babine Lake.  6.1.2  WATER QUALITY  The 20 kg bark experiment  resulted in high concentrations of lignins and  tannins (L-T), colour, and total organic carbon (TOC) and rapid decreases in dissolved oxygen (DO), pollution, determined demand (BOD)  pH, and alkalinity.  Similar trends of increased organic  by chemical oxygen demand (COD) and biochemical oxygen  measurements, resulted from bark leaching experiments  by Sproul  and Sharpe (1970), Benedict et al. (1974a), and Gellman (1971). However, there are significant differences in experimental performed  between  design, bark concentrations, and analyses  the Babine Lake study and previous research.  The 20 kg bark experiment  was the highest density (volume  of bark :  volume of water) performed during the Babine Lake experiments. The bark was a 60 : 40 mixture of fresh (recently or 5.4  peeled)  pine and spruce in 3700 litres of water  g of bark/I. This bark loading density was much lighter (200  compared to experiments  times)  performed by Sproul and Sharpe (1970), Asano ef al.  (1974), and Gellman (1971). Table 6.3  compares relative bark concentrations involved  in the various leaching studies and several water quality  measurements.  Table 6.3  A  comparison  quality measurements between several  Gellman  Sproul (1970)  Parameters Bark  of water  -  studied  softwood, hardwood  (1971)  softwood, hardwood  researchers bark  Asano (1974) lodgepole pine  -  leaching  experiments.  Benedict (1974a)  Babine  softwood, hardwood -  (1984)  lodgepole pine, white spruce  Exp. desig nO)  -  leaching of barkpile  leaching of fresh  hardwood and  and aged  softwood  and  hardwood  -  dist. water( )  water( )  Duration of study  -  0  0 - 8 5  Temperature  -  20  -  2 0 0 g bark/I  Extraction  Bark  water  density* ) 6  4  -  69  days  *C  fresh bark  distilled days  room temp.  25  160  25%,  g fresh bark  water  1 0 - 4 0  hours  "C 18%,  10%  static enclosure  leaching of fresh  mix  5  mix/I  -  softwood  and  aged  and  hardwood  -  distilled  -  0  -  room  -  N/A  -  85  softwood  leaching  of  lake water (4)  water  0 - 3 1  days  4  temp.  *  5.4  -  days 12  g fresh  bark /I  wood chip;water volume  1. COD  (mg/l)( ) 2  Sample  (3)  day  0  8.7  0  2510  1  1000 2520  2200  2100  20  3190  2900  3000  121.6  30  1320  3500  3500  1  day  0  780  0  0  10  990  600  600  20  1350  1250  1 100  30  1320  1500  1500  (1) zz brief description of study involved in static laboratory leaching of bark = data acquired via extrapolation of reseachers' studies (2) =  16.8  (mg/l)  Sample  (3)  138.7  10  2. BOD  conversion of L - T carbon content of the T O C to equivalent C O D (Appendix  A)  60:40  pine and spruce  mix  (4)  =  no  added  (5)  =  no  description  poison (dist.=distilled)  (6)  =  wet  weight  of water  of bark  *C  mixed  78  From Table 6.3, there are numerous differences between the Babine Lake bark study and the data collected by the other researchers. These differences include the following : 1. duration of the study, i.e. 40 hours to 85 days, 2. types and species of bark used, i.e. softwood and hardwood mixtures, and 3. concentration of bark leached and the leaching water, i.e. the bark concentrations of other researchers was 200 times greater than the bark concentration of the Babine Lake study.  Although the 1984 experiment  did not measure C O D or BOD, the T O C  measurements were converted into equivalent C O D values to allow comparisons between  my results and previous research (Table 6.4 ; Appendix A). The trends of  increased C O D and BOD with high concentrations of bark observed by Sproul and Sharpe (1970), Gellman (1971), Benedict et al. (1974a), and the Babine 20 kg bark experiment,  indicated that bark in an aquatic environment can be a serious point  source of organic pollution. The 1 kg bark experiment  or 0.3 g of bark/I, was the  lightest concentration of bark used in the Babine Lake leaching studies, and, even at this relatively low level of bark, water quality measurements of T O C and L-T increased and the DO decreased (Table  6.2).  The L-T leached from the Babine Lake 20 kg bark experiment  contributed to  the red-clay colour of the water, however, because these complex compounds are relatively resistant to rapid microbial degradation, other extractives were probably responsible for the rapid depletion of oxygen. Duval (1980) describes a biphasic BOD where  an increase in BOD in the first few days rapidly decreased, replaced  by a long term or ultimate BOD. The initial high BOD is mostly exerted by sugars and other soluble, utilizable organics leached from the bark. The benthal bark decomposition experiment  by Sproul and Sharpe (1970) gives a twofold  explanation  Table 6.4 Calculations to determine the % of total organic  carbon (TOC) comprised  of lignins and tannins  (L-T) and resultant  chemical oxygen demand (COD).  20 kg Treatment  L-T  (day)  (mg/i)  0  0.6  as carbonO)  (mg/l)  TOC  %  TOC as L-T carbon (^)  %  (mg/l)  COD  7.8  3.8  0.7  55.8 60.3  190.9 406.9  17 24  155.0 330.0 327.5 345.0  148.5 293.5  175.5 184.9  360.0  51.4  425.3  31  370.0  198.3  390.0  50.8  456.1  10  0.3  53.6% of tannic acid standard is carbon portion of L-T that is carbon -r TOC calculated equivalent COD attributed to L-T (Appendix  -  A)  -  ( )  (mg/l)  83.1 176.9  5  (1) = (2) = (3) =  L-T  -  3  80  to the rapid rise and decline in the initial B O D . Firstly, the initial BOD is exerted by sugars, rapidly utilized so that after the first 4 days of the experiment there is very little measurable sugar. The second explanation involves a component of the leaching mechanism, whereby, after surface organics have gone into solution, the leaching rate (a function of changes in the concentration gradient with time) would have decreased and this "...slower rate of leaching would then show up as lower BOD  values." (Sproul and Sharpe, 1970). The long term BOD is exerted  by the gradual decomposition of lignins and  other chemically complex structures by lignoceliulolytic bacteria and fungi (Raabe, 1968  ; Benner ef al. 1986). Although, the exact composition of the T O C was not known in the Babine  Lake bark studies, calculations determined that approximately 54% of the T O C was contributed by L-T organic carbon (Table 6.4).  However, it was assumed that  extractives leached from the inner bark (mono- and polysacchrides) contributed to the oxygen deficit created by the heterotrophic population (Section 6.1.3). The acidic nature of these L-T materials was most likely responsible for the initial lowering of the pH. However, after 10 days a slight increase in pH was discernible in all bark treatments.  It was possible that acidic groups comprising a  portion of the chemical composition of lignin and tannin molecules, effectively titrated the alkalinity as they were leached from the bark. Also, microbial modification of some of the macromolecules would slowly generate some CO2 which would come to equilibrium as  H C O y  (pH of 6.8 to 7.3)  and help to  generate some alkalinity. Sproul and Sharpe (1970) found during laboratory simulated benthic deposits (aerated columns) that the pH increased due to the production of CO^  =  and  H C O y  by benthic microorganisms.  The initial decrease in alkalinity, observed in the 20 kg bark treatment and subsequent increase with time, contradicted the inorganic carbon (TIC)  data which  81  exhibited a rapid increase from initial sampling to Day 10, followed by a gradual decline to the end of the experiment. To explain the discrepancy between the two methods of measuring inorganic carbon, alkalinity and TIC were compared on an absolute carbon basis (Table 6.5).  It was possible that some of the TIC was locked  up in cellular material not readily titrated by the alkalinity determination. From Table 6.5, the TIC measurement was greater than the alkalinity concentration ; it was possible that the TIC determination was removing labile carboxy groups from the L-T compounds that would not be measured by the alkalinity  titration.  A more probable explanation for the disparity between alkalinity and TIC values may be found in the definition of alkalinity. The alkalinity measurement is the sum of the total titratable bases in a water sample ; this includes the carbonate (C0  = 3  ), bicarbonate (HCO^"), and hydroxide (OH")  the dissolved carbon dioxide ( C 0 ) 2  molecules. It does not include  or carbonic acid ( H C 0 ) 2  3  components of the  carbonic acid system. The TIC measurement includes dissolved C 0  2  plus the  alkalinity values, therefore, the TIC minus that carbon as alkalinity equals the carbon present in the water as C 0 2  At the beginning of the 20 kg bark experiment, the pH was 7.30 and the alkalinity was 43 mg/l as CaCO^. After 10 days the pH dropped to 5.55  and the  alkalinity carbon effectively converted to TIC. The Henderson-Hasalbach equation (Equator. 6.1)  can calculate the relative proportion of alkalinity to C 0  dropped 7.30  to  pH  =  pK  2  as the pH  5.55.  n  +  log H C 0 " 3  H CO 2  where: prC  =  6.35  s  (6.1)  82  Table 6.5 A comparison of absolute inorganic carbon concentrations between alkalinity and TIC for the 20 kg bark enclosure of 1984.  Days  Alkalinity (mg Inorg. C/l)  0 5 10 17 24 31  Total Inorganic Carbon (mg Inorg. C/l)  5.3 4.0 2.4 3.5 4.1 3.7  6.7 28.5 23.5 12.5 16.0  therefore,  7.30  =  6.35  +  log H C C y H C0 2  8.91  =  3  HC0 " 3  H C0 2  3  There are 9 molecules of H C O y for each molecule of H CC> 2  however,  by day 10 of the experiment,  drop in the carbon ratio.' Therefore,  5.55  =  6.35  +  log H C O y H C0 2  0.15  =  HCCy H C0 2  3  3  3  at pH  7.3,  the decreased pH resulted in a subsequent  83  At the lower pH of 5.55,  there were 6.6 molecules of r ^ C O ^ for every molecule  of HCOj", therefore, the measurable alkalinity decreased. The difference between TIC and alkalinity was possibly a function of the definition of alkalinity and not a discrepancy between either measurement. Therefore, the subtracted difference between TIC and the alkalinity data in Table 6.5 is an indicaton of the amount of dissolved C 0 generate C 0  6.1.3  2  2  in the water sample (heterotrophy will  as assimilated organics are respired).  HETEROTROPHY  The microbial activity for the 20 kg bark enclosure had peaked by 3 weeks and then started to decline. This increase in heterotrophy, especially when compared to the normal growth rates of bacteria, was relatively slow. The slow uptake rate of radiolabelied glucose may have been attributed  to the inhibitory nature of the  tannins, dilution of the radioisotope or both. Microbial activity was measured by the uptake rate of radioactive glucose ; if there was unlabelled glucose leached from the bark (hydrolyzed from carbohydrates) it could effectively dilute the labelled glucose and give a lower uptake rate. Once the natural unlabelled glucose was metabolized the uptake rate of the added labelled glucose would increase. This problem could only be resolved by monitoring actual glucose or sugar concentrations over the experimental period and determining an actual turnover time. The dilution of labelled glucose by unlabelled glucose leached from the bark would explain the rapid rate of oxygen depletion compared to the relatively slow uptake rates of the bacteria during the first 2 weeks of the experiment. Also, the rapid decrease in phytoplankton biomass, dieoff, could also have stimulated microbial activity with unlabelled nutrients.  84  The decrease in heterotrophy by the end of the 20 kg bark and 8 logs experiments may have been a result of nutrient limitation. By the end of the experiment, readily available glucose and other carbohydrates, would be sufficiently depleted to cause potential nutrient  limitation resulting in a decrease in uptake  rates. The initial increase in heterotrophy and subsquent decline was similar to the initial segment of the biphasic BOD described by Sproul and Sharpe (1970). The second stage BOD would be accomplished by microorganisms capable of utilizing the chemically complex structural components of bark. The degradation of lignified plant materials into digestible by-products is performed by a limited population of bacteria (procaryotes) and fungi (eucaryotes). Benner ef al. (1986) states that bacteria are the predominant degraders of the lignin and polysaccharide components of lignocellulosic detritus in aquatic environments and fungi are apparently the major decomposers of terrestrial lignocellulosic materials (Kirk ef al. 1977). Previous research on microbial degradation of plant matter in aquatic environments by Federle and Vestal (1982) and Lee ef al. (1980) indicate a biphasic utilization of plant detritus ; an initial high fungal biomass with an increasing bacterial biomass over time. The colonization and degradation of aquatic plant material' predominantly by either bacteria or fungi is apparently a function of the physical (i.e. temperate or tropical) and chemical (i.e.  acidic or alkaline)  environment.  The main conclusion is that bacteria and fungi are capable of degrading lignocellulosic detritus with some of the degradative end products (lignin and polysaccharides) incorporated into microbial biomass (Benner ef al. 1986). Therefore, it was possible that a portion of the lignins leached from the bark experiments may have been assimilated into microbial biomass provided enough time had elasped for the establishment of a population of lignocellulolytic microorganisms.  There was a higher glucose uptake rate in the 5 kg bark and 8 log experiments when compared to the 20 kg bark experiment. This suggested either inhibitory or dilution effects by the higher bark loading on microbial activity. However, the 1 kg bark treatment showed lower microbial activity which could have meant that lower concentrations of utilizable organic solutes from the bark were limiting the growth of microorganisms and their ability to take up glucose. Thus, there appeared to be a balance between solutes necessary to stimulate microbial activity and leachates which cause some inhibition at higher concentrations. The inhibition experiments conducted with laboratory leachate from 2.7 g white spruce bark/I (equivalent 0.05)  to 10 kg of bark/3700 I water) resulted in a very significant (a =  decrease in the microbial uptake of radiolabelled glucose. This indicated that  bark leachates from white spruce where inhibitory to the metabolism of the sewage and pond water microorganisms. In the enclosures, this inhibition may not be as pronounced because the microbes present will have had time to adapt to the leachate properties. The assimilation process would result in a shift in the species composition of the microbial community. Such a change in bacterial composition was apparent in the log study (Section  6.1.4  PHYTOPLANKTONIC ALGAL POPULATION  There were several problems encountered while conducting the phytoplanktonic primary productivity tests. The major concern was the high radioactive counts in the formaldehyde bottle (blank) compared to the live samples. The high counts for the blanks may have been the result of improperly washed filters. The high blank counts resulted in low productivity found in the lake =  0.7  (average  rjg C/l/h over 21 days for the log. raft study) compared to values obtained  by other researchers, i.e.  =  2.3  tig C/l/h (Stockner and Shortreed, 1975)  at a  86  station north of Topley Landing (Figure 2.1). Therefore, the phytoplanktonic primary production measurement was not repeated for the 1985  log study due to  inherent  difficulties associated with the technique. The observed reduction in chlorophyll _a_ and phytoplanktonic primary productivity in the 20 kg bark enclosure was similar to results obtained by Howard et al. (1979). Howard ef al. (1979) did experiments to determine the effect of kraft softwood pulp mill effluent the effluent  colour on phytoplanktonic primary production. Although  colour concentration (pH maintained  >  9) studied by Howard ef al.  (1979) was not as high compared to the Babine Lake 20 kg bark enclosure experiment,  there was a decrease in phytoplanktonic primary production, attributed  to shading effects on the effluent colour. The decrease in phytoplanktonic biomass and productivity in the 20 kg bark enclosure may be the result of the inhibitory and possible toxic nature of the resin acids (pH  =  5.55)  and a decrease in light transmission, a result of the highly  coloured waters (tannins). determine  However,  I was unable to partition the results to  the major contributing factor in the annihilation of the phytoplanktonic  primary producers.  6.2  ENCLOSURE EXPERIMENTS WITH LOGS - 1985  Discussion of the log enclosure study data was limited by the number of analysis that produced measurable data. For example, as stated in Section 5.2.2. there was no significant change in any of the enclosures in' alkalinity, pH, total Kjeldhal nitrogen, total inorganic carbon, or phytoplanktonic algal numbers compared to the control enclosures. Also, total phosphorus, volatile fatty acids, and carbohydrates were below the detection limits of the analytical techniques. However, the low concentrations of VFA and carbohydrates were the result of their rapid  87  utilization since the literature indicates these materials are present in wood extractions. Therefore, those data which were detected, measured, and deemed significantly different compared to any combination of interactions (Section 5.2) are discussed in the following sections.  6.2.1  WATER QUALITY Dissolved oxygen (DO)  There was a greater decrease in DO levels for the 3 and 5 pine and spruce log treatments in July compared to the 3 and 5 log enclosure experiments of June. The COD concentrations increased with time and number of logs in both months. However, there was very little difference in the leachate concentration between months (COD measurement) ; this indicated that the warmer water temperature of July did not influence the rate of extractive leaching. Therefore, the low DO levels in the July enclosures were probably the result of a combination of factors ; a higher biochemical oxygen demand (BOD) and a decrease in the oxygen solubility with an increase in water temperature. Firstly, a higher BOD would be caused by an increase in utilizable organic substrate or bacterial biomass/activity or both. Although the concentration of oxidizable materials was similar (no significant difference) in June and July, the concentration of TOC was significantly lower in all the July spruce log experiments compared to June ; this suggested that a larger fraction of the July TOC was oxidizable, indicating more organic substrate available for the heterotrophs. An increase in COD or TOC does not guarantee an increase in BOD because the organic matter being oxidized may have a complex chemical structure and, therefore, biologically difficult to degrade.  88  Unfortunately,  the concentration of VFA's and carbohydrates present in the  enclosures were below the detection limits, thus, we were unable to compare either the organic makeup of the June and July leachates or determine  if the oxygen  depletion was a direct function of an increased BOD. If BOD data were available, then the ratio of BOD : C O D and the BOD k-rate would provide a generalization of the biodegradability of the  leachate.  Although the bacterial biomass at all log concentrations for both the spruce and pine enclosures had significantly decreased in July from the June concentrations, the heterotrophic species between  uptake  rates (activity) for acetate were similar for both tree  June and July. Rapid turnover of the bacterial population would  have caused an increase in oxygen consumption, while maintaining  a relatively  constant biomass. Bacterial activity is discussed in detail in Section 6.2.2. Secondly, by calculating the oxygen saturation percentage (DO values divided by the oxygen saturation 6.6),  concentration for that particular water temperture,  it was possible to determine  Table  that the sharp decrease in July pine and spruce  D O levels could be attributed to a combination of lower oxygen solubility, an available organic carbon source, and bacterial  activity. Total organic carbon (TOC) and chemical oxygen demand  There was a significant temporal  (COD)  decrease in the amount of TOC leached  from all the spruce log enclosures in July compared to the June values. The reason for this decrease in spruce T O C concentrations in July may be a function of when the logs were cut (1 week prior to the commencement of the June and July experiments).  Different harvest times may result in a difference  in the availability  water soluble organic matter and the microorganisms ability to utilize various components of the TOC. Also, I was measuring a steady state concentration of  of  Table  6.6  Dissolved  oxygen  saturation for the 1985  log enclosure  studies at Babine Lake.  Time (days)  Tree species  Study period  No. of logs  0  2  5 Oxygen Saturation  Pine  Spruce  =  15  20  25  (percent)  June  C 1 3 5  116.7 124.7 110.2 114.1  110.5 109.5 96.1 93.2  106.6 103.7 93.9 92.4  111.6 109.7 91.1 86.9  98.9 93.7 80.4 74.9  92.7 84.7 76.6 71.3  96.0 99.4 91.3 87.4  July  C 1 3 5  105.6 106.7 105.6 105.6  107.6 110.9 102.0 100.9  96.4 94.3 84.1 80.0  95.4 87.2 69.7 63.6  104.3 96.7 63.4 59.8  102.3 88.6 57.9 47.7  95.9 85.3 32.0 26.6  June  C  111.8  117.7  109.5  95.9  109.5  105.4  94.5  1 3  103.8 106.6  113.6 104.6  107.6 102.8  91.7 83.8  107.6 94.6  97.0 89.5  85.6 82.0  5  107.6  105.6  101.8  78.9  91.9  87.7  80.2  C 1 3 5  104.3 104.3 105.4 105.4  101.5 101.3 98.5 99.7  104.5 103.4 97.0 93.8  109.9 105.4 89.7 85.2  100.0 94.6 72.8 60.9  103.3 92.4 79.3 57.6  98.9 89.1 65.2 36.4  July  C  10  control enclosure  (no logs)  90  TOC and, therefore, higher temperatures  not accounting for rapid removal of utilizable  organics due to  and microbial activity, thus, the leaching rate may not have  changed. The lignins-tannins (L-T) composition of the T O C was different between the bark and log experiments.  In the 20 kg bark experiment,  the was attributed to L-T carbon (Table 6.4). However, pine and spruce log experiments, approximately  S  50% of the T O C of  by the end of the July 5  3.2% and 8.3% of the T O C  measured was L-T carbon (see Table 6.4 for calculations). This indicated a difference in the chemical composition of the T O C between  the bark and log leachates. The  difference in leachate composition would influence the microbiological dynamics of the enclosure ecosystem. Both Schaumburg (1973) and Pease (1974) had similar increases in C O D concentrations with high log densities : water ratios compared to the Babine Lake study. Schaumburg (1973) performed  7 day static log leaching experiments with 2  log sections of Douglas fir, ponderosa pine, and hemlock measuring BOD, BOD k-rate, C O D , BOD : C O D ratio, and reducing sugars. The log sections were approximately  15.5 inch (in.) diameter x 20 in. length and were submerged in tanks  containing approximately  100 I poisoned water. After all logs had their exposed  ends sealed, the logs were then treated,  i.e. for Douglas fir, 1 log with bark was  submerged in one tank and the other fir log had its bark removed before being submerged in another tank. Schaumburg (1973) concluded the following from the softwood log experiments  after removal of the poison :  1. an increase in BOD, BOD k-rate, C O D , BOD : C O D , and reducing sugars, with time, for all 3 species, and 2. that most logs without bark had higher concentrations of the above measurements, indicating that the exposed cambium layer leached more  91  organics and nutrients than the unaltered logs.  The three significant differences between the Babine Lake experiments and Schaumburg's (1973) was (i) the volume of log : volume of water, i.e. 5 pine logs for Babine had a combined volume of 4.5 ft^/3700 litres or 0.035 m^/m^ water, much smaller than 0.6 m^/m^ (Schaumburg, 1973), (ii) the physical state of the log, i.e. peeled, and (iii) poisoned water which allowed the accumulation of readily utilizable organics, i.e. sugars. Although the Babine Lake data showed an increase in C O D with time, the lack of sugar and BOD data limit the scope and accuracy of applying the findings to in situ situations.  6.2.2  BACTERIAL POPULATION Numbers, biomass, and  The uptake rate of  1 4  hetertrophy  C acetate in the June experiments was greater in the  spruce enclosures compared to pine enclosures, however, pine logs had a higher dissolved oxygen demand than spruce. It was possible that the bacterial population in the pine enclosures was capable of utilizing a wider and more available range of unlabelled organic matter for metabolism and growth than the spruce microbial population. Also, the spruce heterotrophs may have been more adept in taking up simple organic molecules, i.e. acetate, than the pine microorganisms. Therefore, discrepancies in the uptake rates between the tree species were possibly a function of the chemical composition of the extractives and bacterial species composition. The uptake of acetate in the 1 and 3 pine log experiments in July was (i) greater than occurred in June, and (ii) higher compared to the 5 pine logs  92  enclosure in July. However, the dissolved oxygen concentration in the 5 pine logs enclosure was 2.5 mg/l by day 25, thus, it would appear that the microbial population was creating a dissolved oxygen demand. The lower rate of acetate uptake in the 5 pine logs experiment was probably a function of isotope dilution due to possible leaching of higher levels of acetate from the logs. It was difficult to rationalize how the lower bacterial biomass of July, for all log concentrations of pine and spruce, was able to maintain uptake rates similar to or greater than the June heterotrophic values. There are several possible explanations for the observed discrepancies. The smaller bacterial biomass that developed in July had a better ability to assimilate the radioisotope demonstrated by their higher uptake ratios of Mg of carbon uptake : Mg of carbon biomass (Table 6.7). The assimilation ability of the smaller morphological forms can be attributed  to a number  of factors. Firstly, the smaller bacteria have a higher surface area : cell volume ratio compared to the filamentous forms, and secondly, higher water temperature. Assuming that the bacterial uptake of acetate follows Michaelis-Menten enzyme kinetics, the aforementioned anomaly may be caused by the effects of  temperature  on enzymatic reactions. The rate of enzyme-catalyzed reactions generally increases with temperature, within the temperature  range in which the enzyme is stable and  retains full activity. The rate of most enzymatic reactions approximately doubles for each 10 ° C rise in temperature  (Q-JQ —  The average water temperture temperature  2.0 ; Lehninger, 1982).  for the June experiments was 9 °C. The  increased to 15.6 ° C for the July studies and, although, the bacterial  biomass decreased in July (compared to June), the higher water  temperature  probably increased the rate of enzyme reactions and acetate uptake. Thirdly, if a greater proportion of the acetate was respired by the microorganisms in July, it would have given a lower net uptake by the cellular material. Also, a faster turnover of the cellular material and recycling of a limited  Table 6.7 A comparison  of the uptake of carbon per ug of bacterial biomass for the 1985 log experiments. 14  Sample days  Tree species  Month  Number of logs  0  10  25  (Mg C/Mg C/h x 14  Pine  Spruce  C  =  10" ) 3  June  C 1 3 5  5.10 6.10 2.30 2.90  -  0.91 0.49 0.91 0.61  July  c 1 3 5  0 0.01 0.64 1.40  -  12.20 2.20 0.43  12.00 12.0 1.62 2.30  June  C 1 3 5  0.74 0 0.51 0.65  0.89 1.40 4.10 1.60  3.30 3.70 1.70 0.68  July  C 1 3 5  0.76 0.34 0.26 0.48  3.70 6.40 6.90 0.63  2.20 9.90 11.00 4.80  control (no logs)  nutrient pool in the enclosures could help to keep the bacterial biomass lower and more active. Lower oxygen concentrations in July support the observed overall higher metabolic activity. Fourthly, predation by heterotrophic protozoans on the bacterial biomass in July could also help explain the observed results. The predatory action of protozoa on the bacterial community is a widely recognized trophic link in aquatic systems.  94  There are abundant field observations and laboratory evidence that heterotrophic microflagellates and ciliates are active predators of bacteria in a variety of environments, i.e. marine and freshwater ecosystems, and activated sludge treatments plants (Sherr and Sherr, 1984 ; Azam ef al. 1983 ; Fenchel, 1977 and 1982 ; Cude, 1979). Sherr and Sherr (1984) state that in situ bacterioplankton production utilizes 10% - 50% of total phytoplanktonic productivity, and although bacterial numbers and biomass tend to be relatively low and constant, bacterial productivity indicates the biomass is turning over rapidly. Heterotrophic protozoan grazing is a major consumer of bacterioplankton. Laboratory evidence has found that microflagellates and ciliates consume 10 to 250 bacteria per protozoan cell per hour (Sherr and Sherr, 1984 ; Berk ef al. 1976 ; Fenchel, 1982). Cude (1979) studied the microflora composition of an activated sludge continuous culture system. In laboratory experiments with defined inocula, morphological structure and cell size of the bacteria were an important factor in selectivity of protozoan prey items. For example, filamentous, spiral, and floe forms of bacteria are resistant to protozoan grazing (compared to single-cell bacteria) by virtue of morphology. Therefore, defined bacterial cultures with a protozoan population are predominantly filamentous. These observations exemplify that species diversity can be increased by increasing the complexity of the system. Thus, certain bacterial populations wouid be overgrown by better adapted bacteria if competition for substrates were the only selection factor (Gude, 1979). For example, in protozoa free cultures, the single celled bacteria would have a competitive advantage over filamentous forms in substrate aquisition. However, the same population could compete successfully if an additional selection factor, i.e. predation, were introduced into the system (Cude, 1979). The results of Gude's (1979) experiments offer a possible explanation to the observed discrepancies between June and July log enclosure studies. It is possible  95  that protozoan predation  upon singled-celled bacteria enabled the  filamentous  population to become the dominant bacterial species during the June time period. In July, the bacterial biomass was smaller (compared to June) and comprised predominantly  of coccoids (0.5  um to 0.75  urn.) with only a few visible strands of  filaments. This suggests possible intense protozoan grazing, low relatively bacterial numbers and biomass with high secondary production. protozoan biomass for the 1985 above explanation  constant  Unfortunately,  enclosure studies was not measured, therefore,  the  is based on conjecture and theory. K i n e t i c u p t a k e study  .  (i) Limitations of the k i n e t i c uptake study  There were several drawbacks with the kinetics study ; 1. the chemical composition of the leachate was not 2. an unknown in situ acetate  determined,  concentration. Although the in situ solute  concentration was less than 0.1  Mg/', isotope dilution was a possible  problem. If unlabelled acetate was leached from the logs in concentrations greater than 20 ug/l, it could have effectively  dilute the labelled acetate and  give a lower uptake rate, and 3. the values obtained from this study described the dynamics of the heterotrophic  population for the July study period. For example, the  values of the bacterial biomass were a function of water temperature, bags (nutrient limitation), leachate composition, and initial stocking concentrations of bacteria, phytoplankton, and zooplankton.  kinetic sealed  96  The results of the experiment experimental  cannot be superimposed upon the June  results because of differences in the aforementioned  physical, chemical,  and biological conditions. For example, the July bacterial biomass for spruce and pine was significantly smaller compared to the June biomass, therefore, values for V  m a x  ,  T^, and  probably would not apply to the  the July  heterotrophic  dynamics of the June experiments.  (ii) Pine  The increased V  m a x  value for the 5 pine logs enclosure suggested that the  composition of the heterotrophic  community had increased compared to the initial  population. The increase in heterotrophic  biomass was probably the result of a  population of bacteria having been introduced into the enclosure by the pine logs which proliferated  under the conditions of leachate generation (Figures 5.16(A) and  5.16(B)). The increased biomass and acetate  data confirm an increase in the  population. The drop in T^ coupled with the increase in V bacterial biomass in the (i.e.  max  bacterial  -/Kj. indicated that the  5 pine logs enclosure had a high affinity for the  extractives  acetate) leached from the logs and that the extractives were utilizable. This was  confirmed by the low levels of volatile fatty acids in the enclosure.  (iii) Spruce  Although the V pine experiment,  of the spruce kinetic experiment  was the same for the  the result of an increase in bacterial biomass (Figure 5.15  Table 5.10), the K difference  m a x  t  and  and T^ values for spruce were higher ; this indicated a  in log leachate composition between  chemical difference between  the two species of trees. A  log leachates was confirmed by comparing both the  97  C O D values, where more oxidizable material was leached from pine vs spruce (i.e. 47 mg/l C O D and 32 mg/l C O D after 10 days, respectively) and the affinity ratio of V  /K ITlaX  (5 spruce logs was half that of the pine value). This difference in I  extractive composition dictated the degree or intensity of heterotrophic utilization of the leachate. For example, the bacterial population in the spruce log enclosure was not able to utilize the log leachate as quickly as the population that developed in the pine enclosure (as indicated by lower dissolved oxygen).  (ii) I m p l i c a t i o n s  The biomass,  14  C-acetate uptake, and kinetic uptake data provided  information on the microbial dynamics of log storage in a slow flushing environment. The bacterial study, coupled with the DO, C O D , and TOC results, indicated : 1. temporal differences in bacterial biomass and chemical composition of log extractives, 2. a quantitative  and qualitive difference in the organic matter leached from  pine and spruce, that directly affected the ability of heterotrophs to utilize the leachate, 3. the leachate from pine had a faster turnover time and higher degree of biodegradability than spruce and the chemical composition of the pine extractives had more oxidizable matter than spruce ; this suggested that pine leachates were more susceptible to microbial degradation than spruce log leachates (supported by the lower dissolved oxygen values for pine compared to spruce), and 4. the low T  and high V  /K. ratio for pine and spruce indicated a potential  eutrophication problem in a log storage facility.  98  6.2.3  PHYTOPLANKTON BIOMASS  There was no significant increase or decrease in the phytoplanktonic algal biomass for the enclosure experiments. However, there was significantly lower phytoplankton biomass present in the July spruce enclosures compared to the June values. By comparing the lake and control chl _a_ data, the difference between June and July chl a values may be the initial stocking concentration of the phytoplankton. The 1985  studies indicated that there was no apparent toxic effects  from the logs on phytoplanktonic biomass.  6.3  COMPARISON BETWEEN THE ENCLOSURE STUDIES AND THE 3 YEAR FIELD STUDY BY WESTWATER RESEARCH  In conjunction with the 1984  and 1985  enclosure studies, Westwater Research  Center conducted a 3 year investigation commencing in June 1983 to compare pre-impact data (prior to the initiation of fall logging and log transportation at the Morrison Arm dump site and. Topley Landing dewatering site) to post-impact conditions studied from May to August of 1985 1985a and 1985b). Intensive and the dewatering sites (Figure 2.1).  (Levy ef al. 1984,  and Levy ef al.  sampling stations were set up at both the dump site  site with results from the affected areas compared to reference  Colour, turbidity,  L-T, conductivity, DO, temperature,  C O D , and  bacterial biomass measurements were conducted at depths of 0, 1, and 2 m on May 15, May 27, June 11, June 21, and August 8,  1985.  The water quality results from both active sites were a function of location and lake morphology. Water quality measurements at the dewatering  site did not  show any degradation when compared to the reference stations or to the pre-impact data collected in 1983  (Levy ef al. 1984). It was concluded that the  99  unsheltered dewatering site, adjacent to the main basin of the lake, was subjected to strong winds that blow the long fetch of the basin which resulted in good water circulation and a well mixed water mass (characterized by a weak thermocline ; Levy et al. 19856). However, the water mass in the sheltered dump bay at Morrison Arm became stratified and inhibited water circulation, thus, creating a potential precursor for water quality degradation. Figures 6.1 and 6.2 depict the enclosed, sheltered log storage bay at Morrison Arm. For example, the post impact water quality at the dump and storage site deteriorated by May 27 when DO dropped to from  >  &  5 mg/l  8 mg/l on May 15 (Levy ef al. 1985b). The D O deficit coincided with an  increase in C O D in the surface waters (average of 40 mg/l for sampling depth range of 0 - 2 meters) compared to the reference site average of 24 mg/l (0 - 2 meters). The D O concentration at the 2 meter reference site remained constant at approximately 10 mg/I. The rapid warming of the surface waters in these protected areas of Morrison Arm dump site created ideal conditions which stimulated microbial activity to utilize organic materials leached from the logs resulting in deterioration of water quality. By July 16, all logs had been moved to the dewatering site at Topley Landing ; the DO in the dumping bay rebounded to ^ 6,  8 mg/l by August  1985. Assuming that a uniform bundle of logs is only 50% submerged in the  Morrison Arm storage area and a thermocline at 2 m (leachate mixing in the top 2 m of water), the 166,000 m  3  of logs stored in Morrison Arm, 1985, would  represent a loading level of approximately 0.4  m^/m  3  water (Hall, unpubl.). This is  an order of magnitude higher than the 5 log enclosures where the log loading level was 0.035 m / m 3  3  water. Although, the log volume : water volume was greater  at Morrison Arm than the enclosures, the decreased DO concentrations in the enclosures was paralleled by a similar decrease in DO at Morrison Arm ; this  100  Figure 6.1 Aerial photograph of the Morrison dump site in mid-winter, 1984.  101  Figure 6.2 Aerial p h o t o g r a p h of the M o r r i s o n d u m p site o n M a y 14, 1985.  implied that similar leaching mechanisms and trophic level interactions were occurring under experimental  conditions and in situ.  7.  CONCLUSION  The objective of the bark and log studies was to determine the effects of log dumping and storage on the Babine Lake aquatic environment, specifically, changes in water quality and microbiology. A major concern generated by the environmental impact studies was the dissolved oxygen deficit. The depletion of D O was a function of heterotrophic activity, chemistry of the wood and bark, the physical limitations of the enclosures, and oxygen solubility (function of water temperature). The sealed (no water-renewal) construction of the enclosures approximated the morphological scenairo of a shallow, sheltered, non-circulating aquatic ecosystem, and allowed the buildup of organic carbon leached from stored logs and log debris. The quality and quantitity of extractives leached from the logs and bark provided a readily available and utilizable organic source for an enterprizing population of bacteria. Microbial activity combined with the leached organic carbon produced a D O demand. From the enclosure experiments, I would hypothesize that the storage of logs in the enclosed, protected bay of Morrison Arm would result in organic enrichment, D O depletion, and degradation of water quality. The storage facilities and morphology of Morrison Arm provided ample conditions for the processes of organic enrichment and oxygen depletion. For example, the shallow and protected bay of the storage facility would restrict wind generated reaeration of the surface waters, and with the low volumes of water exchange (tracer studies revealed a flow of 3 m^/hr ; Power, 1987), would allow for an accumulation of organic material. These two conditions would contribute to a potentially severe D O deficit which could adversely affect habitat quality of the aquatic environment of Morrison Arm.  102  103  From the bark studies, it was found that bark and logs were significant contributors to the degradation of the enclosure environment. Therefore, in a log dumping and storage operation trees that have lost bark during the bundling and dumping phase of operations create a pollution potential via (i) concentration gradient of leached organic compounds above the 2 m thermocline, and (ii) loose bark and benthal accumulation. The 3 year study of Morrison Arm by Westwater  Research Centre  found that the benthic accumulation of bark had resulted in the annihilation of the benthic community. In conclusion, I would recommend for future  log storage operations :  1. more stringent criteria for the siting of log dumping and storage facilities, i.e. locate the storage sites in areas where there is adequate water circulation to prevent the buildup of organics and the exertion of a BOD to such a degree that low oxygen conditions can develop, 2. rapid removal of logs from constricted circulation areas especially if warm epilimnetic conditions develop, and 3. possible aeration of the dump and storage areas to oxygenate the water and destratify the system, i.e. this is easily accomplished in winter logging areas where an aeration system is already in place to keep the storage bay ice free in winter.  8. APPENDIX A  1. C a l c u l a t i o n of m i c r o b i a l u p t a k e of l a b e l l e d  i) specific activity of acetate  =  ii) molecular weight of acetate iii) specific activity  =  57.6 mCi/mmole =  59, therefore  0.976 uCi/ug of acetate  C a l c u l a t i o n e x a m p l e ; heterotrophy  i) 100 X =  acetate.  experiments  15,000 dpm  ii) 1 ml of isotope = iii) 1 uCi =  2.2 x 10  150,000 dpm 6  dpm  iv) 0.976 MCi x 2.2 x 10 v) 150,000/2,145,200 =  6  =  2,145,200 dpm/Mg acetate  0.06985 Mg acetate  added to each syringe  vi) acetate  uptake expressed as Mg solute/l/hr was determined  uptake  dpm of syringe x 0.6985 Mg Ac. x _1_ hours x 10 m l  =  150,000 dpm  where ; Ci = dpm =  3  by ;  9 ml  T  x 1000 ml 10 ml  Currie =  disintegrations per minute  10 ml is the sum of 9 ml sample water +  104  1 ml isotope.  105  2. Bacteria numbers (counts per  ml)  Calculation example ; June, Day 25 for 5 pine logs  i) total filter count (within a grid of 50 units x 50 units) ii) total number of fields iii) mean field count (i  = ii)  =  46.6 =  v) areal microscope conversion factor* vi) total bacterial count/ml (iii =  (area of filter)  326  7  iv) volume of sample water filtered  *  =  1 ml =  40,000  iv x v) =  1.863  x 10**  -4- (area of counting grid) for that particular microscope  power  3,  Bacteria biomass  i) measure (randomly)  dimensions of rods, cocci, and filaments, keeping tally of  the number of rods and cocci ; count the number of times a filament crosses (intercepts)  the grid.  ii) assume volume of cocci = iii) assume volume of rod = where  ; r = I  =  sphere ; therefore, volume of sphere =  r^7r4/3  cylinder ; volume of cylinder = r^-irl  radius length  iv) use the following equation to determine the volume of rods and cocci in sample volume of bacteria in sample =  (Av. cell vol. rods) x (%  (Av. cell vol. coccoids) x (%  of rods)  of coccoids))  +  106  where ; Av. =  average  cell =  bacterium  vol.  volume  =  C a l c u l a t i o n e x a m p l e ; June, Day 25 for 5 pine logs  v) Total number (no.)  of bacteria measured =  no. of rods measured =  13 or 76%  no. of cocci measured =  4 or 24%  vi) grams of carbon/ml =  (0.75  Mm  3  x 0.76)  =  (0.696 Mm /cell) x 5.6 x 10"  =  3.9 x 1 0  3  - 1 3  17  + 13  (0.524 u m  3  x  0.24)  g of carbon/Mm * ) 3  1  g of carbon/cell x. counts/ml (1.863 x 10 , 6  bacteria counts/ml from previous Section (vi) = where ; (1)  =  7.26  x 10"^ g carbon/ml for rods and cocci  5.6 x 10"  (Bratbak,  13  average carbon content/vol. of bacteria  1985)  C a l c u l a t i o n e x a m p l e ; June, Day 25 for 5 pine logs  vii) assume volume of filaments  =  viii) tally no. of grids counted =  volume of cylinder 4  ix) tally no. of filaments per grid ; i.e. first grid had 1 filament x) count no. of times the filament(s) intercepts the grid ; i.e. 9 xi) length of filament (within grid)  =  9 x 5  =  45 Mm  Mm (width of 1 grid unit)  xii) for grids with more than 1 filament, summate the lengths of all filaments xiii) total length of filaments from the 4 grids =  410 Mm  107  xiv) average length/grid (xiii  viii) =  xv) average length/ml =  Mm x 40,000 (areal conversion factor)  102.5  =  4.1 x 10  xvi) average filament vol./ml = xvii) average g carbon/ml =  Mm  6  1.16  1.16  102.5 Mm  3  x 10^ Mm /ml 3  x 10^ Mm /ml x 5.6 x 10"^ 3  carbon/Mm =  Total bacterial biomass =  6.49 x 10"  (xvii) +  (vi)  =  1.37 x 10"  =  1370 x 10'  =  6  7  g carbon/ml  6.49 x 10"  7  +  g carbon/I  4. Percentage of T O C as tannic acid (L-T measurement)  i) tannic acid =  ^y^^2^4d  ii) molecular weight  =  iii) molecular weight C  1701 =  12  % tannic acid that is carbon = = (912  1701)  x 100  =  g  3  g carbon/ml  6  3  76 X 12 912 g, therefore  53.6% of tannic acid is carbon  7.26  x 10"  7  108 5. Conversion of L-T carbon of T O C into equivalent C O D  (i) C O D =  C +  0  o  2  C0  2  where : C = T O C 0  2  =  COD  (ii) know 53.6% of L-T is carbon and that the oxidization of L-T is represented by, CvcH^O-. /o 52 4 b  +  660  o  0  therefore, 2112  g of 0  912 mg/l C =  2112 mg/l  1 mg/l of C =  76CO + 2 0  2  2  26H„0 2  is required to oxidize 912 g of C.  2.3 mg/l 0  0 2  2  (COD)  By multiplying the L-T carbon by 2.3, the C O D for the L-T portion of the T O C can be determined.  9. APPENDIX B The microbial kinetics of solute uptake was performed assuming the following assumptions : 1. Michaelis-Menten uptake kinetics, substrate-saturation curve (Wright and Hobbie, 1965  ; Hobbie and Wright, 1965  ; Allen, 1968), where the rate  equation for a 1 substrate enzyme catalyzed reaction is described by Equation B.1.  V  = o  V S —max— K  t  where : V V  + S  max =  Q  S = K  t  (B.1)  =  =  maximum uptake rate r  substrate uptake rate substrate concentration active transport constant,  2. steady-state conditions, where the rate of forming the enzyme-substrate complex is equal to the rate of breakdown of the enzyme-substrate complex (Button, 1986), 3. natural substrate concentration is equal to zero (Hall ef al. 1972) concentrations <  0.1  ; VFA  ug/l from analytical measurements,  4. uniformly labelled substrate is added to a mixed, random population of heterotrophs, and 5. no significant removal of substrate by phytoplankton (Wright and Hobbie, 1965).  109  110  (i) D e f i n i t i o n of the k i n e t i c values  a. M a x i m u m  uptake rate o r V  max  (ug/I/h)  V, is an indicator or measure of the relative size of a population of max heterotrophs able to utilize a given substrate, therefore, V proportional to biomass. V different  max  is considered to be  enables one to compare the heterotrophic activity of  max  bodies of water and is a good indicator of pollution (high V  eutrophication ; Gocke, 1977a ; Wright and Hobbie, 1966). The V function of (i) temperature,  m a x  m a x  =  value is a  (ii) population size, composition, and physiological state  of the heterotrophic population, and (iii) availability of substrate.  b. T u r n o v e r t i m e or T^ (hours)  Turnover time is the time required by a natural population of heterotrophs for the complete uptake of a concentration of substrate equivalent to the concentration. Thus, T^ is an indicator of the intensity of heterotrophic  natural  utilization  and a measure of the biological importance of a substrate (Gocke, 1977a and 1977b ; Allen, 1967). For example, a low T^ indicates that the substrate is rapidly utilized. High T  t  values suggests (i) that the substrate is not utilized to any large  extent, (ii) only a small population of heterotrophs are present, (iii) zooplankton grazing.  c. Active transport constant or  The Michaelis-Menten K  {  (Mg/l)  is used as an indicator of the affinity of an  organism for a substrate ; a lower K  indicating higher affinity (Wright and Hobbie,  111  1966 ; Law and Button, 1977 ; Healey, 1980). The Michaelis-Menten rate equation (Equation B.1) defines K^. as the substrate concentration at which the initial reaction velocity is half maximal (1/2 V  =  x  K  t  ; Lehninger, 1982).  is independent of  the number of heterotrophs but is a function of the heterogeniety of a heterotrophic population (Hobbie and Wright, 1965). The term V ' /K. is used to best describe the affinity (attraction) between max t ' acetate (substrate) and bacteria (Healey, (slope of the rate equation of V  q  1980 ; Button, 1986). The V  m a x  /K  t  value  vs S ) evaluates the competitve ability of an  organism in a nutrient limited system. An increased ratio of V strong affinity for the substrate present.  m a x  / K ^ indicates a  10. ABBREVIATIONS  1. i.e.  =  example  2. D O  =  dissolved oxygen  3. C O D  =  chemical oxygen demand  4. BOD =  biochemical oxygen demand  5. TOC =  total organic carbon  6. TIC  =  total inorganic carbon  7. L-T =  lignins and tannins  8. 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