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Biopharmaceutical properties of solid dosage form 1968

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THE BIOPHARMACEUTICAL PROPERTIES OF SOLID DOSAGE FORMS The Operating C h a r a c t e r i s t i c s of a Continuous Flow D i s s o l u t i o n Apparatus by WENDY WENG WAH WOO B. S. P., U n i v e r s i t y of B r i t i s h Columbia, 1 9 6 6 A THESIS SUBMITTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF Master of Science i n Pharmacy (M. S. P.) i n the Facult}' of Pharmacy We accept t h i s t h e s i s as conforming to the req u i r e d standard. THE UNIVERSITY OF BRITISH COLUMBIA A p r i l , 1 9 6 S In p r e s e n t i n g t h i s t h e s i s i n p a r t i a l f u l f i l m e n t o f t h e r e q u i r e m e n t s f o r an advanced degree a t the U n i v e r s i t y o f B r i t i s h C olumbia, I agree t h a t t h e L i b r a r y s h a l l make i t f r e e l y a v a i l a b l e f o r r e f e r e n c e and Study. I f u r t h e r agree t h a t p e r m i s s i o n f o r e x t e n s i v e c o p y i n g o f t h i s t h e s i s f o r s c h o l a r l y p u r p o s e s may be g r a n t e d by the Head o f my Department or by h.i.'S r e p r e s e n t a t i v e s . I t i s u n d e r s t o o d t h a t c o p y i n g or p u b l i c a t i o n o f t h i s t h e s i s f o r f i n a n c i a l g a i n s h a l l not be a l l o w e d w i t h o u t my w r i t t e n p e r m i s s i o n . Department o f *7U OLA^iOi-H^ The U n i v e r s i t y o f B r i t i s h Columbia Vancouver 8, Canada Date f 7 - H - L? fr. i i ABSTRACT A completely automatic continuous f l o w d i s s o l u t i o n procedure was developed and t e s t e d . P e r t i n e n t d i s s o l u t i o n c o n d i t i o n s were i n v e s t i g a t e d and chosen to study the d i s s o l u t i o n c h a r a c t e r i s t i c s of seven brands of phenylbutazone t a b l e t s . A pumping system enabled the simulated d i g e s t i v e f l u i d to flow from,the d i s s o l u t i o n v e s s e l i n t o the fl o w c e l l of a r e c o r d i n g spectrophotometer f o r a continuous r e c o r d i n g of the drug c o n c e n t r a t i o n i n the d i s s o l u t i o n medium, which was g r a d u a l l y changed from an a c i d i c medium t o a ba s i c one. From the " i n v i t r o t t data obtained by t h i s t e s t procedure, a T^Q^ value of 120 minutes was chosen as a l i m i t of acceptance f o r the t e s t products. The " i n v i v o " c h a r a c t e r i s t i c s of s i x of the brands were compared w i t h those observed f o r a pharmaceutically acceptable product. Of the seven t e s t products, only f o u r were acceptable on the b a s i s of both the " i n v i t r o " and the " i n v i v o " data. C o r r e l a t i o n of the " i n v i t r o " and the " i n v i v o " data r e s u l t e d i n " l e a s t squares" l i n e s w i t h negative slopes. M. Pernarowski, Ph. D. Supervisor i i i TABLE OF CONTENTS Page I . INTRODUCTION 1 I I . LITERATURE SURVEY 5 I I I . THE "IN VITRO" CHARACTERISTICS OF PHENYLBUTAZONE (a) The S o l u b i l i t y of Phenylbutazone i n Buff e r e d S o l u t i o n s 28 (b) Determination of the I s o s b e s t i c P o i n t 2 9 (c) Determination of A b s o r p t i v i t y Value 3 0 IV. THE OPERATING CHARACTERISTICS OF A CONTINUOUS FLOW DISSOLUTION APPARATUS (a) D e s c r i p t i o n of the Apparatus 35 (b) C a l i b r a t i o n of the Recorder 3 6 (c) The E f f e c t of S t i r r i n g Rate on D i s s o l u t i o n 3 9 (d) The E f f e c t of Pumping Rate on D i s s o l u t i o n 42 (e) Test Procedure 45 (f) R e s u l t s 4 6 V. THE "IN VIVO" CHARACTERISTICS OF PHENYLBUTAZONE TABLETS 5 6 V I . "IN VIVO" - "IN VITRO" CORRELATION 6 1 i v Page V I I . DISCUSSION 67 V I I I . SUMMARY 70 IX. BIBLIOGRAPHY 71 LIST OF TABLES Page I . The E f f e c t of S t i r r i n g Rate on D i s s o l u t i o n 41 I I . The E f f e c t of Pumping Rate on D i s s o l u t i o n 43 I I I . D i s s o l u t i o n Data f o r Seven Brands of Phenylbutazone Tablets 50 ^50% anc* ^9<yfo V " a l u e s f ° r Seven Brands of Phenylbutazone Tablets 52 V. Areas under Blood Curves f o r Seven Brands of Phenylbutazone Tablets 6 0 v i LIST OF FIGURES Page 1. The E f f e c t of pH on the S o l u b i l i t y of Phenylbutazone 32 2 . The I s o s b e s t i c P o i n t of Phenylbutazone 33 3 . C a l i b r a t i o n Curve f o r Phenylbutazone i n Simulated I n t e s t i n a l F l u i d at 240 mu 34 4 . Diagram of Continuous Flow D i s s o l u t i o n •Apparatus 37 5 . C a l i b r a t i o n Curve f o r Recorder E x t e r n a l to the Spectronic 505 33 6 . pH Changes at a Pumping Rate of 60 ml./min. 44 7 a . D i s s o l u t i o n P r o f i l e s f o r Two Brands of Phenylbutazone Tablets 47 7 b . D i s s o l u t i o n P r o f i l e s f o r Two Brands of Phenylbutazone Tablets 43 7 c . D i s s o l u t i o n P r o f i l e s f o r Three Brands of Phenylbutazone Tablets 49 V l l Page 8. Concentration of Phenylbutazone i n Serum a f t e r A d m i n i s t r a t i o n of Product A to Subject 1 59 9 . C o r r e l a t i o n of T^Q^ Values w i t h " I n Vi v o " Data f o r Subject 1 63 1 0 . C o r r e l a t i o n of T^Q^ Values with "In Vi v o " Data f o r Subject 1 64 1 1 . C o r r e l a t i o n of T^Q^ Values w i t h "In V i v o " Data f o r Subject 2 65 1 2 . C o r r e l a t i o n of T^Q^ Values with " I n Vi v o " Data f o r Subject 2 66 v i i i ACKNOWLEDGEMENT The author wishes to express her sincere g r a t i t u d e to Dr.. M. Pernarowski f o r suggesting and s u p e r v i s i n g t h i s study. The author i s indebted to Mr. R.0. S e a r l f o r h i s many h e l p f u l suggestions throughout the research. Thanks i s a l s o due t o Mrs. J . Naylor f o r her t e c h n i c a l a s s i s t a n c e i n the l a b o r a t o r y . I . INTRODUCTION Tablet d i s i n t e g r a t i o n i m p l i e s that the s o l i d dosage form has broken up i n t o smaller p a r t i c l e s . However, i t does not n e c e s s a r i l y mean that the a c t i v e i n g r e d i e n t has been rel e a s e d from the primary p a r t i c l e s . This was f i r s t pointed out i n the e a r l y 1950's by s e v e r a l researchers (23, 28) and more r e c e n t l y by'Levy and Hayes (13) and Yen (34). By the mid 1950 Ts, Chapman and h i s co-workers (5, 6), i n separate st u d i e s on products c o n t a i n i n g r i b o f l a v i n and p-amino s a l i c y l i c a c i d , showed that p h y s i o l o g i c a l a v a i l a b i l i t y c ould be p r e d i c t e d from the d i s i n t e g r a t i o n times determined by the o f f i c i a l d i s i n t e g r a t i o n t e s t (30). However, t h e i r r e s u l t s d i d suggest that the more important c r i t e r i o n to consider i n r e l a t i o n to a v a i l a b i l i t y i s the release of drug from the primary drug p a r t i c l e . Even though the c o r r e l a t i o n between d i s i n t e g r a t i o n time and d i s s o l u t i o n has r e c e n t l y been shown to be i n v a l i d (1, 9, 13), the d i s i n t e g r a t i o n time i s s t i l l an i n d i c a t i o n of drug re l e a s e because, i f the s o l i d dosage form remains i n t a c t , the surface area f o r d i s s o l u t i o n i s l i m i t e d . As shown by the Noyes-Whitney Equation (13), the amount of drug d i s s o l v e d i s dependent on the surface area of the d i s s o l v i n g s o l i d . da _' KS ( Cs - C ) ctt " where a = amount of drug d i s s o l v e d i n time t 2 S = surface area of the d i s s o l v i n g s o l i d K = s o l u t i o n r a t e constant of the s o l i d C1, - con c e n t r a t i o n of the d i s s o l v i n g s o l i d i n the medium Cs = co n c e n t r a t i o n of the d i s s o l v i n g s o l i d i n the d i f f u s i o n l a y e r surrounding the so l u t e p a r t i c l e s . D i s s o l u t i o n i s the process whereby s o l i d p a r t i c l e s go i n t o s o l u t i o n . I t can be considered as s p e c i f i c types of heterogeneous r e a c t i o n s i n which a mass t r a n s f e r i s e f f e c t e d through the net r e s u l t of escape and d e p o s i t i o n of so l u t e molecules at the s o l u t e - s o l v e n t i n t e r f a c e . From t h i s l a y e r of d i s s o l v e d p a r t i c l e s , d i f f u s i o n o c c u r s s c a r r y i n g the d i s s o l v e d s o l u t e from the i n t e r f a c e i n t o the main body of the d i s s o l u t i o n medium. Because the r e a c t i o n at the i n t e r f a c e i s much slower than the convection t r a n s p o r t process, i t u l t i m a t e l y determines the r a t e of d i s s o l u t i o n (32). Even though i t i s g e n e r a l l y recognised to-day that d i s i n t e g r a t i o n times do not n e c e s s a r i l y bear a r e l a t i o n - ship to the " i n v i v o " a c t i o n of the t a b l e t , the d i s i n t e g r a t i o n time t e s t i s s t i l l the only o f f i c i a l " i n v i t r o " check on drug r e l e a s e from t a b l e t s . I t has been s t a t e d that t h i s t e s t i s inadequate and should be replaced by a d i s s o l u t i o n t e s t . Since the e a r l y 1960's,. s e v e r a l workers have designed d i s s o l u t i o n t e s t s ranging from Levy's beaker method (13) to completely automated systems (24). 3 The great v a r i a t i o n s i n the equipment used f o r " i n v i t r o " d i s s o l u t i o n s t u d i e s can be a t t r i b u t e d to the wide d i f f e r e n c e s i n the p h y s i c a l forms of the t e s t preparations as w e l l as to the attempts to d u p l i c a t e " i n v i v o " c o n d i t i o n s . However, b a s i c a l l y they a l l c o n s i s t of a v e s s e l f o r the d i s s o l u t i o n medium and the t e s t p r e p a r a t i o n , a means of a g i t a t i o n , a sampling system and a means of c o n t r o l l i n g the temperature of the d i s s o l u t i o n medium. Most of the " i n v i t r o " d i s s o l u t i o n s t u d i e s have been done i n a s t a t i c system, i . e . , the d i s s o l u t i o n medium remains unchanged throughout the e n t i r e d i s s o l u t i o n run. This system may encounter a s a t u r a t i o n problem e s p e c i a l l y w i t h the l e s s s o l u b l e drug p r e p a r a t i o n s . Several researchers (7, 22., 33) have used a continuous f l o w system to c a r r y out d i s s o l u t i o n s t u d i e s . This type of system e l i m i n a t e s the p o s s i b i l i t y of s a t u r a t i o n of the medium by the drug. I t may a l s o simulate " i n v i v o " c o n d i t i o n s more than the s t a t i c system because under " i n v i v o " c o n d i t i o n s the d i g e s t i v e j u i c e s are continuously being secreted and absorbed by the mucosal c e l l s of the g a s t r o - i n t e s t i n a l t r a c t . I n an attempt t o i n v e s t i g a t e the p o t e n t i a l s of t h i s type of system, an apparatus f o r continuous flow was developed with,a gradual change i n the pH of the medium, from that of g a s t r i c j u i c e to i n t e s t i n a l j u i c e . This system was i n v e s t i g a t e d and using a standard product, p e r t i n e n t 4 d i s s o l u t i o n c o n d i t i o n s were e s t a b l i s h e d f o r t h i s study. Under these c o n d i t i o n s , seven brands of commericially a v a i l a b l e phenylbutazone t a b l e t s were t e s t e d . The " i m v i t r o " d i s s o l u t i o n data obtained was then c o r r e l a t e d w i t h " i n v i v o " data obtained f o r s e v e r a l s u b j e c t s . 5 I I . LITERATURE SURVEY The o f f i c i a l U.S.P. d i s i n t e g r a t i o n apparatus ( 30 ) has the basic p a rts needed i n a d i s s o l u t i o n t e s t . Workers have e i t h e r used the d i s i n t e g r a t i o n apparatus i t s e l f or modified i t to c a r r y out d i s i n t e g r a t i o n as w e l l as d i s s o l u t i o n s t u d i e s . The apparatus c o n s i s t s of a basket rack assembly that can be r a i s e d and lowered at a constant frequency r a t e of between twenty-eight to t h i r t y - t w o c y c l e s per minute through a distance of not more than s i x cm. and not l e s s than f i v e cm. The volume of the d i s s o l u t i o n medium i n the t h e r m o s t a t i c a l l y c o n t r o l l e d v e s s e l (35°C - 39°C) i s such that at.the highest point of the upward stroke the wire mesh remains at l e a s t 2 . 5 cm. below the surface of the medium and descends to not l e s s than 2 .5 cm. from the bottom of the v e s s e l on the downward str o k e . The basket rack ss;? assembly c o n s i s t s of s i x v e r t i c a l l y h e l d open-ended tubes whose lower ends are covered by a sieve of v a r y i n g mesh s i z e . Each tube i s provided w i t h a s l o t t e d and pe r f o r a t e d c y l i n d r i c a l d i s k of p l a s t i c . The di s k j u s t f i t s the tube and moves up and down w i t h the movement of the basket as i t i s r a i s e d and lowered. Using t h i s apparatus, Schroeter and h i s co-workers ( 25 ) determined the d i s i n t e g r a t i o n times and d i s s o l u t i o n r a t e s of sev e n t y - s i x l o t s of compressed t a b l e t s , i n c l u d i n g 6 an anti-inflammatory s t e r o i d , a sulfonamide, an a n t i - d i a b e t i c agent and an a s p i r i n - p h e n a c e t i n combination. Without stopping the a g i t a t i o n of the U.S.P. t a b l e t d i s i n t e g r a t i o n apparatus, a l i q u o t s of the d i s s o l u t i o n medium were withdrawn at d i f f e r e n t i n t e r v a l s f o r a n a l y s i s . Time f o r 50% of the drug to go i n t o s o l u t i o n (t^o^ ) o r the amount of drug i n s o l u t i o n at a s p e c i f i c time was used as the c r i t e r i o n f o r the r a t e of d i s s o l u t i o n . D i s i n t e g r a t i o n times were obtained w i t h and without the use of the p l a s t i c d i s k s . These workers found f o r the s t e r o i d a h i g h l y s i g n i f i c a n t l i n e a r c o r r e l a t i o n between the r a t e of d i s s o l u t i o n and the average d i s i n t e g r a t i o n time without the use of the p l a s t i c d i s k s . This c o r r e l a t i o n was absent when d i s k s were used. For the sulfonamide t a b l e t s , a s i m i l a r c o r r e l a t i o n was observed when sodium c h l o r i d e was present as an i n g r e d i e n t . Both the a n t i - d i a b e t i c agent and the a s p i r i n - p h e n a c e t i n combination showed no s i g n i f i c a n t c o r r e l a t i o n between d i s s o l u - t i o n r a t e and d i s i n t e g r a t i o n time. The r e s u l t s i n d i c a t e d that there was an extreme s p e c i f i c i t y i n the absence or presence of a r e l a t i o n s h i p between r a t e of d i s s o l u t i o n and d i s i n t e g r a t i o n times. When a q u a n t i t a t i v e r e l a t i o n s h i p e x i s t e d , the " l e a s t squares" l i n e s r e l a t i n g the v a r i a b l e s were found to vary widely depending upon the p a r t i c u l a r drug as w e l l as on the other adjuvants i n the t a b l e t . The use of p l a s t i c d i s k s i n the 7 d i s i n t e g r a t i o n t e s t was observed to mask r e a l d i f f e r e n c e s between the v a r i o u s l o t s t e s t e d . Middleton and h i s co-workers (19), using the U.S.P. d i s i n t e g r a t i o n apparatus, found a s i g n i f i c a n t r e l a t i o n s h i p between d i s i n t e g r a t i o n time, d i s s o l u t i o n r a t e and p h y s i o l o g i c a l a v a i l a b i l i t y of r i b o f l a v i n i n sugar-coated t a b l e t s . The t a b l e t s were immersed f o r t h i r t y minutes i n simulated g a s t r i c f l u i d and the remainder of the time i n simulated i n t e s t i n a l f l u i d . Without stopping the a g i t a t i o n of the apparatus, a l i q u o t s of the simulated d i g e s t i v e f.Mi f l u i d s were withdrawn at var i o u s i n t e r v a l s f o r a n a l y s i s . From the r e s u l t s , these workers concluded that the d i s i n t e g - r a t i o n time provided a v a l i d i n d i c a t i o n of the r a t e at which r i b o f l a v i n went i n t o s o l u t i o n from the sugar-coated t a b l e t s , thus e i t h e r d i s i n t e g r a t i o n time or d i s s o l u t i o n r a t e can provide a u s e f u l estimate of the p h y s i o l o g i c a l a v a i l a b i l i t y of r i b o f l a v i n from sugar-coated t a b l e t s . However, t h i s r e l a t i o n s h i p may not e x i s t f o r other drugs. I n t h e i r study of the d i s s o l u t i o n r a t e s of eighteen brands of commericially a v a i l a b l e tolbutamide t a b l e t s i n g a s t r i c f l u i d , Brudney and h i s co-workers (1) found that there Was no c o r r e l a t i o n between d i s s o l u t i o n r a t e s and the d i s i n t e g r a t i o n times. D i s i n t e g r a t i o n time was determined by the o f f i c i a l method of the Food and Drug D i r e c t o r a t e ( 3 ). D i s s o l u t i o n was determined by the method described by Levy and Hayes (13) w i t h m o d i f i c a t i o n s . S i x l i t r e s of 0.1 N.HC1 c o n t a i n i n g 0.2% NaCl was used as the d i s s o l u t i o n medium. The s t i r r i n g r a t e was maintained at 250 r.p.m. and the temperature O n c o n t r o l l e d at 37 C + 0.5 C. Twenty-five ml. samples of the medium were withdrawn through a nylon f i l t e r pad at f i v e minute i n t e r v a l s over a s i x t y minute p e r i o d . F o l l o w i n g each withdrawal, twenty-five ml.' of f r e s h medium was added to maintain a constant volume. Lu and h i s co-workers (18), i n t h e i r comparative , study o f ' t w e n t y - s i x brands of commericially a v a i l a b l e tolbutamide t a b l e t s , found that there was no c o r r e l a t i o n between d i s i n t e g r a t i o n time and d i s s o l u t i o n r a t e i n g a s t r i c f l u i d as had been reported e a r l i e r ( 1 ) , but that there was a s t a t i s t i c a l l y s i g n i f i c a n t c o r r e l a t i o n between d i s i n t e g - r a t i o n time and d i s s o l u t i o n r a t e i n i n t e s t i n a l f l u i d . The d i s i n t e g r a t i o n times were determined by us i n g the Food and Drug D i r e c t o r a t e method. The d i s s o l u t i o n r a t e s were deter- mined i n both g a s t r i c and i n t e s t i n a l f l u i d s of pH 1.5 and 7.5 r e s p e c t i v e l y . Two and a h a l f l i t r e s of the s o l u t i o n was heated to 37°C i n a t h r e e - l i t r e g l a s s j a r . The t a b l e t was placed at the bottom of the j a r and the s o l u t i o n s t i r r e d by a blade s t i r r e r set at one in c h from the bottom of the j a r at 300 r.p.m. In two of the l o t s , the d i s i n t e g r a t i o n time exceeded the Food and Drug D i r e c t o r a t e l i m i t of s i x t y minutes, whereas the other twenty-four l o t s d i s i n t e g r a t e d w e l l w i t h i n .this l i m i t . D i s s o l u t i o n r a t e s were c a l c u l a t e d 9 on the b a s i s of the amount i n mg. d i s s o l v e d i n one hour and the time r e q u i r e d to reach 2 0 $ and 4 0 $ d i s s o l u t i o n i n g a s t r i c f l u i d and 5 0 $ and 9 0 $ d i s s o l u t i o n i n i n t e s t i n a l f l u i d . The r e s u l t s showed a considerable v a r i a t i o n i n the d i s s o l u t i o n r a t e s among the d i f f e r e n t brands t e s t e d . These f i n d i n g s tend to r e i n f o r c e the observations of Schroeter and h i s co-workers ( 2 5 ) that the existence of a r e l a t i o n s h i p between d i s s o l u t i o n and d i s i n t e g r a t i o n time depends on the p a r t i c u l a r drug as w e l l as on the c o n d i t i o n s under which the d i s s o l u t i o n and d i s i n t e g r a t i o n t e s t s are c a r r i e d out. Among the var i o u s f a c t o r s t o be considered i n the design of a d i s s o l u t i o n t e s t , the means and i n t e n s i t y of a g i t a t i o n are v a r i e d most and have been reported to have a s i g n i f i c a n t e f f e c t on the d i s s o l u t i o n r a t e . Hamlin and h i s co-workers (S) used three d i f f e r e n t methods to study constant surface p e l l e t s of two polymorphic forms of methylprednisolone compressed at high pressure. One form had greater water s o l u b i l i t y and hence should have a grea t e r i n i t i a l d i s s o l u t i o n r a t e i n water. The f i r s t method was that of Nelson ( 2 1 ) , a method i n which fr e e convection, d i f f u s i o n c o e f f i c i e n t and concen- t r a t i o n were s t a t e d to be the r a t e determining f a c t o r s . The only m o d i f i c a t i o n was that i n s t e a d of determining weight l o s s of the p e l l e t s , the f l u i d surrounding the s t e r o i d p e l l e t s was assayed s p e c t r o p h o t o m e t r i c a l l y . In the second method, the p e l l e t s were h e l d i n a polyethylene holder at 10 the center of ov a l f o u r f l u i d ounce b o t t l e s c o n t a i n i n g 120 ml. of deionized water. The b o t t l e s were attached to the machine described by Wurble (31). A f t e r r o t a t i o n f o r d i f f e r - ent i n t e r v a l s of time, the f l u i d surrounding the p e l l e t was assayed s p e c t r o p h o t o m e t r i c a l l y . The dimensions of the p e l l e t before and a f t e r exposure to the d i s s o l u t i o n medium were a l s o determined. The machine was r o t a t e d at 6 r.p.m. and at 12 r.p.m. The t h i r d method was s i m i l a r to the second except that the machine described by Souder and Ellenbogen (29) was used i n s t e a d of the Wurble machine. The b o t t l e s were r o t a t e d at 40 r.p.m. during exposure i n a'constant temperature bath h e l d at 37°C. I t was found that when the i n t e n s i t y of a g i t a t i o n or the v e l o c i t y of the d i s s o l u t i o n medium across the d i s s o l v - i n g s o l i d was of a low order of magnitude, a s i g n i f i c a n t d i f f e r e n c e i n d i s s o l u t i o n r a t e s was observed f o r both forms of the s t e r o i d . When the speed of r o t a t i o n of the wheel i n the Wurble machine was increased from 6 r.p.m. to 12 r.p.m., the r a t e s of d i s s o l u t i o n of the two polymorphs became equal. R o t a t i o n at 40 r.p.m. i n the machine described by Souder and Ellenbogen showed no observable s i g n i f i c a n t d i f f e r e n c e i n the d i s s o l u t i o n r a t e s of the two polymorphs. I n the " i n v i v o " s t u d i e s c a r r i e d out by implanting the p e l l e t s i n r a t s , the two polymorphs showed s i g n i f i c a n t l y d i f f e r e n t r a t e s of weight l o s s . The " i n v i v o " data tend to i n d i c a t e that the " i n v i t r o " t e s t i n which there was a r e l a t i v e l y low i n t e n s i t y 11 of a g i t a t i o n c o r r e l a t e d best with the " i n v i v o " r e s u l t s . Levy (11), i n h i s study of the e f f e c t s of a g i t a t i o n on d i s s o l u t i o n , used two methods tha t v a r i e d g r e a t l y i n a g i t a t i o n i n t e n s i t y - the beaker method (13) and the o s c i l l a t i n g tube method. In the second method, a p l e x i g l a s s c y l i n d e r w i t h a 100-mesh s t a i n l e s s s t e e l wire screen on the bottom was attached to the basic u n i t of the U.SI'P. d i s i n t e g r a t i o n apparatus and immersed i n a beaker c o n t a i n - i n g 300 ml. of 0.1 N.HC1 at 37°C With the apparatus i n motion, a t a b l e t was dropped i n t o the c y l i n d e r and the medium sampled at i n t e r v a l s by a f r i t t e d - g l a s s immersion f i l t e r tube f o r a n a l y s i s . Two p r o p r i e t a r y brands'of a s p i r i n were t e s t e d . One contained a l k a l i n e a d d i t i v e s w h i l e the other d i d not. With the beaker method, which u t i l i z e d a very low a g i t a t i o n r a t e and permitted the s o l i d p a r t i c l e s to remain as an aggregate on the bottom of the beaker, the t a b l e t s c o n t a i n i n g a s p i r i n and the a l k a l i n e a d d i t i v e s d i s s o l v e d much more r a p i d l y than the p l a i n a s p i r i n t a b l e t s . This was i n agreement w i t h the " i n v i v o " a b s o r p t i o n t e s t s c a r r i e d out. The d i s s o l u t i o n h a l f - t i m e of one was approx- imately three times that of the other. On the other hand, there was p r a c t i c a l l y no d i f f e r e n c e i n the d i s s o l u t i o n r a t e s of the two t a b l e t formulations when t e s t e d by the o s c i l l a t i n g tube method, which i n v o l v e d r e l a t i v e l y high a g i t a t i o n i n t e n s i t i e s and caused the s o l i d p a r t i c l e s to be dispersed. 12 Levy a t t r i b u t e d t h i s d i f f e r e n c e to the f a c t that i n an aggregate of d i s i n t e g r a t e d t a b l e t s o l i d s , the a l k a l i n e components of the buf f e r e d a s p i r i n t a b l e t caused an increase i n the pH of the micro-environment from pH 1 to about pH 5.6. This increase i n pH r e s u l t e d i n a more r a p i d d i s s o l u t i o n of the a s p i r i n p a r t i c l e s . This e f f e c t was absent when a s p i r i n and the a l k a l i n e components were p h y s i c a l l y separated by i n t e n s i v e a g i t a t i o n . The method developed by Levy and Hayes (13) i n t h e i r study of the d i s s o l u t i o n r a t e of p l a i n and bu f f e r e d a s p i r i n t a b l e t s has been used by Levy and other workers i n d i s s o l u t i o n s t u d i e s w i t h only minor m o d i f i c a t i o n s . The d i s s o l u t i o n assembly c o n s i s t e d of a 400-ml. Pyrex G r i f f i n beaker immersed i n a constant temperature bath adjusted o o to 37 C + 0.1 C. A three-blade, 5-cm. diameter polyethylene s t i r r e r was attached t o an e l e c t r o n i c a l l y c o n t r o l l e d s t i r r - i n g motor. Two hundred a n d . f i f t y ml. of 0.1 N.HG1 was placed i n the beaker and allowed, to e q u i l i b r a t e t o 37°C and the a c e t y s a l i c y l i c a c i d t a b l e t was dropped along the side of the beaker. The polyethylene s t i r r e r was immersed i n the d i s s o l u t i o n medium to a depth of 27 mm. and a c c u r a t e l y centered by means of a guide. The s t i r r i n g r a t e was 59 r.p.m. - j u s t s u f f i c i e n t to keep the s o l u t i o n homogeneous but low enough to a l l o w the s o l i d s of the d i s i n t e g r a t e d t a b l e t to remain as an aggregate at the centre of the bottom of the beaker w i t h i n an area of one or two sq. cm. Seven ml. samples 13 were taken at f i v e , t en, twenty and t h i r t y minutes by means of a f r i t t e d - g l a s s immersion f i l t e r tube of medium p o r o s i t y . S i x brands of p l a i n a c e t y s a l i c y l i c a c i d t a b l e t s , one brand of the b u f f e r e d drug, and one brand of calcium a c e t y s a l i c y - l a t e carbamide complex product were t e s t e d . S i x t a b l e t s were subjected to the d i s s o l u t i o n t e s t ; another s i x to d i s i n - t e g r a t i o n t e s t i n 0.1 N'.HCl by the U.S.P. method. The r e s u l t s showed that there was a great d i f f e r - ence i n the' s o l u t i o n r a t e s of the v a r i o u s brands of p l a i n t a b l e t s . The t a b l e t w i t h the calcium s a l t d i s s o l v e d at a f a s t e r r a t e than a l l the other products t e s t e d . This was expected due t o the greater s o l u t i o n of the s a l t i n aqueous medium. They found that the d i s s o l u t i o n curve d i d not progress a r i t h m e t i c a l l y and that i t would take much longer f o r the second h a l f of the drug to go i n t o s o l u t i o n . The r a p i d l y d i s s o l v i n g products e x h i b i t e d longer d i s i n t e g - r a t i o n times than the more slow l y d i s s o l v i n g products. I t was concluded that the d i s i n t e g r a t i o n time which i s often a l l u d e d to as an index of drug a v a i l a b i l i t y i s no c r i t e r i o n of a v a i l a b i l i t y or r a t e of s o l u t i o n . When d i s i n t e g r a t i o n times are abnormally long, the r a t e of s o l u t i o n and absorp- t i o n of a drug w i l l be s e r i o u s l y a f f e c t e d since d i s i n t e g r a t i o n time i s j u s t the time r e q u i r e d by. the t a b l e t to f a l l apart i n t o reasonably small p a r t i c l e s and not the time r e q u i r e d f o r the drug to go i n t o solution.. 14 Using the method j u s t described (13), Levy and h i s co-workers (17) c o r r e l a t e d the e f f e c t of s t i r r i n g r a t e s w i t h the i n t e s t i n a l absorption of three d i f f e r e n t dosage forms of a s p i r i n . These dosage forms d i f f e r e d markedly i n drug absorption r a t e as w e l l as i n the p r i n c i p a l mechanism i n v o l v e d i n the release of drug to the d i s s o l u t i o n medium. One was a r a p i d l y d i s i n t e g r a t i n g t a b l e t c o n t a i n i n g a s p i r i n as micro-encapsulated p a r t i c l e s , another was a rapidly- d i s i n t e g r a t i n g " p l a i n " a s p i r i n t a b l e t and the t h i r d was a r a p i d l y d i s i n t e g r a t i n g t a b l e t c o n t a i n i n g a s p i r i n and a l k a l i n e a d d i t i v e s . D i f f e r e n t s t i r r i n g r a t e s i n a c l o c k - wise d i r e c t i o n u s i n g the p r e c i s i o n s t i r r i n g apparatus of Levy and Tanski (15) were used w i t h 0.1 N.HC1 as the d i s s o l - u t i o n medium. The r e s u l t i n g comparison of d i s s o l u t i o n at 60 r.p.m. with plasma l e v e l s obtained by " i n v i v o " s t u d i e s showed that the " i n v i t r o " d i s s o l u t i o n from the micro-encapsulated p a r t i c l e s was too r a p i d r e l a t i v e to the other dosage forms. D i s s o l u t i o n from the p l a i n t a b l e t s was found to be very much more s e n s i t i v e than from the micro-encapsulated p a r t i c l e s which r e l e a s e drug by a d i f f u s i o n process. At 50 r.p.m., i t was found that the r a t i o of " i n v i v o " absorption r a t e s from the p l a i n t a b l e t s and the micro-encapsulated p a r t i c l e s was equal to the r a t i o of the d i s s o l u t i o n r a t e s . At 30 r.p.m., the more r a p i d l y absorbed p l a i n t a b l e t s were found to d i s s o l v e 15 more sl o w l y than the much more slow l y absorbed micro-encap- s u l a t e d p a r t i c l e s . In t h i s study, i t was observed that a change of only 20$ i n s t i r r i n g r a t e s made a d i f f e r e n c e between s u c c e s s f u l c o r r e l a t i o n or f a i l u r e w i t h " i n v i v o " data. Changes i n the composition of the d i s s o l u t i o n medium was subsequently reported to have s i m i l a r e f f e c t s . Besides s t i r r i n g r a t e , Levy and h i s co-workers (16) found that c e r t a i n f o r m u l a t i o n f a c t o r s have an e f f e c t on the d i s s o l u t i o n r a t e of the a c t i v e i n g r e d i e n t . Using the method described ( 1 3 ) , these workers observed that the d i s s o l u t i o n r a t e of s a l i c y l i c a c i d i n compressed t a b l e t s increased w i t h decreasing granule s i z e but t h i s increase was not s t r i c t l y p r o p o r t i o n a l to the increase i n the apparent surface area of the granules. The increase i n d i s s o l u t i o n r a t e from the t a b l e t s c o n t a i n i n g 20 - 40 mesh and 40 - 60 mesh granules was not as great as the increase i n the apparent surface area since the d i s i n t e g r a t e d t a b l e t p a r t i c l e s remain aggregated on the beaker bottom due to the low i n t e n s i t y of a g i t a t i o n . The much greater d i s s o l u t i o n of the t a b l e t s w i t h 60 - 30 mesh granules was due to the f a c t that these granules were small enough to be dispersed somewhat i n the medium despite the low i n t e n s i t y of a g i t a t i o n , thus p e r m i t t i n g the moving solvent to come i n contact w i t h a gr e a t e r p o r t i o n of the p o t e n t i a l l y a v a i l a b l e s u r f a c e . An increase of s t a r c h content i n the t a b l e t s from 5 - 20$ r e s u l t e d i n an increase 16 i n the d i s s o l u t i o n r a t e of s a l i c y l i c a c i d , probably because of more r a p i d and thorough d i s i n t e g r a t i o n of the granules. D i s s o l u t i o n r a t e s were a l s o found to increase w i t h an increase i n the precompression pressure of the t a b l e t s . This may be due to the f r a c t u r i n g of the drug p a r t i c l e s at the higher s l u g g i n g pressure, thus y i e l d i n g s maller primary p a r t i c l e s . Fragmentation of the more h i g h l y compressed granules during subsequent t a b l e t i n g may a l s o occur. F i n a l l y , the s o f t e r granules obtained at the lower precompression pressures are more l i k e l y to undergo bonding during t a b l e t - i n g and thus y i e l d l a r g e r granules. In t h e i r study of the e f f e c t s of l u b r i c a n t s on d i s s o l u t i o n r a t e s of t a b l e t s , Levy and Gumtow (12) used the beaker method (13) and the r o t a t i n g d i s k method ( 1 4 ) . These workers found th a t magnesium s t e a r a t e , a hydrophobic l u b r i - cant, decreased a p p r e c i a b l y the d i s s o l u t i o n r a t e of s a l i c y l i c a c i d t a b l e t s compressed from granules of the pure drug while sodium l a u r y l s u l f a t e , a h y d r o p h i l i c l u b r i c a n t , had the opposite e f f e c t . This enhancing e f f e c t of sodium l a u r y l s u l f a t e was even gr e a t e r w i t h t a b l e t s made from granules tha t contained s a l i c y l i c a c i d and s t a r c h . Experiments w i t h n o n d i s i n t e g r a t i n g d i s k s of s a l i c y l i c a c i d i n d i c a t e d that the more commonly used hydrophobic l u b r i c a n t s (magnesium s t e a r a t e , aluminum s t e a r a t e , s t e a r i c a c i d , t a l c ) decreased the e f f e c t i v e drug-solvent i n t e r f a c i a l area and thereby decreased the r a t e of d i s s o l u t i o n of the drug, while 1 7 water-soluble l u b r i c a n t s (sodium o l e a t e , sodium l a u r y l s u l f a t e ) d i d not have t h i s e f f e c t . The d i s s o l u t i o n r a t e enhancing e f f e c t of sodium l a u r y l s u l f a t e was not due to any m o d i f i c a t i o n of the micro-environmental pH or s o l u b i l i z - a t i o n by m i c e l l e s , but r a t h e r to the b e t t e r p e n e t r a t i o n of the solvent i n t o the t a b l e t s and t h e i r component granules and the r e s u l t i n g greater a v a i l a b i l i t y of drug surfa c e . Levy and S a h l i ( 1 4 ) , i n t h e i r comparative study of the g a s t r o - i n t e s t i n a l absorption of a c e t y s a l i c y l i c a c i d and i t s aluminum s a l t , c o r r e l a t e d the u r i n a r y s a l i c y l a t e e x c r e t i o n w i t h the d i s s o l u t i o n r a t e obtained by the r o t a t - i n g d i s k method. The compressed t a b l e t s were mounted on P l e x i g l a s holders w i t h p a r a f f i n wax so that only one surface of the t a b l e t was exposed. The holder was connected to an e l e c t r o n i c a l l y c o n t r o l l e d p r e c i s i o n s t i r r i n g motor. A two hundred ml. q u a n t i t y of the d i s s o l u t i o n medium was placed i n a 500-ml. three-neck round-bottom f l a s k , which was immersed i n a constant temperature bath adjusted to 37°C. A f t e r temperature e q u i l i b r a t i o n , the t a b l e t attached to the holder was immersed i n t o the d i s s o l u t i o n medium to a depth of one i n c h , w i t h the s t i r r e r turned on at 555 r.p.m. A l i q u o t s of f i v e or ten ml. were removed from the f l a s k at appropriate i n t e r v a l s of time f o r a n a l y s i s . A s i m i l a r volume of medium was added to maintain a constant volume. The t a b l e t was weighed before and a f t e r the d i s s o l u t i o n run as a check on the assay. The c o n d i t i o n s of the experiment was such that 18 the c o n c e n t r a t i o n of drug i n the medium was maintained at a small f r a c t i o n of i t s t o t a l s o l u b i l i t y . In 0.1 N.HC1, the d i s s o l u t i o n r a t e of a c e t y s a l i c y - l i c a c i d was found to be 65.1 mg./hr.cm. and that of aluminum a c e t y s a l i c y l a t e i n terms of s a l i c y l i c a c i d was 8.88 mg./hr.cm. S i m i l a r d i f f e r e n c e s were found when a l k a l i n e medium was used. S i m i l a r l y , " i n v i v o " s t u d i e s showed that u r i n a r y s a l i c y l a t e e x c r e t i o n from subjects t a k i n g aluminum a c e t y s a l i c y l a t e was markedly l e s s than that from subjects t a k i n g the a c i d . Since the a bsorption of s a l i c y l a t e s i s r a t e - l i m i t e d by t h e i r d i s s o l u t i o n r a t e i n g a s t r o - i n t e s t i n a l f l u i d s , these workers concluded that the l e s s r a p i d absorption and subsequent e x c r e t i o n of aluminum a c e t y s a l i c y l a t e was probably due to i t s slow d i s s o l u t i o n . Recently, S e a r l and Pernarowski (26) evaluated'cthe d i s i n t e g r a t i o n times and d i s s o l u t i o n r a t e s of twenty-three brands of phenylbutazone t a b l e t s . The " i n v i v o " c h a r a c t e r - i s t i c s of three brands were compared w i t h that observed f o r a c l i n i c a l l y acceptable product by a comparison of the l e v e l s of the drug i n the blood a f t e r o r a l a d m i n i s t r a t i o n . D i s i n t e g - r a t i o n times were determined by usi n g the Erweka t a b l e t d i s i n t e g r a t i o n apparatus. The procedure i s i n a p u b l i c a t i o n i s s u e d by the Food and Drug D i r e c t o r a t e ( 4 ) . S i x t a b l e t s were placed i n the apparatus and the mean d i s i n t e g r a t i o n times c a l c u l a t e d . D i s s o l u t i o n r a t e s were determined by an 19 apparatus s i m i l a r t o that of Levy and Hayes (13). Two and a h a l f l i t r e s of simulated i n t e s t i n a l f l u i d i n a t h r e e - l i t r e g l a s s j a r was allowed to e q u i l i b r a t e i n a constant temper- ature water bath set at 37°C + 1°C. One t a b l e t was placed i n a c y l i n d r i c a l wire basket (2 .2 cm. i n diameter and 2.8 cm. i n length) which was attached below the i m p e l l e r of a s t i r r - i n g s haft (three-blade, Teflon-coated p r o p e l l e r , 5 cm. i n diameter). The shaft which was connected to a F i s h e r S t e d i - Speed s t i r r e r was i n s e r t e d to a depth of ten cm. below the surface of the l i q u i d and r o t a t e d at e x a c t l y 100 r.p.m. i n a clockwise d i r e c t i o n . Ten ml. a l i q u o t s were taken at f i f t e e n - m i n u t e i n t e r v a l s f o r the. f i r s t hour, at t h i r t y - m i n u t e i n t e r v a l s f o r the second hour and hourly t h e r e a f t e r f o r seven hours. R e s u l t s showed that nine of the twenty-three products had d i s i n t e g r a t i o n times of more than t h i r t y minutes. Twelve products (some w i t h d i s i n t e g r a t i o n times greater than t h i r t y minutes and some wi t h d i s i n t e g r a t i o n times l e s s than t h i r t y minutes) were s e l e c t e d f o r more extensive study. Of these twelve products, three were found to have a d i s i n t e g - r a t i o n time of more than t h i r t y minutes w i t h a range of more than t h i r t y minutes. From the d i s i n t e g r a t i o n time data, i t would appear that the drug i n these t a b l e t s may not he a v a i l a b l e to the p a t i e n t . D i s s o l u t i o n s t u d i e s showed that four of the twelve products had TCQ^ value of more than 20 120 minutes but most of t>he twenty-three products d i s i n t e g - r a t e d and r e l e a s e d t h e i r phenylbutazone content to the medium q u i c k l y . A standard T^Q^ value of 120 minutes was chosen by these workers on the b a s i s of the data a v a i l a b l e . I t was not p o s s i b l e to c o r r e l a t e the " i n v i v o " data to t h a t obtained " i n v i t r o " . For the t a b l e t s examined, the best c o r r e l a t i o n was s t i l l to t a b l e t d i s i n t e g r a t i o n time. However, maximum d i s i n t e g r a t i o n times would be more meaningful than mean d i s i n t e g r a t i o n times. Nash and Marcus (20) used a p e r i o d i c solvent exchange method to study d-amphetamine s u l f a t e " s u s t ained r e l e a s e " capsules and tripelennamine h y d r o c h l o r i d e "sustained r e l e a s e " t a b l e t s . The t e s t sample was put i n t o a 600-ml. Buchner type funnel w i t h a medium p o r o s i t y f r i t t e d d i s k f i l t e r bed and g e n t l y a g i t a t e d i n 400 ml. of simulated g a s t r i c j u i c e . F i n e r p o r o s i t y f i l t e r s prevent proper drainage of the f l u i d while coarser types leak during sampling p e r i o d s . P o s i t i v e a i r pressure was maintained between sampling periods to prevent leakage. A f t e r t h i r t y minutes, 200 ml. was drawn o f f by vacuum through the f i l t e r bed i n t o the s u c t i o n f l a s k and c o l l e c t e d i n a 250 ml. beaker. A . f o u r - i n c h standard two-way stop cork fused i n t o the bottom of the s u c t i o n f l a s k f a c i l i t a t e d the removal of f l u i d samples during the course of each run without u p s e t t i n g the p o s i t i v e pressure at the bottom of the f i l t e r bed. An a d d i t i o n a l 200 ml. p o r t i o n of simulated g a s t r i c f l u i d was added to the funnel to replace 21 the withdrawn volume. Further samples were withdrawn at one and a h a l f , two, th r e e , f i v e , seven and twenty-four hour i n t e r v a l s , each time w i t h replacement of the f l u i d . Norby (22) developed a continuous solvent exchange system to study the re l e a s e p a t t e r n of "su s t a i n e d r e l e a s e " t a b l e t s . One u n i t of the drug was placed i n a beaker c o n t a i n - i n g ten ml. of d i s s o l u t i o n medium at t h e r m o s t a t i c a l l y c o n t r o l l e d temperature. This s o l u t i o n was kept at a constant l e v e l w i t h the r e s e r v o i r of f r e s h medium to ensure a constant volume i n the d i s s o l u t i o n v e s s e l . The medium from the d i s s o l u - t i o n v e s s e l flowed through tubing which had a cott o n f i l t e r at the open end i n t o a c o l l e c t i n g v e s s e l at a r a t e ( 0 - 1 0 ml./min.) c o n t r o l l e d by a magnetic v a l v e . There' was an automatic mechanism c o n t r o l l i n g t h i s v a lve which c o n t r o l l e d the f l o w r a t e through the system to give a reprod- u c i b l e t e s t . The opening and c l o s i n g of the valve was c o n t r o l l e d by an e l e c t r o n i c a l l y d r i v e n mechanical, device. The s t i r r e r was power d r i v e n . A screen around the p r o p e l l e r p r o t e c t e d the t e s t ' p r e p a r a t i o n from coming i n contact w i t h the blades. The s t i r r e r was moving f a s t enough so that the drug was moving f r e e l y and d i d not s t i c k to the sides or the bottom of the d i s s o l u t i o n v e s s e l . Concentration of the medium was determined e i t h e r by a n a l y s i n g a f r a c t i o n of the c o l l e c t e d medium or by a continuous r e c o r d i n g on a spectro- photometer. 22 I n t h e i r study of the e f f e c t of compression pressure on d i s s o l u t i o n , Ganderton and h i s co-workers (7) used two ° methods to t e s t compressed t a b l e t s of phenindione of d i f f e r - ent f o r m u l a t i o n s , a l l c o n t a i n i n g 100 mg. c r y s t a l l i n e l a c t o s e w i t h 15 mg. of. potato s t a r c h dispersed w i t h i n the granules, over a pressure range of 60 - 2500 Kg.cm. on a s i n g l e punch machine. I n the f i r s t method, the d i s s o l u t i o n v e s s e l was a 2 - l i t r e beaker ( c o n t a i n i n g 1.5 l i t r e s of d i s s o l u t i o n medium) immersed i n a water bath. The medium was s t i r r e d by a perspex paddle 11 cm. i n diameter, held 0.5 cm. above the bottom of the v e s s e l and r o t a t i n g at 56 r.p.m. The two blades of the paddle were 2.5 cm. deep and p i t c h e d at 45° to promote a x i a l mixing. Two d i a m e t r i c a l l y opposed b a f f l e s 1.3 cm. wide were f i x e d i n the v e s s e l . The t e s t t a b l e t was placed i n a cube of lOOsmesh s t a i n l e s s s t e e l gauze of sides 1.5 cm., r i g i d l y suspended i n the bath 4 cm. from the paddle a x i s and 2 cm. below the surface of the l i q u i d . The t e s t was performed i n 0.001 N.NaOH at 37°C F i v e ml. samples were withdrawn through a f i l t e r tube at s u i t a b l e i n t e r v a l s - over a p e r i o d of one hour, d i l u t e d and assayed. The second method was a continuous d i s s o l u t i o n process, using a c y l i n - d r i c a l perspex c e l l 5.1 cm. i n diameter. A 100-mesh, concave, s t a i n l e s s s t e e l gauze was f i x e d across the c e l l and the t a b l e t h e l d l i g h t l y at the centre w i t h a v e r t i c a l p i n . Water bu f f e r e d at pH 7 was admitted through the centre of the c e l l 23 base and d i r e c t e d r a d i a l l y on the gauze below the t a b l e t . When the t a b l e t was wetted, the r e t a i n i n g p i n was removed and the l i q u i d i s s u i n g from the top of the c e l l was c o l l e c t e d o and assayed. The t e s t was c a r r i e d out at pH 7 and 20 G to a l l o w the d i r e c t assay of the emerging s o l u t i o n without f u r t h e r d i l u t i o n . The f i r s t l i t r e of s o l u t i o n was c o l l e c t e d and assayed. The t e s t l a s t e d approximately eleven minutes, g i v i n g a mean l i q u i d v e l o c i t y i n the c e l l of 0.075 cm./sec. D i s i n t e g r a t i o n t e s t s were c a r r i e d out usi n g the method described i n the B r i t i s h Pharmacopeia 1963. From the r e s u l t s , i t was found that the speed of d i s i n t e g r a t i o n p r o g r e s s i v e l y decreased as the compression pressure increased. The d i s s o l u t i o n r a t e s f e l l s t e e p l y at the low compression pressures and then r i s e to form a peak at pressures which v a r i e d w i t h f o r m u l a t i o n from 500 to 800 Kg. cm. . The rat e then decreased to give an extended high pressure r e g i o n i n which d i s s o l u t i o n r a t e was independ- ent of both pressure and fo r m u l a t i o n . Although very weak and e a s i l y penetrated by the d i s s o l u t i o n medium, t a b l e t s produced at very low pressures d i d not break up e x t e n s i v e l y during the t e s t . L i t t l e fragmentation had occurred so that p a r t i c l e l o s s and d i s s o l u t i o n r a t e were low. At higher pressures, p e n e t r a t i o n s t i l l occurred q u i c k l y and s t r e s s r e l e a s e and the l o s s of small a i r bubbles caused much more d i s r u p t i o n . The van der Waals forces h o l d i n g the t a b l e t together i n the dry s t a t e were i n e f f e c t u a l i n the presence of a l i q u i d of high d i e l e c t r i c constant and p e n e t r a t i o n by the d i s s o l u t i o n medium caused the t a b l e t t o break up. With f u r t h e r increase i n pressure, rebonding of the m a t e r i a l occurred and a stronger and denser t a b l e t was formed, which was l e s s e a s i l y penetrated. P a r t i c l e l o s s and d i s s o l u t i o n r a t e were, t h e r e f o r e , depressed. U l t i m a t e l y , high s t r e n g t h and low p e n e t r a t i o n prevented break-up of the t a b l e t , and d i s s o l u t i o n occurred only from the surface of the t a b l e t and was t h e r f o r e independent of any fo r m u l a t i o n v a r i a b l e s . At low pressure, the d i s s o l u t i o n r a t e of t a b l e t s increased as the f i l l e r s i z e or the granule s i z e decreased. These r e s u l t s i n d i c a t e d t h a t these f a c t o r s g r e a t l y a f f e c t the s i z e of the p a r t i c l e s l i b e r a t e d by p e n e t r a t i o n and break-up. I n the case of the f i l l e r , decrease i n s i z e increased the homogeneity of the o r i g i n a l mix, opposing the formation of la r g e agglomerates of phenindione. Decrease i n granule s i z e modified the d i s p o s i t i o n of the e x t e r n a l s t a r c h , a f a c t o r which has been shown to g r e a t l y a f f e c t p e n e t r a t i o n and break-up of the t a b l e t . Smaller granules would a l l o w i t s more e f f e c t i v e d i s t r i b u t i o n as a h y d r o p h i l i c or antibonding l a y e r . Both these e f f e c t s w i l l disappear at high pressures of compaction when break-up i s depressed and s o l u t i o n occurs only from the surface of the t a b l e t . 2 A continuous f l o w system was designed by Woo ( 33 ) to study eleven brands of commericially a v a i l a b l e t o l b u t - amide t a b l e t s . The d i s s o l u t i o n v e s s e l was a t w o - l i t r e a s p i r a t o r b o t t l e . The rubber stopper at the bottom o u t l e t of the b o t t l e c a r r i e d a short piece of gl a s s t u bing that was connected on the i n s i d e of the d i s s o l u t i o n v e s s e l to a piece of rubber tu b i n g . A piece of very f i n e gauze that covered the open end of the rubber tubing acted as the f i l t e r f o r the outflowing medium. The g l a s s tube that proje ed out of the v e s s e l was connected to a T-glass tube, one arm of which was j o i n e d to an a i r supply, while the other arm l e d to a 500-ml. s u c t i o n f l a s k v i a rubber tu b i n g . The rubber stopper at the top of the d i s s o l u t i o n v e s s e l c a r r i e d a thermometer and an i n l e t tube from a r e s e r v o i r of si m u l - ated g a s t r i c f l u i d . The d i s s o l u t i o n v e s s e l was set on a pyro-magnetstir, that kept the medium i n the v e s s e l at a temperature of 37°G + 0.5°C, and caused the magnetic s t i r r e r i n the d i s s o l u t i o n v e s s e l to s t i r the medium at va r y i n g speeds. The d i s s o l u t i o n v e s s e l was f i l l e d w i t h one and a h a l f l i t r e s of simulated g a s t r i c f l u i d . A f t e r the o medium had e q u i l i b r a t e d to 37 C, a t e s t t a b l e t o f . t o l b u t - amide ( 500 mg.) was dropped i n t o the d i s s o l u t i o n v e s s e l . The s t i r r e r was turned on to give a vigorous s t i r r i n g i n t e n s i t y . A f t e r an i n i t i a l ten minutes, the clamp on the 2 6 tube to the s u c t i o n f l a s k was p a r t i a l l y unscrewed to a l l o w a f l o w r a t e of 500 ml./30 minutes, when s u c t i o n was a p p l i e d . At the same time, the i n f l o w r a t e from the r e s e r v o i r was adjusted to d e l i v e r 500 ml./30 minutes, by p a r t i a l l y unscrew- i n g the clamp on the d e l i v e r y tube. This ensured that the volume, i n the d i s s o l u t i o n v e s s e l was kept constant. During the changing of the s u c t i o n f l a s k s ( a f t e r each t h i r t y - m i n u t e i n t e r v a l ) both the i n f l o w and outflow tubes were clamped o f f by a second clamp. As the f l u i d flowed through the f i n e gauze f i l t e r , p a r t i c l e s of the t e s t t a b l e t c o l l e c t e d on the f i l t e r and caused i t to be clogged up, thus slowing the outflow r a t e . This was overcome by t u r n i n g on the a i r supply o c c a s s i o n a l l y to blow the p a r t i c l e s o f f the gauze back i n t o the d i s s o l u t i o n medium. The d i s s o l u t i o n t e s t was c a r r i e d on f o r a p e r i o d of three hours, c o l l e c t i n g i n t o t a l a volume of three l i t r e s . I n d i v i d u a l analyses were c a r r i e d out on each p o r t i o n of d i s s o l u t i o n medium c o l l e c t e d . D i s i n t e g r a t i o n t e s t s were c a r r i e d out on these tolbutamide t a b l e t s using the o f f i c i a l U.S.P. d i s i n t e g r a t i o n apparatus (30). The d i s i n t e g r a t i o n s t u d i e s were c a r r i e d out without the use of the p l a s t i c d i s k s . The v a r i o u s brands of tolbutamide t a b l e t s seemed to f o l l o w a s i m i l a r p a t t e r n of d i s s o l u t i o n . In the i n i t i a l ten minutes, most of the t a b l e t s broke up i n t o very f i n e p a r t i c l e s or f l a k e s . The rel e a s e of the a c t i v e i n g r e d i e n t i n c r e a s e d g r a d u a l l y t i l l a maximum was reached i n the f i r s t h a l f hour. The l e v e l remained constant f o r about an hour and then g r a d u a l l y decreased due to a d i l u t i o n process by the incoming s o l v e n t , thus g i v i n g a p l a t e a u - l i k e curve. Even though the d i s s o l u t i o n p a t t e r n was s i m i l a r f o r the d i f f e r e n t brands, the total.amount of the a c t i v e i n g r e d i e n t , r e l e a s e d i n three l i t r e s of s o l u t i o n ranged from 60 mg. to 270 mg. No c o r r e l a t i o n w i t h the d i s i n t e g r a t i o n times could be found, confirming an e a r l i e r report by Brudney and h i s co-workers (1). However, the d i s s o l u t i o n t e s t was c a r r i e d out on only one t a b l e t of each product, so d e f i n i t e conclusions about the t a b l e t q u a l i t y could not be made. The d i s s o l u t i o n t e s t described was very time consuming and r e q u i r e d constant s u p e r v i s i o n . Moreover, the f i l t e r i n g system was not too e f f i c i e n t and tend to blog up towards the end of the d i s s o l u t i o n run and hence would be inadequate f o r longer d i s s o l u t i o n runs. 28 I I I . THE "IN VITRO" CHARACTERISTICS OF PHENYLBUTAZONE The drug being i n v e s t i g a t e d i s phenylbutazone and i s used f o r the r e l i e f of j o i n t pain caused by rheumatoid a r t h r i t i s , gout, b u r s i t i s and other r e l a t e d d i s o r d e r s . Phenylbutazone ( 1 , 2 - D i p h e n y l - 4 - b u t y l - 3 , 4 P y r a z o l i d i n e d i o n e ) , a weak organic a c i d of pKa 4 . 4 , i s very s l i g h t l y s o l u b l e i n water ( l e s s than 0 . 7 mg./ml. at 25°C) but f r e e l y s o l u b l e i n a l c o h o l (50 mg./ml.), acetone, ether and e t h y l a c e t a t e . Phenylbutazone powder tends to agglomerate and f l o a t on the surface of an aqueous medium. Attempts at r e c r y s t a l l i z a t i o n y i e l d e d long needles that f l o a t on the surface of aqueous s o l u t i o n s . Hence, B u t a z o l i d i n (Geigy) t a b l e t s (which have been reported to give good c l i n i c a l response ( 2 , 27)) were chosen as a standard f o r t h i s i n v e s t i g a t i o n . (a) The S o l u b i l i t y of Phenylbutazone i n B u f f e r e d S o l u t i o n s I t i s necessary, during the d i s s o l u t i o n t e s t , to change from an a c i d i c medium of pH 1.2 t o a pH 6 . 2 . Since the s o l u b i l i t y of phenylbutazone i s a f f e c t e d by the p.H of the d i s s o l v i n g medium, s o l u b i l i t y s t u d i e s were c a r r i e d out i n b u f f e r s of d i f f e r e n t pH valu e s . Procedure - Using the Smith, K l i n e & French d i s i n t e g r a t i o n apparatus ( 2 9 ) , phenylbutazone powder i s put i n t o each of s i x ' b o t t l e s c o n t a i n i n g 50 ml. of b u f f e r (two samples of each b u f f e r are used). The b o t t l e s are suspended i n a water bath maintained at 37°C and r o t a t e d so as to provide good mixing f o r a per i o d of twenty-four hours. At the end .of t h i s p e r i o d , the s o l u t i o n s are f i l t e r e d q u i c k l y , d i l u t e d i f necessary w i t h the b u f f e r and assayed spectro- p h o t o m e t r i c a l l y . R e s u l t s - The s o l u b i l i t y curve (Figure 1 ) , drawn from the data obtained, showed that the s o l u b i l i t y of phenylbutazone i s very low i n b u f f e r s of low pH value s . However, once the pH value reached 6, the s o l u b i l i t y i ncreased g r e a t l y . (b) Determination of the I s o s b e s t i c P o i n t The i s o s b e s t i c point of a s o l u t i o n i s the wave- le n g t h at which changes i n the pH of the s o l u t i o n do not a f f e c t the absorbancy (As) reading of the s o l u t i o n . Since the c o n d i t i o n s of t h i s d i s s o l u t i o n t e s t i n v o l v e a change from an a c i d i c to a basic medium, the wavelength at which absorbancy readings are to be recorded must be at the i s o s b e s t i c p o i n t . Procedure - Weigh a c c u r a t e l y 100 mg. of phenyl- butazone powder and d i s s o l v e ^ i n 100 ml. of 95% ethanol. Ten ml. a l i q u o t s of t h i s s o l u t i o n are d i l u t e d a c c u r a t e l y to 1000 ml. w i t h b u f f e r s of va r i o u s pH values (ranging from pH 1.2 to pH 7.5)'. The spectrum of these s o l u t i o n s are recorded on a rec o r d i n g spectrophotometer (Spectronic 505, Bausch and Lomb). From the spectra of the s o l u t i o n s , 30 an i s o s b e s t i c p o i n t was observed at between 238 mu and 245 mu. The absorbancy of these s o l u t i o n s are then read on a Beckman DU spectrophotometer from 238 mu to 245 mu w i t h each s o l u t i o n blanked against the r e s p e c t i v e b u f f e r . A p l o t of the absorbancy readings f o r the s o l u t i o n s showed t h a t the wavelength that i s c l o s e s t to being the i s o s b e s t i c - point f o r phenylbutazone i s 240 mu (Figure 2 ) . This point was chosen as the wavelength at which a l l subsequent absorbancy readings were recorded. (c) Determination of the A b s o r p t i v i t y Value The a b s o r p t i v i t y ( a s ) i s a constant f o r a p a r t i c u l a r s o l u t e i n a p a r t i c u l a r solvent at a c e r t a i n wavelength. According to Beer's Law, i t i s the slope of the l i n e r e l a t i n g the absorbancy of a s o l u t i o n and i t s c o n c e n t r a t i o n . As = a sbc where As i s the absorbancy of the s o l u t i o n a s i s the a b s o r p t i v i t y b i s the c e l l l e n g t h i n cm. c i s the concen t r a t i o n i n gm./L. Procedure -.Weigh a c c u r a t e l y 100 mg. phenylbutazone poxtfder and d i s s o l v e i n 100 ml. of 95% ethanol. D i l u t e a l i q u o t s of f i v e to twenty ml. to 1000 ml. w i t h Simulated I n t e s t i n a l F l u i d U.S.P. The absorbancy of each s o l u t i o n i s read on a Beckman DU spectrophotometer at 240 mu, using Simulated I n t e s t i n a l F l u i d U.S.P. as a blank. 31 R e s u l t s - The absorbancy readings were p l o t t e d and a s t r a i g h t l i n e was obtained. The a b s o r p t i v i t y was c a l c u l a t e d from the slope of t h i s l i n e and found to be 41.5 (Figure 3 ) . 3 2 2 4 0 2 4 3 2 4 6 0 Wavelength (mu) Figure 2 . The Isosbestic Point of Phenylbutazone 34 0.80 0.60 % 0.40 0.20 As= a sbc a q - As = 41.5 5 10 15 20 Concentration (mg./L.) F i g u r e 3. C a l i b r a t i o n Curve f o r Phenylbutazone i n Simulated I n t e s t i n a l F l u i d at 240 mu 35 IV. THE OPERATING CHARACTERISTICS OF A CONTINUOUS FLOW DISSOLUTION APPARATUS (a) D e s c r i p t i o n of the Apparatus The d i s s o l u t i o n v e s s e l (a o n e - l i t r e three-neck round-bottom f l a s k ) i s immersed i n a constant temperature o o water bath at 37 C t 0 . 5 C. Dipping i n t o the c e n t r a l neck of the f l a s k i s the shaft of a F i s h e r Stedi-Speed adj u s t a b l e s t i r r e r (Model 12) with a four-blade i m p e l l e r blade (3 cm. i n diameter) and a c y l i n d r i c a l wire basket at i t s bottom. The basket which i s A cm. long and 2 . 5 cm. i n diameter i s made from 10-mesh s t a i n l e s s s t e e l wire c l o t h . A g l a s s tube which a c t s as the i n l e t tube f o r the d i s s o l u t i o n medium dips through the r i g h t side neck i n t o the d i s s o l u t i o n v e s s e l to a depth of 10 cm. Connected to t h i s tube i s a two-way stop cork, one arm of which i s connected to a r e s e r v o i r of simulated g a s t r i c j u i c e (2 gm. NaCl + 7 ml. HCl i n 1000 ml. d i s t i l l e d water, pH 1 .2) w h i l e the other arm i s j o i n e d to a r e s e r v o i r of simulated i n t e s t i n a l j u i c e ( 6 . 8 gm. KH 2 P0^ + 1 .52 gm. NaOH i n 1000 ml. d i s t i l l e d water, pH 7 . 5 ). Both r e s e r v o i r s of d i s s o l u t i o n medium are placed on heaters to keep the s o l u t i o n at 37°C 0 . 5 ° C An i n v e r t e d s i n t e r e d g l a s s funnel (30 ml. capacity) of coarse p o r o s i t y dips i n t o the l e f t side arm of the d i s s o l - u t i o n v e s s e l . I t i s j o i n e d to a short l e n g t h of g l a s s tube c a r r y i n g a s i n t e r e d t i p (coarse p o r o s i t y ) , that dips i n t o the d i s s o l u t i o n v e s s e l to a depth of '5.0 cm. A combination g l a s s e l e c t r o d e (not i l l u s t r a t e d i n F i g u r e 4) l e a d i n g from a reco r d i n g potentiometer (Potent- iograph E336A) and a short piece of g l a s s tube, which ac t s as the o u t l e t tube, dip i n t o the broad end of the s i n t e r e d g l a s s f u n n e l . The o u t l e t tube i s j o i n e d by Tygon tubing to an a d j u s t a b l e pump (Cole Palmer A-769) from which tubing leads to a fl o w c e l l (1 cm. i n length) i n a rec o r d i n g spectro- photometer (Spectronic 505, Bausch & Lomb). A recorder ( V a r i c o r d Model 43) i s attached to the spectrophotometer f o r a d i r e c t r e c o r d i n g of the concentration of the s o l u t i o n passing through the flow c e l l . From the fl o w c e l l , the s o l u t i o n i s d e l i v e r e d i n t o a t w e l v e - l i t r e covered g l a s s v e s s e l , that a c t s as the c o l l e c t i n g v e s s e l . (b) C a l i b r a t i o n of the Recorder Procedure - The recorder i s set on the 25 and mV range. I t i s zeroed w i t h g a s t r i c j u i c e i n the sample c e l l and a i r as the blank. S o l u t i o n s of known concent r a t i o n of phenyl- butazone are read i n the sample c e l l of the spectrophotometer at 240 mu and the chart readings on the recorder s c a l e are recorded. From the known concentrations and the chart readings on the recorder s c a l e , a graph i s p l o t t e d (Figure 5 ) . gure 4. Diagram of Continuous Flow D i s s o l u t i o n Apparatus ISPECTRONIC 5Q5 • t DISCHARGE COLE PALMER A - 7 6 9 ADOUSTABLE PUMP t FILTERING DEVICE FOR CONTINUOUS FLOW SINTERED GLASS FUNNEL (COARSE 30 mi.) SINTERED TIP (COARSE) VARICORD MODEL FISHER STEDI* SPEED STIRRER TO STIRRING SUCTION / SHAFT T £ S T F U J | D x GLASS TUBE TWO WAY STOPCOCK TEST FLUID H GLASS TUBE ONE LITER FLASK BASKET 38 150 to •H Ti crj CD ca •p u crj X ! o 100 10 20 Concentration (mg. phenylbutazone/L^) F i g u r e 5. C a l i b r a t i o n Curve f o r Recorder E x t e r n a l to the Spectronic 505 (c) The E f f e c t of S t i r r i n g Rate on D i s s o l u t i o n The s t i r r i n g r a t e has been reported by many researchers (8, 11, 17) to have a considerable e f f e c t on the d i s s o l u t i o n r a t e . Levy (11) has s t a t e d that too i n t e n s i v e an a g i t a t i o n r a t e i s to be avoided since X-ray photographs have shown tha t t a b l e t p a r t i c l e s tend to remain as an aggregate on the stomach surface. This i m p l i e s that the mixing process i n the stomach i s very low. However, i n the small i n t e s t i n e , the p a r t i c l e s appeared to be very w e l l dispersed i n d i c a t i n g that the mixing i n t h i s r e g i o n i s quite vigorous. Since phenylbutazone i s n e a r l y i n s o l u b l e i n s o l u - t i o n s of pH 1.2 but very s o l u b l e i n s o l u t i o n s of pH 7.5, i t would appear that the absorption of phenylbutazone occurs mainly under the b a s i c c o n d i t i o n s of the small i n t e s t i n e . Hence, a s t i r r i n g r a t e that causes the drug p a r t i c l e s to be w e l l dispersed throughout the medium may be d e s i r a b l e i n the d i s s o l u t i o n t e s t under study. Consequently, s t i r r i n g r a t e s of 65, 100 and 115 r.p.m. i n the forward d i r e c t i o n were i n v e s t i g a t e d using the designed d i s s o l u t i o n apparatus and t e s t procedure. These s t i r r i n g r a t e s were i n v e s t i g a t e d w i t h the s t i r r e r at d i f f e r - ent depths i n the d i s s o l u t i o n medium, using B u t a z o l i d i n t a b l e t s as the t e s t p r e p a r a t i o n , and at a constant pumping ra t e of 60 ml./minute. Each r a t e was t e s t e d i n d u p l i c a t e . 40 R e s u l t s - The data obtained from the d i s s o l u t i o n runs are t a b u l a t e d i n Table I . I t can be seen th a t when the s t i r r i n g i s not enough to move the p a r t i c l e s , the r e l e a s e r a t e of the drug i s low even though the s o l u t i o n i n the d i s s o l u t i o n v e s s e l appears to be homogeneous. The low rel e a s e r a t e i s due t o the f a c t that the t a b l e t p a r t i c l e s remain as an aggregate on the d i s s o l u t i o n v e s s e l bottom, thus present- i n g a l i m i t e d surface area to the d i s s o l u t i o n medium. When the basket i s 6 Cm. from the v e s s e l bottom, the s t i r r i n g i s i n s u f f i c i e n t even at the high s t i r r i n g r a t e of 115 r.p.m. At 3.5 cm. from the bottom of the v e s s e l , the s t i r r i n g i s adequate except at the lox^est r a t e of 65 r.p.m., when about h a l f of the t a b l e t p a r t i c l e s remain aggregated on the v e s s e l bottom. The other two s t i r r i n g r a t e s caused the p a r t i c l e s to move through the medium, the higher ra t e of 115 r.p.m. g i v i n g a more vigorous movement. At a depth of 1 cm. from the v e s s e l bottom, the s t i r r i n g e f f e c t i s qui t e vigorous even at the r a t e of 65 r.p.m. To avoid t h i s s t i r r i n g e f f e c t , t h i s depth was not chosen f o r product t e s t i n g . The depth f i n a l l y chosen was 3.5 cm. w i t h a s t i r r - i n g r a t e of! 100 r.p.m. r a t h e r than 115 r.p.m. The former r a t e produced the d e s i r e d s t i r r i n g e f f e c t . At t h i s speed, the p a r t i c l e s move sl o w l y through the medium. 41 Table I . The Effect of S t i r r i n g Rate on Dissolution S t i r r i n g rate Depth of basket Mg. phenylbutazone (forward direction) from vessel bottom i n solution i n 3 hours 1 cm. .87.5 mg. 65 r.p.m. 3*5 cm. 62.1 mg. 6 cm. 55.4 mg. 1 cm. 83.1 mg. 100 r.p.m. 3.5 cm. 83.5 mg. 6 cm. 75.0 mg. 115 r.p.m. 1 cm. 3.5 cm. 6 cm. 90.0 mg. 85.0 mg. 78.5 mg. 42 (d) The E f f e c t of Pumping Rate on D i s s o l u t i o n Three pumping r a t e s were stud i e d f o r t h e i r e f f e c t on d i s s o l u t i o n r a t e . The d i s s o l u t i o n t e s t was c a r r i e d out w i t h B u t a z o l i d i n t a b l e t s as the t e s t p r e p a r a t i o n and w i t h the basket at 3.5 cm. fronr the v e s s e l bottom s t i r r i n g at a r a t e of 100 r.p.m. The pumping r a t e s s t u d i e d were 50 ml./min., 60 ml./min. and 70 ml./min. Rates lower than 50 ml./min. were not s t u d i e d due to instrum e n t a l l i m i t a t i o n s i n reading the more concentrated s o l u t i o n s . R e s u l t s - The data obtained are ta b u l a t e d i n Table I I . This data shows that there i s no s i g n i f i c a n t d i f f e r - ence i n the rel e a s e of the drug i n the three-hour p e r i o d . The pH changes i n the medium are a f f e c t e d by the pumping r a t e to a s l i g h t extent. Higher pumping r a t e s gave a much f a s t e r pH change from a c i d i c to bas i c pH. At the pumping r a t e of 60 ml./min., the sharp r i s e i n pH from about pH of 2 to pH of 6 . 2 occurred w i t h i n t w e n t y - f i v e minutes (Figure 6), whi l e at a pumping ra t e of 70 ml./min., i t occurred w i t h i n twenty minutes. However, at 70 ml./min., the f i n a l volume of solvent i s too great to be handled e a s i l y . At 50 ml./min., the pH change occurred i n t h i r t y minutes but at t h i s r a t e , the s o l u t i o n i s becoming too saturated to be read on the spectrophotometer. A pumping rat e of 60 ml'./min. was t h e r e f o r e , f i n a l l y chosen f o r the d i s s o l u t i o n t e s t . 43 Table I I . The E f f e c t of Pumping Rate on D i s s o l u t i o n Pumping r a t e Mg. of phenylbutazone i n s o l u t i o n 1 hour 2 hours 3 hours 50 ml./min. 1. 4.0 mg, 2. 3.9 mg. 56.0 mg, 51.0 mg. 86.5 mg. 86.4 mg. 60 ml./min. 1. 9.0 mg. 2. 8.3 mg. 70 ml./min. 1. 5.9 mg. 2. 6.7 mg. 68.4 mg. 62.6 mg. 62.2 mg. 63.0 mg. 89.6 mg. 86.4 mg, 86.5 mg. 88.2 mg.  45' (e) Test Procedure With the apparatus set up as described and as shown i n F igure 4, the d i s s o l u t i o n v e s s e l ( c o n t a i n i n g one l i t r e of simulated g a s t r i c j u i c e ) i s allowed to e q u i l i b r a t e i n a constant temperature water bath at 37°C + 0.5°C The e n t i r e system i s f l u s h e d out w i t h simulated g a s t r i c j u i c e to remove a i r from the system. The recorder i s zeroed w i t h simulated g a s t r i c j u i c e i n the sample flow c e l l and a i r as the blank. A s i n g l e phenylbutazone t a b l e t (100 mg.) i s put i n t o the wire basket below the i m p e l l e r blade and the whole apparatus i s set infemotion. The s t i r r i n g r a t e i s 100 r.p.m. i n a forward (clockwise) d i r e c t i o n at a depth of 3.5 cm. from the bottom of the d i s s o l u t i o n v e s s e l . With the i n l e t tube from the r e s e r v o i r of simulated g a s t r i c j u i c e opened, the pump i s turned on to d e l i v e r 60 ml./min. through the e n t i r e system. A f t e r the i n i t i a l t h i r t y minutes, simulated i n t e s t - i n a l j u i c e i s allowed i n t o the system. At t h i s p o i n t , the potentiometer i s switched on to record pH changes as the medium i s g r a d u a l l y changed from an a c i d i c medium to a basi c one. A l i q u o t s from the c o l l e c t i n g v e s s e l are taken at hourly i n t e r v a l s f o r a n a l y s i s on a Beckman DU spectrophoto- meter at 240 mu, w i t h simulated i n t e s t i n a l j u i c e as the blank. As the s o l u t i o n from the d i s s o l u t i o n v e s s e l passes 46 through the flow c e l l i n the spectrophotometer set at 240 mu, the V a r i c o r d records the c o n c e n t r a t i o n . This gives a c o n t i n - uous r e c o r d i n g of the c o n c e n t r a t i o n of the s o l u t i o n as d i s s o l u t i o n occurs. The d i s s o l u t i o n run i s c a r r i e d on f o r three to s i x hours depending on the d i s s o l u t i o n rate ! of the i n d i v i - dual products. At the end of each run, the e n t i r e system i s f l u s h e d out w i t h 95% ethanol and water. Using t h i s t e s t procedure, seven brands of commer- i c i a l l y a v a i l a b l e phenylbutazone t a b l e t s (100 mg.) were t e s t e d i n t r i p l i c a t e . Four of the products are sugar-coated t a b l e t s (Products A, E, W, X) while the other three are e n t e r i c coated t a b l e t s (Products AA, CC, DD^). Each product was t e s t e d i n t r i p l i c a t e . (f) R e s u l t s An average continuous c o n c e n t r a t i o n p l o t was obtained f o r each product from the curves drawn out on the recorder. This p l o t gave the d i s s o l u t i o n p r o f i l e of the product and a l s o allows the c a l c u l a t i o n of the t o t a l amount of drug r e l e a s e d i n any p e r i o d of time (Figures 7 a , b, c ) . The h o u r l y a n a l y s i s on the Beckman DU spectrophotometer a l s o gave the amount of drug rel e a s e d at each hourly i n t e r v a l , thus p r o v i d i n g a check on the amount r e g i s t e r e d by the recorder (Table I I I ) . From the data on the products obtained by the two 4 7 L8 LO I-H e o •H •P cd U •P c © o o o 20 CC 1 2 Time (hours) Figure 7b. D i s s o l u t i o n P r o f i l e s f o r Two Brands of Phenylbutazone Tablets  50 Table I I I . D i s s o l u t i o n Data f o r Seven Brands of Phenylbutazone Tablets Mg. of phenylbutazone i n s o l u t i o n Product 1 hour 2 hours 3 hours 1. 5 . 2 mg. 6 2 . 5 mg. 8 6 . 5 mg. A 2 . 5 . 2 mg. 6 1 . 9 mg. 8 8 . 0 mg. 3 . 5 . 2 mg. 6 5 . 9 mg. 9 3 . 5 mg. 1. 1 .2 mg. 5 2 . 5 mg. 8 6 . 5 mg. E 2 . 1 . 8 mg. 5 0 . 4 mg. 81 . 8 mg. 3 . 1.5 mg. 4 8 . 0 mg. 7 9 . 2 mg. 1. 0 . 8 mg. 4 3 . 2 mg. 8 5 . 3 mg. AA 2 . 0 . 8 mg. 4 3 . 2 mg. 8 6 . 5 mg. 3 . 1 .2 mg. 51-5 mg. 8 9 . 6 mg. 1. 1 .4 mg. 9 1 . 4 mg. 9 6 . 2 mg. CC 2 . 1 .7 mg. 9 5 . 7 mg. 1 0 0 . 0 mg. 3 . 1 .7 mg. 9 0 . 7 mg. 9 5 . 8 mg. 51 Table I I I (Continued) Mg. of phenylbutazone i n s o l u t i o n Product 3 hours 4 hours 5 hours 6 hours 1. 7 . 4 mg. 2 3 . 6 mg. 5 5 . 8 mg. 7 5 . 7 mg. X 2 . 1 6 . 2 mg. 5 5 . 3 mg. 7 8 . 7 mg. 9 3 . 0 mg. 3 . 2 3 . 8 mg. 5 0 . 8 mg. 7 5 . 0 mg. 9 2 . 2 mg. 1. 3 6 . 0 mg. 5 5 . 2 mg. 70 . 6 mg. 8 6 . 3 mg. W 2 . 3 6 . 2 mg. 5 5 . 0 mg. 70 .3 mg. 8 6 . 0 mg. 3 . 3 6 . 3 mg. 5 5 . 3 mg. 70 . 5 mg. 8 6 . 2 mg. 1. 5 . 4 mg. 2 7 . 0 mg. 9 4 . 0 mg. 101 .5 mg. DDX 2 . 4 . 5 mg. 2 2 . 5 mg. 9 1 . 7 mg. 9 9 . 0 mg. 3 . 4 . 5 mg. 2 2 . 3 mg. 9 0 . 9 mg. 9 3 . 0 mg. 52 Table IV. T 5 0 % a n d T 9 0 % Values f o r Seven Phenylbutazone Tablets Brands of Product T $ 0 $ value T 9 0 % value A 100 minutes 180 minutes E 120 minutes 215 minutes W 225 minutes 735 minutes. X 255 minutes 735 minutes AA 125 minutes 185 minutes CC 80 minutes 120 minutes DD-L 225 minutes 300 minutes 53 methods, time f o r 5 0 $ of the drug (T5Q$) and 9 0 $ of the drug (T^ 0^) to go i n t o s o l u t i o n was c a l c u l a t e d and t a b u l a t e d i n Table IV. The four brands of sugar-coated t a b l e t s showed l i t t l e d i s s o l u t i o n i n simulated g a s t r i c f l u i d . However, once the medium reached a pH of 6.2, the co n c e n t r a t i o n of the a c t i v e i n g r e d i e n t increased g r e a t l y . This change i n pH occurred i n twenty-five minutes at the pumping rat e of 60 ml./minute. Products A and E (which d i s i n t e g r a t e d w i t h i n f i f t e e n minutes of the s t a r t of the experiment) showed s i m i l a r d i s s o l u t i o n p a t t e r n s . The c o n c e n t r a t i o n of a c t i v e i n g r e d i e n t continued to increase g r a d u a l l y once the medium reached a pH of 6. A peak was reached i n about one and a h a l f hours. The curve then g r a d u a l l y d e c l i n e d due to a d i l u t i o n process by the incoming s o l v e n t . Product E rele a s e d s l i g h t l y l e s s a c t i v e i n g r e d i e n t from the t a b l e t than d i d Product A. The other two sugar-coated products showed d i f f e r - ent d i s s o l u t i o n p r o f i l e s due to the l a c k of d i s i n t e g r a t i o n . Product W d i d not d i s i n t e g r a t e at a l l even at the end of s i x hours. D i s s o l u t i o n by t h i s product was very low and stayed at the same l e v e l throughout the e n t i r e d i s s o l u t i o n run. The a c t i v e i n g r e d i e n t seemed to d i s s o l v e out from the surface of the t a b l e t , which was smaller but s t i l l i n t a c t 5k at the end of the d i s s o l u t i o n run. Product W showed very l i t t l e • t a b l e t v a r i a t i o n . Product X was found to vary from one t a b l e t to another i n the same l o t . A f t e r t h e . t a b l e t c o a t i n g came o f f a f t e r three hours, l a r g e p a r t i c l e s could be'seen coming o f f and,,at t h i s p o i n t , the con c e n t r a t i o n of the a c t i v e i n g r e d i e n t r e g i s t e r e d on the recorder was seen to increase suddenly but not g r e a t l y . The concen t r a t i o n l e v e l remained approximately the same f o r about an hour, then decreased very s l i g h t l y f o r the r e s t of the time. The d i s s o l u t i o n rate of Products ¥ and X was so low t h a t the d i s s o l u t i o n - r u n had to be c a r r i e d on f o r s i x hours i n s t e a d of three hours. Products A and E re l e a s e d about 90% of the a c t i v e i n g r e d i e n t i n three hours while i t took Products W and X s i x hours to r e l e a s e the same amount of a c t i v e i n g r e d i e n t . The three e n t e r i c coated Products AA, CC and DD-̂ remained i n t a c t t i l l the d i s s o l u t i o n medium reached a pH of 7. Products AA and CC broke up very q u i c k l y i n t o small p a r t i c l e s . Product DD^ d i d not s t a r t to d i s i n t e g r a t e u n t i l about three and a h a l f hours a f t e r the s t a r t of the d i s s o l u - t i o n run. Product CC gave a much f a s t e r d i s s o l u t i o n r a t e than Product AA, wi t h a' concentration peak much higher than those of the other t e s t products and, even e a r l i e r than the sugar-coated products. This was probably due to i t s 55 almost immediate d i s i n t e g r a t i o n i n the a l k a l i n e medium i n t o very f i n e p a r t i c l e s . The d i s s o l u t i o n p r o f i l e f o r Product AA was s i m i l a r to those f o r the sugar-coated Products A and E , except f o r a delay of about twenty-five minutes i n i t s peak due to the e n t e r i c c o a t i n g . The f a l l i n the d i s s o l u t i o n curve of Product CC was very sharp while that of Product AA was much more gradual. Product DD]_ released i t s a c t i v e ingred- i e n t r a p i d l y a f t e r d i s i n t e g r a t i o n occurred, g i v i n g a peak second only to Product CC. Henee, i t would appear that the d i s i n t e g r a t i o n time of the t a b l e t had an e f f e c t on the d i s s o l u t i o n of the t a b l e t when the d i s i n t e g r a t i o n time was e x c e s s i v e l y l o n g . A T^Q^ value of 120 minutes would seem to be a reasonable l i m i t of acceptance of the products th a t could be s a i d to be e f f e c t i v e " i n v i v o " . This i m p l i e s that only f o u r of the products t e s t e d , A, E, AA and CC are acceptable. This " i n v i t r o " e s t i m a t i o n can be supported by the " i n v i v o " data that showed the three Prdducts E , AA and CC were at l e a s t 75$ as e f f e c t i v e as Product A (a product t e s t e d and found to give good c l i n i c a l response). Product. AA would be a b o r d e r - l i n e case by both the " i n v i v o " and the " i n v i t r o " r e s u l t s . 56 V. THE "IN VIVO" CHARACTERISTICS OF PHENYLBUTAZONE TABLETS Burns and h i s co-workers (2) claimed that due t o the a f f i n i t y of phenylbutazone f o r plasma p r o t e i n s as compared t o t i s s u e p r o t e i n s , a great p o r t i o n of administered phenylbutazone i s found i n the plasma. They reported that peak plasma l e v e l s were reached i n about two hours a f t e r o r a l a d m i n i s t r a t i o n , i n d i c a t i n g that the drug was r a p i d l y absorbed from the g a s t r o i n t e s t i n a l t r a c t . Plasma l e v e l s d i d not increase p r o p o r t i o n a t e l y w i t h i n c r e a s i n g doses of the drug but tend to. reach a l i m i t i n g c o n c e n t r a t i o n , which v a r i e d c o n s i d e r a b l y among s u b j e c t s . On repeated d a i l y dosage, the drug.accumulated i n the body w i t h a progressive increase i n the plasma co n c e n t r a t i o n u n t i l a pla t e a u was reached on the t h i r d or f o u r t h day. In a comparative study of com m e r i c i a l l y a v a i l a b l e brands of phenylbutazone t a b l e t s , S e a r l and Pernarowski (26) determined serum l e v e l s a f t e r a d m i n i s t r a t i o n of the products; to human s u b j e c t s . The seven products were administered to nine subjects as shown i n Table V. Each subject was given two t a b l e t s , that i s approximately 200 mg. of phenylbutazone. I n general, the product was given to the subject s h o r t l y a f t e r b r e a k f a s t . Three t o f i v e samples of blood were taken over a 43-hour p e r i o d but only the r e s u l t s to 30 hours are shown. 57 Determination of Phenylbutazone i n Serum F i f t e e n ml. of blood are withdrawn by means of a dry s t e r i l e syringe and needle from a l a r g e v e i n i n the bend of the elbow. The blood i s allowed to c l o t , then c e n t r i f u g e d and the serum i s removed. Two ml. of serum are t r a n s f e r r e d to a 5 0-ml. glass-stoppered c e n t r i f u g e tube. F i v e ml. of 3 N.HC1- s o l u t i o n and 2 0 . 0 ml. of heptane are added and the tube i s shaken f o r t h i r t y minutes i n an auto- matic shaker and c e n t r i f u g e d at 1 2 0 0 r.p.m. f o r f i v e minutes. F i f t e e n , m l . of heptane are withdrawn and t r a n s f e r r e d to a 2.00-ml. b o t t l e . F i v e ml. of 2 . 5 N.NaOH s o l u t i o n are added and s w i r l e d g e n t l y . The s o l u t i o n i s then shaken f o r f i v e minutes and poured i n t o a 5 0-ml. c e n t r i f u g e tube and ce n t r i f u g e d at 1 2 0 0 r.p.m. f o r three minutes. The absorbancy of the sodium hydroxide s o l u t i o n i s read at 2 6 5 mu. The a b s o r p t i v i t y of phenylbutazone at 2 6 5 mu i n 2 . 5 N.NaOH i s 6 5 . 2 . I t may be necessary to d i l u t e the sodium hydroxide s o l u t i o n or to decrease the amount of serum taken f o r a n a l y s i s i f the concent r a t i o n of phenylbutazone i n the serum exceeds 3 0 mg. per l i t r e . A blood sample taken before the a d m i n i s t r a t i o n of the drug serves as a c o n t r o l or "blood-blank". This sample when t r e a t e d as described above, absorbs some u l t r a v i o l e t r a d i a n t energy at 2 6 5 mu. Consequently, a l l subsequent assay values must be adjusted to compensate f o r t h i s "blood-blank". 58 R e s u l t s Peak plasma l e v e l s were obtained i n f i v e r a t h e r than i n two hours as had been reported by Burns and h i s co-workers ( 2 ) . A t y p i c a l blood curve i s shown i n Figure 8. A f t e r the peak r e g i o n , the serum drug l e v e l decreased q u i t e g r a d u a l l y . The areas under the blood curves were determined by a planimeter and are t a b u l a t e d i n Table V. Using Product A as a chosen standard, only three other products, E, AA and CC appeared to be at l e a s t 75% as e f f e c t i v e as Product A, and thus considered acceptable. Since data f o r subjects 3 - 9 was incomplete, i t was not p o s s i b l e t o use i t f o r a comparison. However, a v a i l a b l e data from these subjects tend to i n d i c a t e that response to the products f o l l o w the same trend as i n subjects 1 and 2. 10 20 Time (hours) Figure 8. Concentration of Phenylbutazone i n Serum a f t e r A d m i n i s t r a t i o n u! of Product A to Subject 1 60 Table V. Areas under Blood Curves f o r Seven Brands of Phenylbutazone Ta b l e t s ^sProduct Sub j e c t N v A E W X AA CC DD]_ 1 18.57 14.8 1.82 13.3 12.42 15.33 6.55 2 20.55 15.2 4.61 7.0 15.51 22.84 11.21 3 24.1 26.4 8.9 17.0 — — — 4 — — — — 8.62 — — 5 — — — — 10.48 — — 6 — — — — 23.34 — 7 — — — — • — 11.48 — 8 — — — — — — 3.79 9 — . — — — • — — 4.65 61 V I « "IN VIVO" - "IN VITRO" CORRELATION "In v i t r o " r e s u l t s only serve to reduce the number of samples considered s u i t a b l e f o r " i n v i v o " t e s t i n g . However, they are s i g n i f i c a n t when they can be q u a n t i t a t i v e l y c o r r e l a t e d w i t h " i n v i v o " data. Even though the " i n v i v o " data a v a i l a b l e i s not complete f o r a l l t e s t products, an attempt was made to c o r r e l a t e t h i s data w i t h the T^Q^ and the T^Q^ values obtained by the " i n v i t r o " t e s t procedure. C o r r e l a t i o n was attempted only f o r the two subjects w i t h complete " i n v i v o " data, since data f o r Product A was not a v a i l a b l e f o r the other s u b j e c t s . However, the response shown by these subjects seem to f o l l o w the same trend as tha t of subjects 1 and 2. I t can be assumed, t h e r e f o r e , t h a t the same type of c o r r e l a t i o n could be obtained f o r these s u b j e c t s , t a k i n g i n t o account the p o s s i b l e d i f f e r e n c e s i n the metabolism of the drug i n these s u b j e c t s . By the use of l i n e a r r e g r e s s i o n , the l i n e of " l e a s t squares" drawn f o r each set of data f o r subjects 1 and 2 (Figures 9, 10, 11, 12). The p o i n t s f o r subject 2 seem to f i t b e t t e r on the l i n e of " l e a s t squares" drawn than those f o r subject 1. One of the reasons f o r the d e v i a t i o n s from the " l e a s t squares" l i n e could be th a t some of the " i n v i v o " blood curves were drawn from the inadequate data. Tablet v a r i a b i l i t y ( e s p e c i a l l y f o r Product X) could a l s o have c o n t r i b u t e d to the d e v i a t i o n s . 62 The l i n e s of " l e a s t squares" had negative slopes i n d i c a t i n g an in v e r s e r e l a t i o n s h i p between the " i n v i v o " and the " i n v i t r o " data. The slopes of these l i n e s ranged from -0.042, to -0.075 5 w i t h the slopes of the T^Q^ l i n e s greater than the T^Q^ l i n e s of both s u b j e c t s . The slopes of the l i n e s f o r subject 2 were greater than those of subject 1. Th i s was expected since subject 2 showed greater response to the drug than subject 1. The d i f f e r e n c e between the slopes of the' T^Q$ l i n e and the T^Q$ l i n e of subject 1 was twice t h a t of subject 2.  64  66 67 V I I . DISCUSSION The d i s s o l u t i o n procedure described h e r e i n i s completely automatic. No r o u t i n e a n a l y s i s of the medium i s r e q u i r e d , because a complete re c o r d i n g of the amount of the a c t i v e i n g r e d i e n t r e l e a s e d from the t a b l e t throughout the d i s s o l u t i o n process i s obtained on the recorder. Determination of the t o t a l area under the recorded curve w i l l give the t o t a l amount of drug r e l e a s e d . At the same time, a complete r e c o r d i n g of the pH changes i n the d i s s o l u t i o n medium can be obtained on a re c o r d i n g potentiometer. This r e c o r d i n g enables one to see how the d i s s o l u t i o n process i s a f f e c t e d by the changing pH of the d i s s o l u t i o n medium. A s i n g l e a n a l y s i s of the c o l l e c t e d medium w i l l a l s o give the t o t a l amount of drug r e l e a s e d i n a p e r i o d of time but i t w i l l not give the d i s s o l u t i o n time f o r a spec- i f i e d amount of drug t o go i n t o s o l u t i o n . F u r t h e r , t h i s f i n a l a n a l y s i s does not give the d i s s o l u t i o n p r o f i l e which can be seen to vary from product to product. Even though the apparatus set-up seems to i n v o l v e many components, the b a s i c p a r t s r e q u i r e d f o r the continuous f l o w system are the r e s e r v o i r s of d i s s o l u t i o n medium, the d i s s o l u t i o n v e s s e l , the pump and the c o l l e c t i n g v e s s e l . The re c o r d i n g potentiometer and the spectrophotometer w i t h i t s attached recorder are ac c e s s o r i e s that f u r n i s h more inform- a t i o n and avo i d manual a n a l y s i s of the medium. These 68 a c c e s s o r i e s are not necessary i f one i s i n t e r e s t e d i n the t o t a l amount of drug r e l e a s e d i n a c e r t a i n p e r i o d of time. The b a s i c p a r t s of t h i s d i s s o l u t i o n apparatus are r e l a t i v e l y easy to o b t a i n and assemble and provide a good reproducible method of checking on product q u a l i t y . The inverse r e l a t i o n s h i p e s t a b l i s h e d between the " i n v i v o " and the " i n v i t r o " data obtained by t h i s t e s t procedure tends to i n d i c a t e that t h i s procedure may p o s s i b l y be used i n the p r e d i c t i o n of the " i n v i v o " a v a i l a b i l i t y of the a c t i v e i n g r e d i e n t from the s o l i d dosage form. A T^Q% value of 120 minutes as the l i m i t of acceptance f o r the products seems reasonable on the b a s i s of both the " i n v i v o " and the " i n v i t r o " data. From the " l e a s t squares" l i n e s , i t would seem that products w i t h a T^0% value of more than f i v e hours would not be absorbed i n t o the blood stream at a l l . From both the " i n v i v o " and the " i n v i t r o " data, only f o u r of the seven products t e s t e d , namely A, E, AA and CC, can be accepted as products that w i l l be e f f e c t i v e when administered to p a t i e n t s . Products W and X give such low serum l e v e l s and long T^Q^ values t h a t one can conclude t h a t these products are u n s a t i s f a c t o r y and should not be administered to p a t i e n t s . Product DD^ w i t h s l i g h t l y higher serum l e v e l s i s s t i l l u n s a t i s f a c t o r y . The poor " i n v i v o " r e l e a s e from these products i s most probably due to poor f o r m u l a t i o n , which.is revealed by t h e i r poor d i s i n t e g r a t i o n . c h a r a c t e r i s t i c s . Product W and X do not d i s i n t e g r a t e w h i l the d i s i n t e g r a t i o n time of Product DD-j_ i s abnormally long 70 V I I I . SUMMARY In t h i s i n v e s t i g a t i o n , a completely automatic continuous f l o w d i s s o l u t i o n procedure was developed and t e s t e d . P e r t i n e n t c o n d i t i o n s of d i s s o l u t i o n were studi e d and chosen to t e s t seven brands of commericially a v a i l a b l e phenylbutazone t a b l e t s . "In v i v o " s t u d i e s were c a r r i e d out on these products by determining serum l e v e l s a f t e r the a d m i n i s t r a t i o n of the products to nine s u b j e c t s . From the " i n v i t r o " data obtained by the t e s t d i s s o l u t i o n procedure, a T^Q$ v a l u e °f 120 minutes was chosen as the l i m i t of acceptance f o r the t e s t products. Of the seven t e s t products, only f o u r were acceptable on the b a s i s of both the " i n v i v o " and the " i n v i t r o " data. C o r r e l a t i o n of the " i n v i v o " and the " i n v i t r o " data r e s u l t e d i n " l e a s t squares" l i n e s w i t h negative slopes. This c o r r e l a t i o n i n d i c a t e d the p o s s i b i l i t y of usi n g t h i s " i n v i t r o " d i s s o l u t i o n procedure i n the p r e d i c t i o n of the " i n v i v o " a v a i l a b i l i t y of phenylbutazone from the s o l i d dosage form. 71 IX. BIBLIOGRAPHY 1. Brudney, N., Stewart, D.J. and Eustace, B.T. : Canad. Med. Ass. J . , 90 : 980 (1964). 2. Burns, J . J . , Rose, R.K., Chenkin, T., Goldman', A., S c h u l e r t , A., and Brodie, B.B. : J . Pharmacol. Exp. Ther., 109 : 346 (1953). 3 . Canada, Dept. of N a t i o n a l Health and Welfare, Food and Drug D i r e c t o r a t e : The determination of the d i s i n t e g r a t i o n time of t a b l e t s , Ottawa, 1965. 4 . Canada Food and Drugs Act, and Regulations : Food and Drugs act and r e g u l a t i o n s , l o o s e - l e a f o f f i c e c o n s o l i d a t i o n , Sec.0.015, The Queen's P r i n t e r , Ottawa, 1954. 5 . Chapman, D.G., C r i s a f i o , R. and Campbell, J.A. : J . Pharm. S c i . , 43 : 297 (1954). 6 . Idem : J . Pharm. S c i . , 45 : 374 (1956). 7 . Ganderton, D., Hadgraft, J.W., R i s p i n , W.T. and Thompson, A.G. : Pharm. Acta Helv., 42 : 152 (1967). 8. Hamlin, W.E., Nelson, E., B a l l a r d , B.E. and Wagner, J.G. : J . Pharm. S c i . , 51 : 432 (1962). 9 . Levy, G. : J . Pharm. S c i . , 50 : 388 (1961). 10. Levy, G. : J . Pharm. S c i . , 51 : 947 (1962). 11. Levy, G. : J . Pharm. S c i . , 52 : 1039 (1963). 12. Levy, G. and Gumtow, R.H. : J . Pharm. S c i . , 52 : 1139 (1963) . 13. Levy, G. and Hayes, B.A. : New Eng. J . Med., 262 : 1053 (I960). 14. Levy, G. and S a h l i , B.A. : J . Pharm. S c i . , 51 : 58 (1962). 15. Levy, G. and Tanski, W., J r . : J . Pharm. 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Searl, R.O. and Pernarowski, M. : Ganad. Med. Ass. J . , 96 : 1513 ( 1 9 6 7 ) . 2 7 . Stephens, C.A.L., J r . , Yeoman, E.E., Holbrook, W.P., H i l l , D.F. and Goodin, W.L. : J . A. M. A., 150 : 1084 (1952) . 2 8 . Sperandio, G.J., Evanson, R.V. and DeKay, H.G. : J . Am. Pharm. Ass. ( S c i . Ed.), 37 : 71 ( 1 9 4 8 ) . 29. Souder, J.C. and Ellenbogen, W.C. : Drug Std., 26 : 77 (1953) . 30. United States Pharmacopeia, 16 th. rev., Mack Publishing Co., Easton, Pa., I 9 6 0 . 3 1 . Wurble, M.S. : Am. J . Pharm., 102 : 318 ( 1 9 3 0 ) . 3 2 . Wurster, D.E. and Taylor, P.W. : J. Pharm. S c i . , 54 : 169 ( 1 9 6 5 ) . 3 3 . Woo, W.W. : B. S. P. Thesis, University of B r i t i s h Columbia, Vancouver, 1 9 6 6 . 3 4 . Yen, J.K.C. : Can. Pharm. J . , 97 : 493 ( 1 9 6 4 ) .

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