Die Pharmazie

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D 20485 E

Die Pharmazie

An International Journal

of Pharmaceutical Sciences 3 48. Jahrgang März 1993

Aus dem Inhalt

Übersicht

• Amidinohydrazone in der Arzneistofforschung, Teil 2 Originale

• Synthese von N-(2-Carboxy-thieno-pyridinylformamidinen mit antianaphylaktischer Wirkung

• Synthesis of l,2,4-triazolo[4,3-a](l,3)diazepines

• Neue Thieno-Verbindungen

• Determination of hyrcanoside by FIA

• HPLC-Trennung chiraler Benzo-l,4-diazepine

• In vitro penetration of betamethasone-17-valerate into a multilayer membrane system

• Incorporation of amphotericin B in liposomes decreases its toxicity in mice

• Interaction of ibuprofen with early chick embroygenesis

• New triterpenoids from galls of Pistacia integerrima

• Analytik von Prazosin-Hydrochlorid

• Translocation of protein kinase C in human platelets by patients with uremia

• Quercetin-3-O-sambubioside in Quercus-species

• Acylated flavonol glucosides ofHelichrysum bractea- tum

Erfahrungsbericht

• pH-Messung in W/L-Emulsionssalben

GOVl

Govi-Verlag

Pharmazeutischer Verlag GmbH Frankfurt am Main

Redaktion Chefredakteur S. Pfeifer (Berlin)

Stellvertretender Chefredakteur P . Pflegel (Greifswald) Wissenschaftlicher Beirat G . B e r n ä t h (Szeged)

R. B r a u n (Frankfurt a m M a i n ) D . D . Breimer (Leiden) M . D i t t g e n (Jena) D . D u c h e n e (Paris) G . F r a n z (Regensburg) H . G e b l e r (Hannover) G . H e i n i s c h (Innsbruck) K . H i l l e r (Berlin) H . - D . H ö l t j e (Berlin) L . K n y (Berlin)

L . K r ö w c z y h s k i ( K r a k o w ) W . K u b e l k a (Wien) M . L u c k n e r (Halle/Saale) F . M a r k w a r d t (Erfurt) H . M ö h r l e (Düsseldorf) H . M o r c k (Frankfurt a m M a i n ) E . M u t s c h i e r (Frankfurt a m M a i n ) P . N u h n (Halle/Saale)

H . - H . O t t o ( F r e i b u r g i . Br.) J. Richter (Berlin)

P . C . Schmidt ( T ü b i n g e n ) W . Schunack (Berlin) F . T . S m i t h (Auburn) O . Sticher ( Z ü r i c h ) K . Szendrei (Szeged) K . T h o m a ( M ü n c h e n ) R. V o i g t (Berlin) G . W a g n e r (Leipzig) R. A . de Zeeuw (Groningen)

PHARAT 48 (3) 161-240 (1993)

ISSN 0031-7144 • Pharmazie • Frankfurt/Main • 48 (1993) 3 • S. 161 -240

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Lehrstuhl für Biopharmazie

¹

der Universität Halle und Institut für Pharmazeutische Technologie und Biopharmazie

²

der Universität Heidelberg

Study of t h e i n v i t r o penetration of t h e topical g l u c o c o r t i c o i d betamethasone-17-valerate

f r o m s o l u t i o n - t y p e gels into a multilayer membrane s y s t e m

B A R B A R A B E N D A S

¹

, A. G Ö P F E R I C H

²

, G. LEE

²

and R. NEUBERT

¹

T h e

in vitro

transport of betamethasone-17-valerate (1) into a multilayer membrane system has been inves- tigated. Subsaturated formulations of 1 were studied as formed by m i x i n g appropriate propylene g l y c o l / w a - ter cosolvent systems. T h e A U C (drug concentration in acce ptor membrane as a function of time) and the diffusivity of the d r u g in the vehicle were used to evaluate the results of the

in vitro

transport. T h e importance and relationship between solubility, parti- tion coefficient, and diffusivity for the process of

in vitro

penetration of 1 are discussed.

Untersuchungen zur in vitro-Penetration von Bethame- thason-17-valerat aus Lösungsgelen in ein Mehrschicht- membransystem

Gegenstand der Untersuchungen ist der in v i t r o - T r a n s - port v o n Betamethason-17-valerat (l)aus L ö s u n g s g e l e n in ein M e h r s c h i c h t m e m b r a n m o d e l l . A l s H y d r o g e l e k a - men Systeme mit unterschiedlichen P r o p y l e n g l y c o l / W a s s e r - M i s c h u n g e n zur A n w e n d u n g . Z u r A u s w e r t u n g der Ergebnisse wurden die A U C (Arzneistoffkonzentra- tion i n der A k z e p t o r m e m b r a n als F u n k t i o n der Zeit) und der Diffusionskoeffizienz des Arzneistoffes i m V e h i - kel herangezogen. D i e Bedeutung und der Z u s a m m e n - hang zwischen L ö s l i c h k e i t , Verteilungskoeffizient und Diffusionskoeffizient für die i n v i t r o - P e n e t r a t i o n v o n 1 werden diskutiert.

1. Introduction

T h e importance of the interactions between drug, vehicle, and skin for percutaneous a b s o r p t i o n are complex, as can be appreciated from a study of the available literature [1-6].

There are various parameters that affect the mechanisms responsible for specific d r u g release a n d penetration. O n e way to i m p r o v e the d e r m a l a b s o r p t i o n of drugs is to manipulate their physico-chemical properties by p r o d u c i n g derivatives w i t h increased potential for permeation through the skin barrier. M a n y investigations have been carried out to produce prodrugs w i t h the a i m to increase l i p o p h i l i c i t y . T h i s changes related parameters like p a r t i t i o n coefficient, solubility and even diffusivity [7-16].

T h e d e r m a l uptake properties of a d r u g are also influenced by the choice of the type of vehicle a n d its ingredients. T h u s the interactions of the physico-chemical properties of a d r u g and vehicle play an i m p o r t a n t role for the delivery of the d r u g into the s k i n from its f o r m u l a t i o n [17-20]. F u r t h e r m o r e , the components of the vehicle are themselves able to penetrate into the skin. T h i s can result i n a decreased diffusional resistance of the stratum c o r n e u m [21-23].

In this study, betamethasone-17-valerate (1) i n the mixed cosolvent system propylene glycol ( P G ) / w a t e r has been used to study solution-type gels as t o p i c a l vehicles. T h e influence of P G o n the

in vitro

penetration of the d r u g out of the vehicle into a membrane system has been investigated. T h e ratio of the 1 concentration i n s o l u t i o n ( C

^v

) to its saturated solubility in the gel ( C

s

) was m a i n t a i n e d constant, w i t h the d r u g only being present i n the dissolved state.

2. Investigations, results and discussion

The

in vitro

penetration experiments were run using s o l u t i o n - type formulations of 1 i n simple hydrogels with v a r y i n g content of cosolvent. T o classify the l i p o p h i l i c i t y of 1 its saturated solubility in the acceptor l i p i d d o d e c a n o l ( D D ) , in propylene glycol ( P G ) and i n water was determined (Table 1).

It is slightly soluble i n water, but shows a high solubility in D D . T h e percentage of dissolved 1 in water increases with increasing P G content ( F i g . 1). If the ratio of C

^v

to C

^s

is kept constant for a l l systems investigated, then the same solubility conditions are established. F r o m previous results [31] one can expect that for fixed t h e r m o d y n a m i c activity, the amount penetrated into the membrane w i l l be constant up to a certain value. A b o v e this, the release of 1 w i l l be c o n t r o l l e d by p a r t i t i o n behaviour.

Table 1 Solubility of 1 in D D , P G and water as solvent systems (mean value + S D , n = 4)

D D P G Water

Solubility 12.5 ± 1.0 3.0 ± 0.01 0.012 ± 0.008 [mg/ml]

In this study, the A U C was used as a parameter to estimate the amount a n d rate of

in vitro

penetration of 1 from each gel as a function of time and P G - c o n t e n t ( F i g . 2).

C o n s i d e r a t i o n of the cumulative amount of 1 transported in dependence o n the time shows that w i t h P G content above 5 0 % the amount penetrated after 200 m i n decreases ( F i g . 2).

W h e n the A U C is expressed as a function of P G content (see T a b l e 2) it can be estimated that the

in vitro

penetration is independent of P G content up to 4 0 % . T h e slight increase i n A U C values in the range of 50 and 6 0 % P G are caused by higher amounts of 1 being transported w i t h i n the first 15 m i n (Fig. 3.). T h i s increased penetration rate is caused by convec- tive transport. It has been found that P G is able to penetrate

S o l u b i l i t y [ m g / m l ]

3H

2H

1

^{i , r -}

2 0 4 0 6 0 8 0 1 0 0 P G c o n t e n t [ ° /⁰]

F i g . 1. S o l u b i l i t y o f 1 i n P G / w a t e r m i x t u r e s

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P e n e t r a t e d a m o u n t [ % ] 1 0 0 -

8 0 - . . i 1 . ,

6 0 -

4 0 -

2 0 -

2 0 3 0 4 0 5 0 6 0 7 0 8 0 P G c o n t e n t [ % ]

F i g . 2. In vitro p e n e t r a t i o n o f 1 from s o l u t i o n - t y p e gels as function o f P G

considerably i n the acceptor membranes [32]. D r u g molecules dissolved i n the cosolvent are, therefore, transported with the cosolvent flow into the acceptor. W i t h increasing P G content in the gels, the a m o u n t o f 1 penetrated into the acceptor decreases, a l t h o u g h the extent of convection w o u l d be expect- ed to increase.

T h e study of the p a r t i t i o n behaviour of 1 between vehicle a n d acceptor indicates that the ability of 1 to distribute into the acceptor decreases w i t h increasing P G content o f the d o n o r m e d i u m (Fig. 4). B y c o m p a r i n g the solubility a n d p a r t i t i o n behaviour as functions o f P G content, it is evident that these two contrary processes result i n a n o p t i m a l cosolvent concen- tration for the in vitro penetration o f 1 [33-35]. T h e p a r t i t i o n coefficient w i l l be the most i m p o r t a n t parameter, p r o v i d e d the concentration o f d r u g is i n a constant relation to its saturated solubility a n d it other factors influencing the d r u g release, such as change i n viscosity o f the vehicle, c a n be neglected. T h e decrease i n p a r t i t i o n coefficient seen w i t h higher cosolvent concentration i n the gels arises from a higher affinity of the d r u g for the vehicle, consequently lower uptake rates of 1 are measured.

T h e diffusion coefficients for the 1 i n the vehicles lie i n the range o f 1 0 "

7

c m

2

/ s ( F i g . 5) as is typically found for isotropic hydrogels. T h e y are independent o f the P G content up to 4 0 % , A t higher P G contents, D decreases slightly but remains w i t h i n the same order o f magnitude. T h e higher affinity o f the d r u g for the f o r m u l a t i o n at these large P G contents manifests itself i n lower values for K . In accordance w i t h F i c k ' s First law, these two changes result i n a decrease i n the a m o u n t o f 1 penetrated.

P e n e t r a t e d a m o u n t [%>]

6 0 J

20]

2 0 3 0 4 0 5 0 6 0 7 0 8 0 P G c o n t e n t [%>]

K 1 5 -

1 0 -

5 -

2 0 4 0 6 0 8 0 1 0 0

P G c o n t e n t [ ° /⁰]

F i g . 4. Penetrated a m o u n t o f 1 in dependence o f P G content i n 15 m i n

W e conclude that b o t h A U C a n d diffusivity c a n be used to evaluate the in vitro penetration of a l i p o p h i l i c substance from simple hydrogels (Table 2). T h e c a l c u l a t i o n of diffusivity allows a quantitative estimation of the importance of the A U C .

Table 2 Comparision of the parameters A U C and diffusivity for evaluating the in v i t r o penetration of 1 from hydrogels with varrying P G content

P G content [%] A U C [% • minj D [cm2 s]

20 13690 1.105 E-007

30 13639 1.179 E-007

40 13439 1.173 E-007

50 13866 8.204 E-008

60 13734 8.006 E-008

70 11743 6.4865 E-008

80 11597 6.2296 E-008

3. Experimental

3.1. M a t e r i a l s

Betamethasone-17-valerate (1) a n d p r o p y l e n e g l y c o l ( P G ) were p u r c h a s e d from C O M P h a r m a H a n d e l s - G m b H , H a m b u r g . C h l o r o f o r m , m e t h a n o l a n d c o l l o - d i o n ( 4 % w/w) were p r o v i d e d by L a b o r c h e m i e A p o l d a . D o d e c a n o l ( D D ) , t e t r a z o l i u m blue a n d p o t a s s i u m h y d r o x i d e were p u r c h a s e d from M e r c k , D a r m s t a d t a n d s o d i u m c a r b o x y m e t h y l c e l l u l o s e from F l u k a F e i n c h e m i k a l i e n G m b H , N e u - U l m .

D [ c m2/ s ]

1,200

E - 0 7 -

1. 1 0 0 E - 0 7 - j •

1,000

E - 0 7 - /

9,000 E - 0 8 - / 8,000 E- 0 8 - /

7000E-08-*

6,000 E- 08-' 1 1 1 1 1 1 1 r -

5

10 20

3 0 4 0 P G

Sic

F i g . 3. Penetrated a m o u n t o f 1 i n dependence o f P G content w i t h i n 200 m i n P i g . 5. P a r t i t i o n coefficient o f 1 between vehicle a n d acceptor l i p i d

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3.2. M e t h o d s

3.2.1. Solubility of \ in PG water mixtures

A series o f P G / w a t e r m i x t u r e s was prepared from 10 to 8 0 % P G in 1 0 % (w/w) increments. D r u g was a d d e d to each until a suspension formed, w h i c h was then s h a k e n for 24 h at 32 C + 1 C . T h e samples were centrifuged a n d the supernatant s o l u t i o n s were assayed for their d r u g content. T h e t e t r a z o l i u m blue m e t h o d for g l u c o c o r t i c o i d s was used.

3.2.2. Determination of partition behaviour

T o characterize the p a r t i t i o n b e h a v i o u r o f 1 between the s o l u t i o n - t y p e gels a n d the acceptor m e d i u m ( D D ) the penetration experiments were c a r r i e d out for a finite time (300 min). T h e d r u g content i n the m e m b r a n e s was then determined, a n d the p a r t i t i o n coefficient ( K ) c a l c u l a t e d a c c o r d i n g to N e r n s t :

aa qu

T h e f o l l o w i n g c o n d i t i o n s have to be taken into a c c o u n t : 1. n o n ideal, dilute s o l u t i o n s were used; a n d 2. the f o r m u l a t i o n excipients, e.q. P G , c o u l d penetrate into the acceptor m e d i u m .

3.2.3. Preparation of the gels

A suitable a m o u n t o f s o d i u m c a r b o x y m e t h y l c e l l u l o s e was m i x e d w i t h the a p p r o p r i a t e a m o u n t o f P G a n d the m i x t u r e made up w i t h distilled water. T h e gels were then a d d e d to the d r u g a n d stirred to a h o m o g e n o u s f o r m u l a t i o n .

3.2.4. In vitro membrane transport studies

T h e m u l t i l a y e r m e m b r a n e system used has been described p r e v i o u s l y [24, 25].

A defined a m o u n t o f the f o r m u l a t i o n (10 mg) was a p p l i e d to the acceptor system, w h i c h was attached i n a cell w i t h a fixed a p p l i c a t i o n area (4 c m2) . T h e acceptor system consists o f one l i p i d m e m b r a n e to m a i n t a i n a p p r o x i m a t i v e sink c o n d i t i o n s (acceptor l o a d i n g after c o m p l e t e a b s o r p t i o n of the d r u g :

1 10%). T h e experiments were c a r r i e d out sixfold at 32 C ± 1 C in a t e m p e r a t u r e - c o n t r o l l e d c h a m b e r . A t selected time intervals the m o d e l a p p a r a t u s was r e m o v e d from the c h a m b e r , the p e n e t r a t i o n cells were disassembled, the f o r m u l a t i o n r e m a i n i n g o n the exposed surface was r e m o v e d , a n d the m e m b r a n e s assayed for their content o f 1.

3.2.5. Extraction of the drug

T h e separated m e m b r a n e s were extracted w i t h 2 m l for each sample by s h a k i n g for 30 m i n .

3.2.6. Analytical method

T h e content o f 1 i n the v a r i o u s m e m b r a n e s of the m o d e l was assayed indirectly

x = 0 x =

F o r m u l a t i o n A c c e p t o r c ( x , t ) m ( t )

D

d c ( 0 , t ) , .

— = 0 c ( h , t ) = 0 d x t

f d c ( x , t ) m ( t ) = - D A —

d x d t

0

F i g . 6. S i n k m o d e l for d e s c r i b i n g the diffusional release o f a d r u g from t h i n film i n t o adjacent acceptor

by m e a s u r i n g a b s o r b a n c e after r e d u c t i o n o f t e t r a z o l i u m blue t h r o u g h the o x i d a t i o n o f the k e t o l structure of the g l u c o c o r t i c o i d in a l k a l i n e s o l u t i o n . T h e absorbance of the f o r m a z a n c o m p o u n d o b t a i n e d was measured at 525 n m .

3.2.7. Analysis of data

T h e results were expressed as plots of 1 c o n c e n t r a t i o n in the acceptor m e m b r a n e versus time. T h e statistical m o m e n t s o f each curve were calculated a c c o r d i n g to B r o c k m e i e r [26, 27]. A c c o r d i n g to the literature, the A U C can be used for c h a r a c t e r i z i n g the extent of d r u g penetration into systems such as excised h u m a n s k i n a n d the membranes used here [28, 29].

A d d i t i o n a l l y , the diffusivity o f the d r u g in each f o r m u l a t i o n was calculated.

T h e o r e t i c a l values for the time-dependent mass o f d r u g in the acceptor m e m b r a n e were fitted by the N e l d e r - M e a d m e t h o d to the c o r r e s p o n d i n g e x p e r i m e n t a l l y determined values. T h e c a l c u l a t i o n was based o n a finite- difference m e t h o d [30] to a p p r o x i m a t e n u m e r i c a l l y the diffusion e q u a t i o n for the linear movement o f a d r u g w i t h constant diffusity, D . t h r o u g h a finite plane sheet: D c ( x , t )x x — c(x, t), = 0. where c(x. t) = c o n c e n t r a t i o n in f o r m u l a - tion, x = space c o o r d i n a t e , and t = time. T h e sink m o d e l used is illustrated in F i g . 6 a n d describes the diffusional release o f the d r u g from a t h i n film of f o r m u l a t i o n into an adjacent acceptor m e m b r a n e . T h e latter is taken to act as a sink, resulting i n : c(h. t) = 0. T h e flux of d r u g entering the acceptor m e m b r a n e . J, is equal to that l e a v i n g the f o r m u l a t i o n , i.e. J = d m (x. t) A d t |x h.

References

1 K a t z , M . ; P o u l s e n . B. J . : J . Soc. C o s m e t . 23, 565 (1972) 2 Idson, B . : J . P h a r m . Sei. 64, 901 (1975)

3 L i p p o l d , B . C . : A c t a P h a r m . T e c h n o l . 27, 1 (1981) 4 H o r s c h , W . : P h a r m a z i e 39. 598 (1984)

5 Ziegenmeyer, J . : P h a r m , unserer Zeit 15, 129 (1986)

6 S t ü t t g e n . G . : D i e H a u t als T r a n s p o r t o r g a n für Arzneistoffe, S. 1. Steinkopff V e r l a g , D a r m s t a d t 1990

7 L a n d m a n n , L . : P h a r m , unserer Zeit 20, 155 (1991)

8 F l y n n , G . L . ; Y a l k o w s k y , S. H . : J . P h a r m . Sei. 61, 839 (1972) 9 C o o p e r , E . R . : J . C o n t r o l l e d Release 1, 153 (1984)

10 G u y , R. H . ; C a r l s t r ö m , E . M . ; B u c k s , D . A . W . : H i n z , R. S.: J . P h a r m . Sei.

75, 968 (1986)

11 Y o u n g - H a r v e y , J . A . : Rae, 1. D . ; P i t m a n , I. H . : Int. J . P h a r m . 30, 151(1986) 12 J o h a n s e n , M . : M o l l g a a r d , B . ; W o t t o n , P. K . ; L a r s e n , C ; H o l l g a a r d , A . :

i b i d . 32, 199 (1986)

13 W a r a n i s . R. P . ; S l o a n . K . B . : J . P h a r m . Sei 76, 587 (1987)

14 A l - K h a m i s , K . ; D a v i s , S. S.; Hadgraft, J . : Int. J . P h a r m . 40, 111 (1987) 15 T o j o , K . ; C h i a n g , C . C ; C h i e n , Y . W . : J . P h a r m . Sei. 76, 123 (1987) 16 B u n d g a a r d , H . ; M o r k , N . ; H o l l g a a r d , A . : Int. J. P h a r m . 55, 91 (1989) 17 P r u s t r u p , J . ; F u l l e r t o n , A . ; C h r i s t r u p , L . ; B u n d g a a r d , H . : i b i d . 71,105(1991) 18 L o t h , H . : i b i d . 68, 1 (1991)

19 Asche, H . ; B o t t a , L . ; Rettig, H . ; W e i r i c h , E . G . : A c t a P h a r m . H e l v . 60, 232 (1985)

20 S l o a n , B . K . ; Siver. K . G . ; K o c h , A . M . : J . P h a r m . Sei. 75, 744 (1986) 21 N a n n i p i e r i , E . ; C a r e l l i , V . ; D i C o l o , G . ; G i o r g i , I.; Serafini, M . F . : Int. J .

C o s m e t . Sei. 12, 21 (1990)

22 Hadgraft, J . : P h a r m . Int. 5, 252 (1984)

23 R i t s c h e l . W . A . ; S p r o c k e l . O . L . : D r u g s of today 24, 613 (1988) 24 F ü r s t , W . ; N e u b e r t , R . ; W i l d n e r , R . : P h a r m a z i e 42, 452 (1987) 25 N e u b e r t , R . ; W o h l r a b , W . : A c t a P h a r m . T e c h n o l . 36, 197 (1990) 26 B r o c k m e i e r , D . : A r z n e i m . - F o r s c h . 31, 746 (1981)

27 I d e m : A c t a P h a r m . T e c h n o l . 32, 164 (1986)

28 N e u b e r t , R . ; Bendas, C ; W o h l r a b , W . ; G i e n a u , B . ; F ü r s t . W . : Int. J . P h a r m . 75, 89 (1991)

29 N e u b e r t , R . ; Bendas, B . ; W o h l r a b , W . : E u r . J . P h a r m . B i o p h a r m . 38. 11 (1992)

30 C r a n k , J . ; N i c o l s o n , P. A . : P r o c . C a m b . P h i l . Soc. 43, 50 (1947) 31 L i p p o l d , B . C . : A c t a P h a r m . T e c h n o l . 27, 1 (1981)

32 Bendas, B . ; N e u b e r t , R . : i n p r e p a r a t i o n

33 Patel, A . R . ; V a s a v a d a , R. C : P h a r m . Res. 5, 116 (1988)

34 P o u l s e n , B . J . ; Y o u n g , E . ; C o q u i l l a , V . ; K a t z , M . : J . P h a r m , sei. 57,928 (1968) 35 O s t r e n g a , J . ; Steinmetz, C ; P o u l s e n , B . : i b i d . 60, 1175 (1971)

Received September 4, 1992 Prof. D r . R e i n h a r d N e u b e r t W e i n b e r g w e g 15 4010 H a l l e Saale