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Percutaneous absorption of ketoprofen. I. In vitro release and percutaneous absorption of ketoprofen from different ointment bases
PHARMACEUTIC ACTA HELVETIAE ELSEVIER
Pharmaceutics
Acta Helvetiae 71 (1996) 205-212
Percutaneous absorption of ketoprofen. I. In vitro release and percutaneous absorption of ketoprofen from different ointment bases Z. Giirol a, S. Hekimoglu
a’*, R. Demirdamar
‘, M. $umnu a
aHacettepe UniuersiQ, Faculty of Pharmacy, Department of Pharmaceutical Technology, 06100 Ankara, Turkq b Hacettepe Uniuersity, Faculty of Pharmacy, Department of Pharmacology, 06100 Ankara, Turkey Received 31 July 1995; accepted
19 September
1995
Abstract Ketoprofen (KP) is a potent non-steroidal anti-inflammatory drug which is used for the treatment of rheumatoid arthritis. The oral administration of KP can cause gastric irritation and renal adverse effects. Topical application of the drug can bypass gastrointestinal disturbances and provide relatively consistent drug levels at the site of action. Since the efficacy of an ointment depends on the type of ointment base and the concentration of the drug, four different bases (white petrolatum, cold cream, hydrophilic ointment and Carbopol 940 gel) were used at 1, 3, 5, 7 and 10% concentrations of KP to evaluate the effect of ointment base and concentration. The general rank
order of the drug release was found to be: Carbopol gel > hydrophilic ointment > cold cream > white petrolatum. There was a positive correlation between the concentration of KP and release rate for all bases except Carbopol gel. The in vivo percutaneous absorption of KP from different ointment bases at 3% concentration was studied by carrageenan-induced paw edema in mice. The rank order of the percent edema inhibition was as follows: Carbopol gel 2 hydrophilic ointment > cold cream > white petrolatum. There was a good correlation between the in vitro and in vivo results. Keywords: Ketoprofen;
Drug release; Percutaneous
absorption;
Carrageenan
1. Introduction Ketoprofen (KP) is a non-steroidal anti-inflammatory, analgesic and antipyretic drug used for the treatment of rheumatoid arthritis (Lussier et al., 1976; Peltola, 1976; Zutshi and Mason, 19761, osteoarthritis (Famaey and Colinet, 1976; Kantor, 1986) ankylosing spondylitis (Fossgreen, 1976) and gout (Fossgreen, 1976; Kantor, 1986). It is equally or more potent than the other non-steroidal anti-inflammatory drugs (NSAIDS) with respect to some
* Corresponding
author. Fax: (312) 311 47 77, Tel: (312) 310 15 24.
paw edema test
effects such as anti-inflammatory and analgesic activities (Peltola, 1976; Saxena and Saxena, 1978). Although KP is rapidly absorbed, metabolized and excreted, it causes some gastrointestinal complaints such as nausea, dyspepsia, diarrhea, constipation and some renal side effects like other NSAIDs (Fossgreen, 1976). Therefore, there is a great interest to develop the topical dosage forms of these NSAIDs to avoide the oral side effects and to provide relatively consistent drug levels at the application site for prolonged periods (Babar et al., 1990; Muktadir et al., 1986; Ranman et al., 19901. The main objective of this study is to prepare the topical formulations of KP. For this aim, the effects of type of ointment bases and drug concentration on the in vitro release and percutaneous absorption of KP were evaluated.
0031-6865/96/$15.00 Copyright 0 1996 Elsevier Science B.V. All rights reserved PII SO03 l-6865(96)0001 I-8
206
Z. Giirol et al. /Pharmaceutics
2. Experimental
the aid of triethanolamine. Tiomersal and sodium metabisulfide were added to this solution, then KP solution was added to polymer dispersion and stirred with continous agitation.
2.1. Materials The following chemicals were used as received from the suppliers. Ketoprofen (Drifen Pharm. Co., Turkey), Carbopol 940 (Goodrich), carrageenan (Sigma), paraffine liquid (Merck), Pluronic P94 (Ugine Kuhlmann), potassium dihydrogen phosphate (Merck), sodium hydroxide (Merck), sodium lauryl sulfate (Merck), sodium metabisulfide (Merck), stearyl alcohol (Merck), thiomersal (Sigma), triethanolamine (Merck), Spectrapor 2 regenerated cellulose membrane MW Cut off 14000 (Spectrum). 2.2. Equipment Double beam UV spectrophotometer (Hitachi 220 S), specially designed diffusion cell apparatus (Ildam, Turkey), DSC apparatus (Shimadzu DT-40) dial thickness gauge (Ozaki). 2.3. Preparation
Acta Heluetiae 71 (1996) 205-212
of the formulations
White petrolatum as a hydrocarbon base, cold cream (USP) as an absorption base, hydrophilic ointment (USP) as a water washable base and Carbopol gel as a water-soluble base were selected. All formulations were prepared at 1, 3, 5, 7 and 10% concentrations of KP. White petrolatum ointment was suspension type which was prepared by geometric dilution of KP and melted base in the mortar. The other ointments were solution types which were prepared with the KP solution in water and triethanolamine mixture. Cold cream (USP XXI) and hydrophilic ointment (USP XXI) were prepared by some modifications of the formulations. (Babar et al., 1990; Muktadir et al., 1986; Ranman et al., 1990). 6% of triethanolamine was added to the formulations to dissolve KP in the bases. In addition, methyl and propyl parabens were added to cold cream for preservation. All the aqueous phase ingredients except KP and triethanolamine were dissolved in two-thirds of the water. Aqueous and oil phases were heated to 70 + 5°C separately and then the water phase was added to the oil phase with continuous stirring. The ointment was then cooled to approximately 40-50°C and KP was dissolved in one-third of the water and triethanolamine mixture was added and stirred till the ointment was cooled to room temperature. Carbopol gel base formulation and preparation is as follows: Carbopol940 1.5 g, triethanolamine 9 g, tiomersal 0.01 g, sodium metabisulfide 0.1 g, water qs to 100 g. Carbopol 940 was dispersed in two-third of the water for overnight. KP was dissolved in the remaining water with
2.4. In tiitro release studies Specially designed vertical-type glass diffusion cells with side-arm were used. Donor and receptor compartments were attached to each other by a separating Spectrapor 2 membrane. 1 g of ointment was placed in the donor compartment, the volume of the receptor compartment was 20 ml. pH 7.4 degassed phosphate buffer at 37’C was used as receptor solution. It was stirred magnetically at 600 rpm. The whole system was placed in a constant water bath at 37 k 1°C. Samples were withdrawn at 15, 30, 60, 90, 120, 150 and 180 minute intervals and analyzed for KP contents spectrophotometrically at a wavelength of 260 nm. The volume of the diffusion medium was kept constant by replacing the amount of sample with an equal volume of the diffusion medium. The concentration of KP in each sample was determined by spectrophotometrically. Blank ointment samples were used as a reference and all diffusion studies were the average of six experiments. Solubility of KP in the receptor solution was found as 880.5 mg/ml at 25°C. 2.5. Mathematical
evaluation
of release experiments
Mathematical analysis of the release of KP from different ointment bases was carried out using the Higuchi equation for solution type of ointments (Higuchi, 1962). When the amount of drug released was plotted against the square root of time, a straight line was obtained. The diffusion coefficient of drug (D) was calculated using the following equation: _ Dt
Q=2C,d
7T
Q: amount of drug released per unit area (mg/cm*); C,: initial concentration of drug in the ointment (mg/cm3); D: diffusion coefficient (cm*/& t: time (s). The permeability coefficient of KP was calculated using Eq. 2 (Babar et al., 1990): P=
4 A ’ C, . t
P: permeability coefficient (cm/s); released (mg); A: area of the diffusion C,: initial concentration of drug in the time (s). The partition coefficient of the KP
q: amount of drug membrane (cm*); base (mg/cm”); t: between
the oint-
Z. Gird
et al./ Pharmaceutics
ment base and receptor medium was calculated (Muktadir et al., 1986):
using Eq. 3
Acta Helcetiar
207
71 (1996) 205-212
2.7. Statistical
analysis
The differences in the results of in vitro and in vivo studies were evaluated using one-way ANOVA and Student’s t-test.
P.h
K,: partition coefficient; P : permeability coefficient (cm/s); D: diffusion coefficient (cm’/s); h: thickness of the membrane (cm>.
3. Results and discussion
2.6. Percutaneous
3.1. In L:itro release studies
absorption
in mice
Percutaneous absorption of KP was evaluated for its anti-inflammatory activity on carrageenan-induced paw edema in mice. The carrageenan-induced paw edema test is the most common test to evaluate the anti-inflammatory activity of NSAIDs (Chi and Jun, 1990; Winter et al., 1962). Although rats are used frequently (Chi and Jun, 1990; Gemmell et al., 1979; Graziani et al., 1987; Winter et al., 19621, mice may also be used (Schrier et al., 1987). The edema induced by the injection of carrageenan solution is measured using a plethysmometer (Graziani et al., 1987; Schrier et al., 1987; Winter et al., 1962) or a dial thickness gauge (Kasahara et al., 1985; Safak et al., 1992). Five young male mice weighing 20 & 5 g were used to evaluate the effect of ointment base and drug concentration on the percutaneous absorption of KP. The paw edema was measured via dial thickness gauge. 10 mg ointment was applied to the plantar surface of the left hind paw of the mice by gently rubbing 50 times with finger. After three hours, the paw was measured before carrageenan injection and recorded as control (E,). 0.01 ml of a 1% carrageenan solution was injected subplantarly into the same paw. Three hours after the carrageenan injection the paw was measured again and recorded as edema induced by carrageenan injection (E). The percent swelling of the paw was determined using the following equation (Chi and Jun, 1990). % Swelling
E - E, = x 100 4
% Inhibition
=
x 1-
I
100
% Swelling of control group
3.2. Effect of ointment bases The amount of KP released from different ointment bases at various concentrations are shown in Fig. 1 and Fig. 2. It can be seen that the amount of drug released were lowest from white petrolatum for all concentrations.
AMOUNT
OF RELEASED
(mg)
121
(4)
EC: control measuring; E: paw edema induced by carrageenan injection. Control group of the mice was treated only by the ointment base through the same procedure and the percent edema inhibition was determined using Eq. 5 which is the ratio of average swelling of the drug treated group to the average swelling of the control group (Chi and Jun, 1990). % Swelling of drug treated group
All formulations except white petrolatum ointment were solution types which were checked microscopically. The Higuchi equation (Higuchi, 1962) for solution type ointment was used to evaluate the data, and all graphs were obtained by plotting the amount of drug released against the square root of time.
1 (5)
0
2
4
6
8
10
12
14
Jt (mln)
-
1%
+-
3% W.P.
W.P.
+-
1% cc.
mxm 1% H.O.
-=
1% C.Q.
-6-
3% C.C.
-
+
3% C.Q.
3% H.O.
Fig. I. Effect of ointment bases on the release of ketoprofen W.P., white petrolatum: C.C.. cold cream: H.O., hydrophilic C.G., Carbopol 940 gel.
(IS. 38) ointment;
208
Z. Giirol et al. /Pharmaceutics AMOUNT
OF DRUG RELEASED
(rng)
Acta Heluetiae 71 (1996) 205-212 Table 1 Release kinetics of ketoprofen FORMULATIONS
251 20 /
I
0
d
2
4
6
Jt -
7% W.P.
+
+-
10% W.P. +-
8
10
x--x I
12
14
1% Cold cream 3 % Cold cream 5% Cold cream 7% Cold cream 10% Cold cream 1% Hydrophilic oint. 3% Hydrophilic oint. 5% Hydrophilic oint. 7% Hydrophilic oint. 10% Hydrophilic oint. 1% Carbopol940 gel 3% Carbopol940 gel 5% Carbopol940 gel 7% Carbopol940 gel 10% Carbopol940 gel
D X lo-’
P x 10-7
KXIO-2
4.43 7.02 7.48 9.53 10.07 223.40 375.57 396.27 445.65 484.79 5863.20 1947.60 703.88 364.89 176.31
6.67 9.06 9.96 12.30 12.91 54.23 67.47 69.39 72.80 75.47 228.00 131.10 78.78 59.09 41.81
34.41 29.50 30.43 29.50 29.31 5.55 4.1 I 4.00 3.73 3.56 0.89 1.54 2.56 3.70 5.42
D, diffusion coefficient (cm’/s). P, permeability coefficient (cm/s). K, partition coefficient.
(min)
7% CC.
+
7% H.O.
-
10% CC.
+-
10% H.O. -
Fig. 2. Effect of ointment bases on the release of ketoprofen Abbreviations as in Fig. 1.
7% C.Q. 10% C.G.
(7%, 10%).
The rank order of the drug release from different bases was found as: Carbopol gel > hydrophilic ointment > cold cream > white petrolatum for 1,3 and 5% of KP concentrations (Fig. 1). The results of 5% of KP ointment couldn’t be plotted in Fig. 1 for reasons of clarity of the graphic. The differences between the released amount of drug from the bases were statistically significant (p < 0.05). The rank order was modified slightly for 7 and 10% of KP as seen in Fig. 2. The statistically higher amount of drug was released from hydrophilic ointment for 7 and 10% of KP and Carbopol 940 gel was in the second order for these concentrations (p < 0.05). Cold cream and white petrolaturn ointments released KP in lower levels than the others. The calculated diffusion coefficient, permeability coefficient and partition coefficient of KP for all solution type ointments can be seen in Table 1. It can be seen that, the diffusion coefficient and permeability coefficient of KP are highest in the Carbopol 940 gel and lowest for the cold cream. The partition coefficient values show that the affinity of KP for the base is higher in the cold cream than the other bases.
was a good correlation between the concentration of KP and release rate for all bases except Carbopol 940 gel (Fig. 3). 3% concentration of KP was the optimum concentration for Carbopol 940 gel. It couldn’t be considered as a solubility limitation because the solubility of KP in the receptor solution was found as 880.5 mg/lOO ml at 25°C and it was 1100 mg/lOO ml at 37°C in the literature
RELEASE
RATE
1;
0
2
4
6
CONCENTRATION
3.3. Effect of ketoprofen
8
10
12
of KETOPROFEN
(%)
concentration
The release of KP from different ointment bases showed concentration dependency except Carbopol 940 gel. There
-
White
Petrolatum
-++
Hydrophilic
Olnt.
Fig. 3. Effect of drug concentration
+
Cold Cream
-
Carbopol
940 gel
on the release rate of ketoprofen.
14
209
Z. Giirol et al. / Pharmaceutics Acta Helvetiae 71 (1996) 205-212
Carbopol940 Gel (5 % W)
Ketoprofen
I
Carbopol940
Gel (7 % W)
Carbopot940 Gel
r
Carbopol940
Gel (1 % W’)
Carbopol940
(‘C)
of Carbopol 940 gel at different concentrations
(Bramanti et al., 1980). This could be explained by the hydrogen bond formation between KP, Carbopol 940 and triethanolamine. This interaction becomes stronger for high concentrations showing that KP is held tightly in the base. This situation can be seen in Fig. 4. Differential scanning calorimetric graphs of Carbopol 940 gel containing different percentage of KP show the specific peak of KP at 94°C which can be seen in 1% and 3% of KP, but it dissappears in higher KP concentrations. This may be the result of strong interaction between KP, carbopol 940 and triethanolamine. The released amounts for respective concentrations of
940 Gel (10 % a)
Gel (3 % KP)
TEMPERATURE Fig. 4. DSC thermogram
carbpol
of ketoprofen.
KP are significantly different from each other for white petrolatum, cold cream and hydrophilic ointment ( p < 0.05). This concentration dependency can be observed in Table 1 for the diffusion coefficient, permeability coefficient and partition coefficient values.
4. Percutaneous
absorption
of ketoprofen
4.1. Effect of ointment bases A 3% concentration of KP was used for all bases to evaluate the effect of ointment bases because it was the
2. Giirol et al. /Pharmaceutics
210 PERCENT
Acta Heluetiae 71 (1996) 205-212
INHIBITION
PERCENT
INHIBITION
,’ I /I’ i 60
60 -,’
50
50
40
40
>
,
30
20
10
10
k/’
0
L-
, __/ White
petr.
.i--
._~~ Cold cream
~ ,~ ~_i_
Hydr.oint.
FORMULATIONS Fig. 5. Effect ketoprofen.
of ointment
bases
, ~~~~~ /
Garb.940
0 Hyd.oint.3%
gel
absorption
of
optimum concentration for Carbopol 940 gel. The effect of bases on the percutaneous absorption of KP in mice paw was shown in Fig. 5. The percentage inhibition of KP was 10.96, 14.64, 55.76 and 56.62 for white petrolatum, cold cream, hydrophilic ointment and Carbopol gel, respectively. There was a difference between the white petrolatum and cold cream significantly ( p < 0.05). No significant difference was observed between the inhibition of hydrophilic ointment and Carbopol 940 gel (p > 0.05). But white petrolaturn and cold cream showed lower absorption than the others (p < 0.05). There was a good correlation between the in vitro and in vivo results with respect to the effect of ointment bases. Hydrophilic ointment and Carbopol 940 gel bases which caused the highest percutaneous absorption of KP were evaluated for the percent edema inhibition against carrageenan solution alone. While Carbopol gel base showed 1.43% of inhibition, hydrohpilic ointment base caused 69.07% of inhibition. The reason for the high edema inhibition value of hydrophilic ointment base was the high concentration of sodium lauryl sulfate in its formulation. 4.2. Effect of ketoprofen
Carb.gelB%
Carb.gellO%
FORMULATIONS
(3% KP)
on the percutaneous
Hyd.oint.lO%
concentration
3% and 10% concentrations of both hydrophilic ointment and Carbopol 940 gel formulations were compared to
Fig. 6. Effect of drug concentration ketoprofen.
on the percutaneous
absorption
of
investigate the effect of drug concentration. These concentrations were investigated because they were optimum and maximum concentrations in the bases which showed the maximum amount released in in vitro studies. The percentage inhibition of 3% of KP obtained from hydrophilic ointment and Carbopol 940 gel were found to be 55.76% and 56.62%, respectively. These values were 51.77% and 57.66% for 10% of KP in hydrophilic ointment and Carbopol 940 gel (Fig. 6). No significant difference was observed among the formulations (p > 0.05). It was seen that the percutaneous absorption of KP was not concentration-dependent contrary to the results of in vitro studies. As a result of in vitro and in vivo studies; it is obvious that the ointment bases effect the released amount and percutaneous absorption of KP strongly. In vitro data are in agreement with the in vivo data. Solution type ointments show the higher release rates and higher percutaneous absorption than the suspension type. It is in agreement with the results of the studies with benzocaine (Abd El Magid and Abd El Mohsen, 19901, chlorpheniramine maleate (Velissaratou and Papaieannou, 19891, piroxicam (Babar et al., 1990) and naproxen (Ranman et al., 1990). The general rank order of the percutaneous absorption of KP is found as follows: Carbopol gel 2 hydrophilic ointment > cold cream > white petrolatum. Bramanti et al.
Z. Giirol et al. / Pharmaceutics
(1980) obtained similar results with KP release from different type of ointment and gel bases. Drugs in ointment formulations should reach the skin surface in adequate amounts and sufficient rates in order to provide topical therapy. The release of drugs from ointment bases is a function of solubility of that agent in both base and its surrounding medium (Abd El Magid and Abd El Mohsen, 1990; Idson, 1975). If a drug is insoluble in the base, only the drug particles available at the surface of the base apparently dissolve into an aqueous medium (Idson, 1975). Since KP is not soluble practically in the white petrolatum, this formulation shows the lowest release and lowest absorption. Different amounts of KP release and absorption were obtained from cold cream and hydrophilic ointment formulations, although they are both solution type formulations. Solubilized KP was in the internal phase of cold cream while it was in the external phase of hydrophilic ointment. The oily external phase of the cold cream plays a barrier role around the water droplets and prevents the diffusion of the drug to the receptor phase. This may be the explanation of the lower amount released and absorbed of KP from cold cream than from hydrophilic ointment. The percutaneous absorption is also limited by skin and vehicle interaction besides the release rate of the drug. Although the released amount of KP from Carbopol 940 gel is higher than that of hydrophilic ointment for 3% of KP (Fig. 1), there was no significant difference between them in vivo (Fig. 5). It may be due to the interaction between the skin and hydrophilic ointment base. It was shown that hydrophilic ointment base caused 69.07% of inhibition while Carbopol 940 gel base showed 1.43% inhibition against the carrageenan solution. This was the result of the effect of sodium lauryl sulfate on the skin (Barry, 1988; Froebe et al., 1990; Walters, 1989). Although the effect of KP concentration was found to be significant in in vitro release studies for all bases except Carbopol 940 gel (Fig. 3), no significant difference could be observed in the percent edema inhibition of KP from different bases at 3% and 10% of drug concentration (Fig. 6). Generally, there is a positive correlation between drug concentration and release rate and percutaneous absorption of the drug due to the increase in the thermodynamic activity. The thermodynamic activity of a drug is related to its concentration in the vehicle. In different bases, thermodynamic activity of the drug increases linearly with concentration until it reaches the same limiting value which is the value of the saturated solution (Higuchi, 1982). Furthermore, the skin has a limited capacity for the transportation of some drugs (Akhter and Barry, 1985). Because of these reasons. percutaneous absorption of KP from 3% and 10% of hydrophilic ointment and Carbopol 940 gel formu-
Acta HelretiacJ 71 (199hi 205-212
211
lations was similar. The percent inhibition of KP with these two formulations was about 55% and was nearly the maximum response considered possible for a topical antiinflammatory drug using this method (Chi and Jun, 1990). 3% of KP was found as a critical concentration for the formulations investigated in this study. In conclusion, it can be stated tha;; There was a good correlation between the in vitro release studies from the cellophane membrane and carrageenan-induced paw edema test in mice. KP can penetrate to the skin easily. The best topical formulations for KP were both hydrophilic ointment and Carbopol gel at 3% drug concentration. The anti-inflammatory activity of the best formulations was about 557~.
References Abd El Magid, S.I.A. and Abd El Mohsen. M.G. (1990) In vitro and in viva studies of benzocaine release from ointments. S.T.P. Pharm.. 6, 380-387. Akhter, S.A. and Barry, B.W. (1985) Absorption through human skin of ibuprofen and flubiprofen: Effect of dose variation, deposited drug films, occlusion and the penetration enhancer N-methyl-?-pyrrolidone. J. Phann. Pharmacol., 37, 27-37. Babar. A., Solanki, U.D., Cutie, A.J. and Plakogiannis, F. (1990) Piroxicam release from dermatological bases: In vitro studies using cellulose membrane and hairless mouse skin. Drug Dev. Ind. Pharm., 16, 523-540. Barry, B.W. (1988) Action of skin penetration enhancers - the lipid protein partitioning theory. Int. J. Cosm. Sci., 10. 281-293. Bramanti. G.. Pisatura, G., Mazzi, G., Mura, P. and Papini. P. (1980) Release of drugs in vitro from different dermatological preparations. Boll. Chim. Form., 119, 738-744. Chi, S. and Jun, H.W. (1990) Anti-inflammatory activity of ketoprofen gel on carrageenan-induced paw edema in rats. J. Pharm. Sci.. 79. 974-977. Famaey, J.P. and Colinet, E. (1976) (Suppl.) A double-blind trial of ketoprofen in the treatment of osteoarthritis of the hip. Stand. J. Rheum., 14, 129-132. Fossgreen, J. (1976) (Suppl.) Ketoprofen - A survey of current publications. Stand. J. Rheum. 14. 1l-32. Froebe, C.L., Simion, F.A.. Rhein, L.D., Cagan. R.H. and Kligman (1990) A.. Stratum corneum lipid remove] by surfactants: Relation to in viva irritation. Dermatology. I8 I, 277-283. Gemmell, D.K.. Cottney, J. and Lewis, A.J. (1979) Comparative effects of drugs on four paw oedema models in rat. Agents Actions, 9, 107-116. Graziani, G.. Abbiati, G.A.. Dolfini, E.. Testa. R. and Velo, G.P. (1987) Pharmacokinetic, pharmacodynamic and toxicological properties of naproxen gel in laboratory animals. Pharm. Res., 43, 480-490. Higuchi, W.I. (1962) Analysis of data on the medicament release from ointments. J. Pharm. Sci., 51, 802-804. Higuchi, T. (1982) V. In vitro drug release from ointments and creams. In Brandau, R. and Reisen, P. (Eds.). Dermal and Transdermal Absorption. Wissenschaftliche Verlagsgesellschaft, Germany, pp. 90.
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Idson, B. (1975) Percutaneous absorption. J. Pharm. Sci., 64, 901-921. Kantor, T.G. (1986) Ketoprofen. A rewiew of its pharmacologic and clinical properties. Pharmacotherapy 6, 93-103. Kasahara, Y., Hikino, H., Tsurufuji, S., Watanabe, M.,, and Ohuchi, K. (1985) Anti-inflammatory actions of ephedrines in acute inflammations. Planta Med. 5 1, 325-33 1. Lussier, A., Camerlain, M., Menard, H., Myhal, D. and Wehner, S. (1976) (Suppl.) A double blind cross-over evaluation of ketoprofen and aspirin in rheumatoid arthritis. Stand. J. Rheum. 14, 99-104. Muktadir, A., Babar, A., Cutie, A. J. and Plakogiannis, F. M. (1986) Medicament release from ointment bases. III. Ibuprofen: In vitro release and in vivo absorption in rabbits. Drug Dev. Ind. Pharm., 12, 2521-2540. Peltola, P. (1976) (Suppl.) Long-term treatment with ketoprofen in rhumatoid arthritis. Stand. J. Rheum., 14, 115-l 17. Ranman, M.S., Babbar, A., Patel, N.K. and Plakogiannis, F.M. (1990) Medicament release from ointment bases: V. Naproxen in vitro release and in vivo percutaneous absorption in rabbits. Drug Dev. Ind. Pharm., 16, 651-672. Saxena, R.P. and Saxena, U. (1978) A comparative trial of ketoprofen and ibuprofen in patients with rheumatic disease. Curr. Med. Res. Opin., 5, 484-488.
Schrier, D.J., Moniot, S., Gluckman, M.I. and Gilbertsen, R.B. (1987) The topical anti-inflammatory effects of a topical preparation of meclofenamic acid on carrageenan-induced footpad swelling in mice. J. Pharm. Pharmacol., 39, 57-59. Safak. C., Erdogan, H., Palaska, E., Sunal, R. and Duru, S. (1992) Synthesis of 3-(2-pyridylethyl) benzoxazolinone derivatives: Potent analgesic and anti-inflammatory compounds inhibiting prostaglandin E?. J. Med. Chem. 35, 1296-1299. Velissaratou, A.S. and Papaieannou, G. (1989) In vitro release of chlorpheniramine maleate from ointment bases. Int. J. Pharm., 52, 83-86. Walters, K.A. (1989) Penetration enhancers and their use transdermal therapeutic systems. In Hadgraft, J. and Guy, R.H. (Eds.), Transderma1 Drug Delivery, Marcel Dekker, Inc., New York, pp. 197. Winter, C.A., Risley, E.A. and Nuss, G.V. (1962) Carrageenan-induced edema in hind paw of the rat as an assay for anti-inflammatory drugs. Proc. Sot. Exp. Med., 111, 544-547. Zutshi, D. and Mason, M. (1976) (Suppl.) Ketoprofen in rheumatoid arthritis: Its tolerance and therapeutic effect. Stand. J. Rheum., 14, 77-84.
Pharmaceutics
Acta Helvetiae 71 (1996) 205-212
Percutaneous absorption of ketoprofen. I. In vitro release and percutaneous absorption of ketoprofen from different ointment bases Z. Giirol a, S. Hekimoglu
a’*, R. Demirdamar
‘, M. $umnu a
aHacettepe UniuersiQ, Faculty of Pharmacy, Department of Pharmaceutical Technology, 06100 Ankara, Turkq b Hacettepe Uniuersity, Faculty of Pharmacy, Department of Pharmacology, 06100 Ankara, Turkey Received 31 July 1995; accepted
19 September
1995
Abstract Ketoprofen (KP) is a potent non-steroidal anti-inflammatory drug which is used for the treatment of rheumatoid arthritis. The oral administration of KP can cause gastric irritation and renal adverse effects. Topical application of the drug can bypass gastrointestinal disturbances and provide relatively consistent drug levels at the site of action. Since the efficacy of an ointment depends on the type of ointment base and the concentration of the drug, four different bases (white petrolatum, cold cream, hydrophilic ointment and Carbopol 940 gel) were used at 1, 3, 5, 7 and 10% concentrations of KP to evaluate the effect of ointment base and concentration. The general rank
order of the drug release was found to be: Carbopol gel > hydrophilic ointment > cold cream > white petrolatum. There was a positive correlation between the concentration of KP and release rate for all bases except Carbopol gel. The in vivo percutaneous absorption of KP from different ointment bases at 3% concentration was studied by carrageenan-induced paw edema in mice. The rank order of the percent edema inhibition was as follows: Carbopol gel 2 hydrophilic ointment > cold cream > white petrolatum. There was a good correlation between the in vitro and in vivo results. Keywords: Ketoprofen;
Drug release; Percutaneous
absorption;
Carrageenan
1. Introduction Ketoprofen (KP) is a non-steroidal anti-inflammatory, analgesic and antipyretic drug used for the treatment of rheumatoid arthritis (Lussier et al., 1976; Peltola, 1976; Zutshi and Mason, 19761, osteoarthritis (Famaey and Colinet, 1976; Kantor, 1986) ankylosing spondylitis (Fossgreen, 1976) and gout (Fossgreen, 1976; Kantor, 1986). It is equally or more potent than the other non-steroidal anti-inflammatory drugs (NSAIDS) with respect to some
* Corresponding
author. Fax: (312) 311 47 77, Tel: (312) 310 15 24.
paw edema test
effects such as anti-inflammatory and analgesic activities (Peltola, 1976; Saxena and Saxena, 1978). Although KP is rapidly absorbed, metabolized and excreted, it causes some gastrointestinal complaints such as nausea, dyspepsia, diarrhea, constipation and some renal side effects like other NSAIDs (Fossgreen, 1976). Therefore, there is a great interest to develop the topical dosage forms of these NSAIDs to avoide the oral side effects and to provide relatively consistent drug levels at the application site for prolonged periods (Babar et al., 1990; Muktadir et al., 1986; Ranman et al., 19901. The main objective of this study is to prepare the topical formulations of KP. For this aim, the effects of type of ointment bases and drug concentration on the in vitro release and percutaneous absorption of KP were evaluated.
0031-6865/96/$15.00 Copyright 0 1996 Elsevier Science B.V. All rights reserved PII SO03 l-6865(96)0001 I-8
206
Z. Giirol et al. /Pharmaceutics
2. Experimental
the aid of triethanolamine. Tiomersal and sodium metabisulfide were added to this solution, then KP solution was added to polymer dispersion and stirred with continous agitation.
2.1. Materials The following chemicals were used as received from the suppliers. Ketoprofen (Drifen Pharm. Co., Turkey), Carbopol 940 (Goodrich), carrageenan (Sigma), paraffine liquid (Merck), Pluronic P94 (Ugine Kuhlmann), potassium dihydrogen phosphate (Merck), sodium hydroxide (Merck), sodium lauryl sulfate (Merck), sodium metabisulfide (Merck), stearyl alcohol (Merck), thiomersal (Sigma), triethanolamine (Merck), Spectrapor 2 regenerated cellulose membrane MW Cut off 14000 (Spectrum). 2.2. Equipment Double beam UV spectrophotometer (Hitachi 220 S), specially designed diffusion cell apparatus (Ildam, Turkey), DSC apparatus (Shimadzu DT-40) dial thickness gauge (Ozaki). 2.3. Preparation
Acta Heluetiae 71 (1996) 205-212
of the formulations
White petrolatum as a hydrocarbon base, cold cream (USP) as an absorption base, hydrophilic ointment (USP) as a water washable base and Carbopol gel as a water-soluble base were selected. All formulations were prepared at 1, 3, 5, 7 and 10% concentrations of KP. White petrolatum ointment was suspension type which was prepared by geometric dilution of KP and melted base in the mortar. The other ointments were solution types which were prepared with the KP solution in water and triethanolamine mixture. Cold cream (USP XXI) and hydrophilic ointment (USP XXI) were prepared by some modifications of the formulations. (Babar et al., 1990; Muktadir et al., 1986; Ranman et al., 1990). 6% of triethanolamine was added to the formulations to dissolve KP in the bases. In addition, methyl and propyl parabens were added to cold cream for preservation. All the aqueous phase ingredients except KP and triethanolamine were dissolved in two-thirds of the water. Aqueous and oil phases were heated to 70 + 5°C separately and then the water phase was added to the oil phase with continuous stirring. The ointment was then cooled to approximately 40-50°C and KP was dissolved in one-third of the water and triethanolamine mixture was added and stirred till the ointment was cooled to room temperature. Carbopol gel base formulation and preparation is as follows: Carbopol940 1.5 g, triethanolamine 9 g, tiomersal 0.01 g, sodium metabisulfide 0.1 g, water qs to 100 g. Carbopol 940 was dispersed in two-third of the water for overnight. KP was dissolved in the remaining water with
2.4. In tiitro release studies Specially designed vertical-type glass diffusion cells with side-arm were used. Donor and receptor compartments were attached to each other by a separating Spectrapor 2 membrane. 1 g of ointment was placed in the donor compartment, the volume of the receptor compartment was 20 ml. pH 7.4 degassed phosphate buffer at 37’C was used as receptor solution. It was stirred magnetically at 600 rpm. The whole system was placed in a constant water bath at 37 k 1°C. Samples were withdrawn at 15, 30, 60, 90, 120, 150 and 180 minute intervals and analyzed for KP contents spectrophotometrically at a wavelength of 260 nm. The volume of the diffusion medium was kept constant by replacing the amount of sample with an equal volume of the diffusion medium. The concentration of KP in each sample was determined by spectrophotometrically. Blank ointment samples were used as a reference and all diffusion studies were the average of six experiments. Solubility of KP in the receptor solution was found as 880.5 mg/ml at 25°C. 2.5. Mathematical
evaluation
of release experiments
Mathematical analysis of the release of KP from different ointment bases was carried out using the Higuchi equation for solution type of ointments (Higuchi, 1962). When the amount of drug released was plotted against the square root of time, a straight line was obtained. The diffusion coefficient of drug (D) was calculated using the following equation: _ Dt
Q=2C,d
7T
Q: amount of drug released per unit area (mg/cm*); C,: initial concentration of drug in the ointment (mg/cm3); D: diffusion coefficient (cm*/& t: time (s). The permeability coefficient of KP was calculated using Eq. 2 (Babar et al., 1990): P=
4 A ’ C, . t
P: permeability coefficient (cm/s); released (mg); A: area of the diffusion C,: initial concentration of drug in the time (s). The partition coefficient of the KP
q: amount of drug membrane (cm*); base (mg/cm”); t: between
the oint-
Z. Gird
et al./ Pharmaceutics
ment base and receptor medium was calculated (Muktadir et al., 1986):
using Eq. 3
Acta Helcetiar
207
71 (1996) 205-212
2.7. Statistical
analysis
The differences in the results of in vitro and in vivo studies were evaluated using one-way ANOVA and Student’s t-test.
P.h
K,: partition coefficient; P : permeability coefficient (cm/s); D: diffusion coefficient (cm’/s); h: thickness of the membrane (cm>.
3. Results and discussion
2.6. Percutaneous
3.1. In L:itro release studies
absorption
in mice
Percutaneous absorption of KP was evaluated for its anti-inflammatory activity on carrageenan-induced paw edema in mice. The carrageenan-induced paw edema test is the most common test to evaluate the anti-inflammatory activity of NSAIDs (Chi and Jun, 1990; Winter et al., 1962). Although rats are used frequently (Chi and Jun, 1990; Gemmell et al., 1979; Graziani et al., 1987; Winter et al., 19621, mice may also be used (Schrier et al., 1987). The edema induced by the injection of carrageenan solution is measured using a plethysmometer (Graziani et al., 1987; Schrier et al., 1987; Winter et al., 1962) or a dial thickness gauge (Kasahara et al., 1985; Safak et al., 1992). Five young male mice weighing 20 & 5 g were used to evaluate the effect of ointment base and drug concentration on the percutaneous absorption of KP. The paw edema was measured via dial thickness gauge. 10 mg ointment was applied to the plantar surface of the left hind paw of the mice by gently rubbing 50 times with finger. After three hours, the paw was measured before carrageenan injection and recorded as control (E,). 0.01 ml of a 1% carrageenan solution was injected subplantarly into the same paw. Three hours after the carrageenan injection the paw was measured again and recorded as edema induced by carrageenan injection (E). The percent swelling of the paw was determined using the following equation (Chi and Jun, 1990). % Swelling
E - E, = x 100 4
% Inhibition
=
x 1-
I
100
% Swelling of control group
3.2. Effect of ointment bases The amount of KP released from different ointment bases at various concentrations are shown in Fig. 1 and Fig. 2. It can be seen that the amount of drug released were lowest from white petrolatum for all concentrations.
AMOUNT
OF RELEASED
(mg)
121
(4)
EC: control measuring; E: paw edema induced by carrageenan injection. Control group of the mice was treated only by the ointment base through the same procedure and the percent edema inhibition was determined using Eq. 5 which is the ratio of average swelling of the drug treated group to the average swelling of the control group (Chi and Jun, 1990). % Swelling of drug treated group
All formulations except white petrolatum ointment were solution types which were checked microscopically. The Higuchi equation (Higuchi, 1962) for solution type ointment was used to evaluate the data, and all graphs were obtained by plotting the amount of drug released against the square root of time.
1 (5)
0
2
4
6
8
10
12
14
Jt (mln)
-
1%
+-
3% W.P.
W.P.
+-
1% cc.
mxm 1% H.O.
-=
1% C.Q.
-6-
3% C.C.
-
+
3% C.Q.
3% H.O.
Fig. I. Effect of ointment bases on the release of ketoprofen W.P., white petrolatum: C.C.. cold cream: H.O., hydrophilic C.G., Carbopol 940 gel.
(IS. 38) ointment;
208
Z. Giirol et al. /Pharmaceutics AMOUNT
OF DRUG RELEASED
(rng)
Acta Heluetiae 71 (1996) 205-212 Table 1 Release kinetics of ketoprofen FORMULATIONS
251 20 /
I
0
d
2
4
6
Jt -
7% W.P.
+
+-
10% W.P. +-
8
10
x--x I
12
14
1% Cold cream 3 % Cold cream 5% Cold cream 7% Cold cream 10% Cold cream 1% Hydrophilic oint. 3% Hydrophilic oint. 5% Hydrophilic oint. 7% Hydrophilic oint. 10% Hydrophilic oint. 1% Carbopol940 gel 3% Carbopol940 gel 5% Carbopol940 gel 7% Carbopol940 gel 10% Carbopol940 gel
D X lo-’
P x 10-7
KXIO-2
4.43 7.02 7.48 9.53 10.07 223.40 375.57 396.27 445.65 484.79 5863.20 1947.60 703.88 364.89 176.31
6.67 9.06 9.96 12.30 12.91 54.23 67.47 69.39 72.80 75.47 228.00 131.10 78.78 59.09 41.81
34.41 29.50 30.43 29.50 29.31 5.55 4.1 I 4.00 3.73 3.56 0.89 1.54 2.56 3.70 5.42
D, diffusion coefficient (cm’/s). P, permeability coefficient (cm/s). K, partition coefficient.
(min)
7% CC.
+
7% H.O.
-
10% CC.
+-
10% H.O. -
Fig. 2. Effect of ointment bases on the release of ketoprofen Abbreviations as in Fig. 1.
7% C.Q. 10% C.G.
(7%, 10%).
The rank order of the drug release from different bases was found as: Carbopol gel > hydrophilic ointment > cold cream > white petrolatum for 1,3 and 5% of KP concentrations (Fig. 1). The results of 5% of KP ointment couldn’t be plotted in Fig. 1 for reasons of clarity of the graphic. The differences between the released amount of drug from the bases were statistically significant (p < 0.05). The rank order was modified slightly for 7 and 10% of KP as seen in Fig. 2. The statistically higher amount of drug was released from hydrophilic ointment for 7 and 10% of KP and Carbopol 940 gel was in the second order for these concentrations (p < 0.05). Cold cream and white petrolaturn ointments released KP in lower levels than the others. The calculated diffusion coefficient, permeability coefficient and partition coefficient of KP for all solution type ointments can be seen in Table 1. It can be seen that, the diffusion coefficient and permeability coefficient of KP are highest in the Carbopol 940 gel and lowest for the cold cream. The partition coefficient values show that the affinity of KP for the base is higher in the cold cream than the other bases.
was a good correlation between the concentration of KP and release rate for all bases except Carbopol 940 gel (Fig. 3). 3% concentration of KP was the optimum concentration for Carbopol 940 gel. It couldn’t be considered as a solubility limitation because the solubility of KP in the receptor solution was found as 880.5 mg/lOO ml at 25°C and it was 1100 mg/lOO ml at 37°C in the literature
RELEASE
RATE
1;
0
2
4
6
CONCENTRATION
3.3. Effect of ketoprofen
8
10
12
of KETOPROFEN
(%)
concentration
The release of KP from different ointment bases showed concentration dependency except Carbopol 940 gel. There
-
White
Petrolatum
-++
Hydrophilic
Olnt.
Fig. 3. Effect of drug concentration
+
Cold Cream
-
Carbopol
940 gel
on the release rate of ketoprofen.
14
209
Z. Giirol et al. / Pharmaceutics Acta Helvetiae 71 (1996) 205-212
Carbopol940 Gel (5 % W)
Ketoprofen
I
Carbopol940
Gel (7 % W)
Carbopot940 Gel
r
Carbopol940
Gel (1 % W’)
Carbopol940
(‘C)
of Carbopol 940 gel at different concentrations
(Bramanti et al., 1980). This could be explained by the hydrogen bond formation between KP, Carbopol 940 and triethanolamine. This interaction becomes stronger for high concentrations showing that KP is held tightly in the base. This situation can be seen in Fig. 4. Differential scanning calorimetric graphs of Carbopol 940 gel containing different percentage of KP show the specific peak of KP at 94°C which can be seen in 1% and 3% of KP, but it dissappears in higher KP concentrations. This may be the result of strong interaction between KP, carbopol 940 and triethanolamine. The released amounts for respective concentrations of
940 Gel (10 % a)
Gel (3 % KP)
TEMPERATURE Fig. 4. DSC thermogram
carbpol
of ketoprofen.
KP are significantly different from each other for white petrolatum, cold cream and hydrophilic ointment ( p < 0.05). This concentration dependency can be observed in Table 1 for the diffusion coefficient, permeability coefficient and partition coefficient values.
4. Percutaneous
absorption
of ketoprofen
4.1. Effect of ointment bases A 3% concentration of KP was used for all bases to evaluate the effect of ointment bases because it was the
2. Giirol et al. /Pharmaceutics
210 PERCENT
Acta Heluetiae 71 (1996) 205-212
INHIBITION
PERCENT
INHIBITION
,’ I /I’ i 60
60 -,’
50
50
40
40
>
,
30
20
10
10
k/’
0
L-
, __/ White
petr.
.i--
._~~ Cold cream
~ ,~ ~_i_
Hydr.oint.
FORMULATIONS Fig. 5. Effect ketoprofen.
of ointment
bases
, ~~~~~ /
Garb.940
0 Hyd.oint.3%
gel
absorption
of
optimum concentration for Carbopol 940 gel. The effect of bases on the percutaneous absorption of KP in mice paw was shown in Fig. 5. The percentage inhibition of KP was 10.96, 14.64, 55.76 and 56.62 for white petrolatum, cold cream, hydrophilic ointment and Carbopol gel, respectively. There was a difference between the white petrolatum and cold cream significantly ( p < 0.05). No significant difference was observed between the inhibition of hydrophilic ointment and Carbopol 940 gel (p > 0.05). But white petrolaturn and cold cream showed lower absorption than the others (p < 0.05). There was a good correlation between the in vitro and in vivo results with respect to the effect of ointment bases. Hydrophilic ointment and Carbopol 940 gel bases which caused the highest percutaneous absorption of KP were evaluated for the percent edema inhibition against carrageenan solution alone. While Carbopol gel base showed 1.43% of inhibition, hydrohpilic ointment base caused 69.07% of inhibition. The reason for the high edema inhibition value of hydrophilic ointment base was the high concentration of sodium lauryl sulfate in its formulation. 4.2. Effect of ketoprofen
Carb.gelB%
Carb.gellO%
FORMULATIONS
(3% KP)
on the percutaneous
Hyd.oint.lO%
concentration
3% and 10% concentrations of both hydrophilic ointment and Carbopol 940 gel formulations were compared to
Fig. 6. Effect of drug concentration ketoprofen.
on the percutaneous
absorption
of
investigate the effect of drug concentration. These concentrations were investigated because they were optimum and maximum concentrations in the bases which showed the maximum amount released in in vitro studies. The percentage inhibition of 3% of KP obtained from hydrophilic ointment and Carbopol 940 gel were found to be 55.76% and 56.62%, respectively. These values were 51.77% and 57.66% for 10% of KP in hydrophilic ointment and Carbopol 940 gel (Fig. 6). No significant difference was observed among the formulations (p > 0.05). It was seen that the percutaneous absorption of KP was not concentration-dependent contrary to the results of in vitro studies. As a result of in vitro and in vivo studies; it is obvious that the ointment bases effect the released amount and percutaneous absorption of KP strongly. In vitro data are in agreement with the in vivo data. Solution type ointments show the higher release rates and higher percutaneous absorption than the suspension type. It is in agreement with the results of the studies with benzocaine (Abd El Magid and Abd El Mohsen, 19901, chlorpheniramine maleate (Velissaratou and Papaieannou, 19891, piroxicam (Babar et al., 1990) and naproxen (Ranman et al., 1990). The general rank order of the percutaneous absorption of KP is found as follows: Carbopol gel 2 hydrophilic ointment > cold cream > white petrolatum. Bramanti et al.
Z. Giirol et al. / Pharmaceutics
(1980) obtained similar results with KP release from different type of ointment and gel bases. Drugs in ointment formulations should reach the skin surface in adequate amounts and sufficient rates in order to provide topical therapy. The release of drugs from ointment bases is a function of solubility of that agent in both base and its surrounding medium (Abd El Magid and Abd El Mohsen, 1990; Idson, 1975). If a drug is insoluble in the base, only the drug particles available at the surface of the base apparently dissolve into an aqueous medium (Idson, 1975). Since KP is not soluble practically in the white petrolatum, this formulation shows the lowest release and lowest absorption. Different amounts of KP release and absorption were obtained from cold cream and hydrophilic ointment formulations, although they are both solution type formulations. Solubilized KP was in the internal phase of cold cream while it was in the external phase of hydrophilic ointment. The oily external phase of the cold cream plays a barrier role around the water droplets and prevents the diffusion of the drug to the receptor phase. This may be the explanation of the lower amount released and absorbed of KP from cold cream than from hydrophilic ointment. The percutaneous absorption is also limited by skin and vehicle interaction besides the release rate of the drug. Although the released amount of KP from Carbopol 940 gel is higher than that of hydrophilic ointment for 3% of KP (Fig. 1), there was no significant difference between them in vivo (Fig. 5). It may be due to the interaction between the skin and hydrophilic ointment base. It was shown that hydrophilic ointment base caused 69.07% of inhibition while Carbopol 940 gel base showed 1.43% inhibition against the carrageenan solution. This was the result of the effect of sodium lauryl sulfate on the skin (Barry, 1988; Froebe et al., 1990; Walters, 1989). Although the effect of KP concentration was found to be significant in in vitro release studies for all bases except Carbopol 940 gel (Fig. 3), no significant difference could be observed in the percent edema inhibition of KP from different bases at 3% and 10% of drug concentration (Fig. 6). Generally, there is a positive correlation between drug concentration and release rate and percutaneous absorption of the drug due to the increase in the thermodynamic activity. The thermodynamic activity of a drug is related to its concentration in the vehicle. In different bases, thermodynamic activity of the drug increases linearly with concentration until it reaches the same limiting value which is the value of the saturated solution (Higuchi, 1982). Furthermore, the skin has a limited capacity for the transportation of some drugs (Akhter and Barry, 1985). Because of these reasons. percutaneous absorption of KP from 3% and 10% of hydrophilic ointment and Carbopol 940 gel formu-
Acta HelretiacJ 71 (199hi 205-212
211
lations was similar. The percent inhibition of KP with these two formulations was about 55% and was nearly the maximum response considered possible for a topical antiinflammatory drug using this method (Chi and Jun, 1990). 3% of KP was found as a critical concentration for the formulations investigated in this study. In conclusion, it can be stated tha;; There was a good correlation between the in vitro release studies from the cellophane membrane and carrageenan-induced paw edema test in mice. KP can penetrate to the skin easily. The best topical formulations for KP were both hydrophilic ointment and Carbopol gel at 3% drug concentration. The anti-inflammatory activity of the best formulations was about 557~.
References Abd El Magid, S.I.A. and Abd El Mohsen. M.G. (1990) In vitro and in viva studies of benzocaine release from ointments. S.T.P. Pharm.. 6, 380-387. Akhter, S.A. and Barry, B.W. (1985) Absorption through human skin of ibuprofen and flubiprofen: Effect of dose variation, deposited drug films, occlusion and the penetration enhancer N-methyl-?-pyrrolidone. J. Phann. Pharmacol., 37, 27-37. Babar. A., Solanki, U.D., Cutie, A.J. and Plakogiannis, F. (1990) Piroxicam release from dermatological bases: In vitro studies using cellulose membrane and hairless mouse skin. Drug Dev. Ind. Pharm., 16, 523-540. Barry, B.W. (1988) Action of skin penetration enhancers - the lipid protein partitioning theory. Int. J. Cosm. Sci., 10. 281-293. Bramanti. G.. Pisatura, G., Mazzi, G., Mura, P. and Papini. P. (1980) Release of drugs in vitro from different dermatological preparations. Boll. Chim. Form., 119, 738-744. Chi, S. and Jun, H.W. (1990) Anti-inflammatory activity of ketoprofen gel on carrageenan-induced paw edema in rats. J. Pharm. Sci.. 79. 974-977. Famaey, J.P. and Colinet, E. (1976) (Suppl.) A double-blind trial of ketoprofen in the treatment of osteoarthritis of the hip. Stand. J. Rheum., 14, 129-132. Fossgreen, J. (1976) (Suppl.) Ketoprofen - A survey of current publications. Stand. J. Rheum. 14. 1l-32. Froebe, C.L., Simion, F.A.. Rhein, L.D., Cagan. R.H. and Kligman (1990) A.. Stratum corneum lipid remove] by surfactants: Relation to in viva irritation. Dermatology. I8 I, 277-283. Gemmell, D.K.. Cottney, J. and Lewis, A.J. (1979) Comparative effects of drugs on four paw oedema models in rat. Agents Actions, 9, 107-116. Graziani, G.. Abbiati, G.A.. Dolfini, E.. Testa. R. and Velo, G.P. (1987) Pharmacokinetic, pharmacodynamic and toxicological properties of naproxen gel in laboratory animals. Pharm. Res., 43, 480-490. Higuchi, W.I. (1962) Analysis of data on the medicament release from ointments. J. Pharm. Sci., 51, 802-804. Higuchi, T. (1982) V. In vitro drug release from ointments and creams. In Brandau, R. and Reisen, P. (Eds.). Dermal and Transdermal Absorption. Wissenschaftliche Verlagsgesellschaft, Germany, pp. 90.
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Idson, B. (1975) Percutaneous absorption. J. Pharm. Sci., 64, 901-921. Kantor, T.G. (1986) Ketoprofen. A rewiew of its pharmacologic and clinical properties. Pharmacotherapy 6, 93-103. Kasahara, Y., Hikino, H., Tsurufuji, S., Watanabe, M.,, and Ohuchi, K. (1985) Anti-inflammatory actions of ephedrines in acute inflammations. Planta Med. 5 1, 325-33 1. Lussier, A., Camerlain, M., Menard, H., Myhal, D. and Wehner, S. (1976) (Suppl.) A double blind cross-over evaluation of ketoprofen and aspirin in rheumatoid arthritis. Stand. J. Rheum. 14, 99-104. Muktadir, A., Babar, A., Cutie, A. J. and Plakogiannis, F. M. (1986) Medicament release from ointment bases. III. Ibuprofen: In vitro release and in vivo absorption in rabbits. Drug Dev. Ind. Pharm., 12, 2521-2540. Peltola, P. (1976) (Suppl.) Long-term treatment with ketoprofen in rhumatoid arthritis. Stand. J. Rheum., 14, 115-l 17. Ranman, M.S., Babbar, A., Patel, N.K. and Plakogiannis, F.M. (1990) Medicament release from ointment bases: V. Naproxen in vitro release and in vivo percutaneous absorption in rabbits. Drug Dev. Ind. Pharm., 16, 651-672. Saxena, R.P. and Saxena, U. (1978) A comparative trial of ketoprofen and ibuprofen in patients with rheumatic disease. Curr. Med. Res. Opin., 5, 484-488.
Schrier, D.J., Moniot, S., Gluckman, M.I. and Gilbertsen, R.B. (1987) The topical anti-inflammatory effects of a topical preparation of meclofenamic acid on carrageenan-induced footpad swelling in mice. J. Pharm. Pharmacol., 39, 57-59. Safak. C., Erdogan, H., Palaska, E., Sunal, R. and Duru, S. (1992) Synthesis of 3-(2-pyridylethyl) benzoxazolinone derivatives: Potent analgesic and anti-inflammatory compounds inhibiting prostaglandin E?. J. Med. Chem. 35, 1296-1299. Velissaratou, A.S. and Papaieannou, G. (1989) In vitro release of chlorpheniramine maleate from ointment bases. Int. J. Pharm., 52, 83-86. Walters, K.A. (1989) Penetration enhancers and their use transdermal therapeutic systems. In Hadgraft, J. and Guy, R.H. (Eds.), Transderma1 Drug Delivery, Marcel Dekker, Inc., New York, pp. 197. Winter, C.A., Risley, E.A. and Nuss, G.V. (1962) Carrageenan-induced edema in hind paw of the rat as an assay for anti-inflammatory drugs. Proc. Sot. Exp. Med., 111, 544-547. Zutshi, D. and Mason, M. (1976) (Suppl.) Ketoprofen in rheumatoid arthritis: Its tolerance and therapeutic effect. Stand. J. Rheum., 14, 77-84.