Pharmaceutical Development and Technology, 5(4), 455–464 (2000)

RESEARCH ARTICLE

Topical Liposomal Gel of Tretinoin forthe Treatment of Acne: Research andClinical Implications

Vandana B. Patel,1 Ambikanandan Misra,1 andYogesh S. Marfatia2

1Pharmacy Department, Faculty of Technology and Engineering, Kalabhavan,M. S. University of Baroda, Baroda 390 001, India2Skin-VD Department, S.S.G. Hospital, Faculty of Medicine, M. S. University ofBaroda, Baroda 390 001, India

Received April 6, 1999; Accepted January 13, 2000

ABSTRACT

An attempt was made to pharmaceutically develop a topical liposomal tretinoin (TRE) gel and clini-cally evaluate the developed formulation for the treatment of acne in patients. Liposomes of TREwere prepared using the lipid film hydration technique and the entrapment efficiency of TRE in lipo-somes was optimized to 79.96%. The drug retention in liposomes and in liposomal TRE gel (Carbopol934 gel base) studied at three storage conditions indicated maximum drug retention at refrigerationtemperature. For liposomal TRE gel, reduced drug leakage was observed as compared to that ofliposomes at all three storage conditions. Diffusion studies of plain TRE gel and liposomal TRE gelsuggested prolongation (3.4 times reduction in flux value) of drug diffusion and almost two- foldincrease in skin drug retention after liposomal encapsulation of drug. A comparative double-blindclinical study of the developed liposomal TRE gel, carried out on 30 acne patients over a period of3 months, demonstrated significant enhancement (about 1.5-fold) in drug efficacy. More remarkableimprovement was observed in the treatment of comedones, where the mean percent reduction in lesionsincreased from 62.36% for plain TRE gel to 94.17% for liposomal TRE gel. Erythema and irritationassociated with the use of plain TRE gel was reduced considerably with the use of liposomal TREgel. The findings of this investigation therefore underscore potential utility of commercialization ofliposomal TRE gel in the treatment of acne.KEY WORDS: Acne; Clinical; Gel; In vitro; Liposomes; Topical; Tretinoin.

Address correspondence to Dr. Ambikanandan Misra, Professor of Pharmaceutics. Fax: (0265) 423898. E-mail: misraan@hotmail.com

455

Copyright 2000 by Marcel Dekker, Inc. www.dekker.com

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456 Patel, Misra, and Marfatia

INTRODUCTION

Tretinoin (TRE), also known as all-trans-retinoic acid,the acid form of vitamin A, is reported to be effectivetopically in the treatment of acne (1,2). TRE decreasesand prevents lesions by increasing epithelial cell turnoverand decreasing horny cell adherence (3). Compliance inthe use of TRE is often adversely affected by several un-wanted side effects that include heightened susceptibilityto sunlight, severe local erythema and peeling at the site ofapplication (4), occasional worsening of the acne dur-ing the first 2 weeks of application (5), and lack of full im-pact of treatment for the first 6–8 weeks of application (6).

Liposomes, minute spherical vesicles consisting oflipid bilayers, have been suggested for use as a suitable‘‘carrier’’ or ‘‘localizer’’ for various drugs used in top-ical administration. According to many investigators(7,8), the liposomal drug formulations are able to reduceside effects and incompatibilities and enhance the accu-mulation of drug at the administration site. Hence, an at-tempt was made to formulate liposomal TRE gel withthe view to reduce local irritation and erythema of TREtherapy and to improve therapeutic response, which inturn may improve patient compliance and therapeutic ef-ficacy. Note that although liposomes are promising forpotential use in pharmacotherapy, clinical studies are stillscarce.

Therefore, in the present investigation, the liposomesof TRE were prepared and optimized for the drug entrap-ment efficiency. Liposomal TRE gel was prepared usingcarbopol 934 gel base and was subjected to comparativein vitro drug diffusion studies along with the plain TREgel. An attempt was also made to clinically evaluate theformulation against the plain TRE gel in acne patients.

MATERIALS AND METHODS

Materials

TRE (Sigma Chemical Company, St. Louis, MO), eggphosphatidyl choline (PC) (Centre for Biochemical Tech-nology, New Delhi, India), cholesterol A.R. (CHOL)(S.D. Fine Chemicals Ltd., Boisar, India), α-tocopherol(α-Toco) (E. Merck, Mumbai), carbopol 934 (B.F. Good-rich, Cleveland, OH), triethanolamine (E. Merck), andphenyl mercuric nitrate (BDH, Mumbai, India) were pur-chased and used as such without further purification. Allother chemicals used were of analytical grade. Acetatebuffer, pH 5.0 (ionic strength 0.261), was prepared asdescribed in the Indian Pharmacopoeia 1996 (9).

Preparation of Skin

Human cadaver skin (HCS), from abdominal portionof the bodies of both sexes (aged between 25 and 35years), was obtained from autopsy at the Faculty of Medi-cine, M. S. University of Baroda. The skin was washedthoroughly with water and subcutaneous fat was re-moved. The skin was then stored at 24°C. Full thicknessHCS membrane was prepared by shaving the skin,punching out a disk of approximately 2.5 cm2 area andslicing to 500 µm thickness using a Dermatome 7(Davis). These slices were hydrated with diffusion me-dium [pH 5.0 acetate buffer with 20% poly(ethylene gly-col) (PEG) 400] for 24 hr at room temperature prior touse.

Preparation of Liposomes

Liposomes were prepared essentially according to thelipid film hydration technique reported by Bangham etal. (10). Films of TRE, PC, and CHOL containing α-Toco [1% w/w of PC taken (11)] in the ratios shown inTable 1 were prepared in a 100-ml round-bottomedquickfit flask by rotary flash evaporation. The contentsdissolved in methylene chloride (5 ml) were evaporatedto thin, dry film at 180 rpm under vacuum (10 in. of Hg)and at the temperature of 25 6 2°C. The dried film washydrated with 5 ml distilled water for 2 hr at room tem-perature, sonicated for size reduction for three 2-min in-tervals using an ice bath (about 0°C) with a probe sonica-tor (model RP 120, Ralsonics, India), and hydrated for24 hr at refrigerated temperature (2–8°C). The liposomes(MLVs) thus formed were then separated from free drugby ultracentrifugation (C-24, Remi, India) at 3.3 3 106

3 g and washed three times with purified water.

Preparation of Gels

Carbopol 934 (1% w/w) was dusted onto distilled wa-ter containing 0.001% phenyl mercuric nitrate while themixture was stirred, and left overnight to settle for 24hr. Triethanolamine (0.5 ml) was then added with gentlestirring to avoid inclusion of air (12). The pH of the gelbase thus obtained was 5.6.

Plain TRE gel and liposomal TRE gel were preparedby incorporating plain TRE or liposomal TRE pellet, re-spectively, into carbopol gel base by trituration so as toobtain 0.025% w/w TRE-containing gels.

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Tretinoin Liposomal Gel for Acne 457

Table 1

Effect of Composition of Liposomes on Drug Entrapment Efficiencies

EntrapmentPC/CHOL TRE/PC/CHOL Efficiencya (%) Free TREb (%)

Batch Molar Ratio Molar Ratio Mean (6SD) Mean (6SD)

TRE 1 2.47:1 1:3.84:1.55 31.07 (0.66) 64.03 (0.21)TRE 2 4.94:1 1:7.68:1.55 40.06 (0.04) 54.92 (0.19)TRE 3 10.19:1 1:15.84:1.55 79.96 (0.26) 15.77 (0.29)TRE 4 5.10:1 1:15.84:3.11 55.04 (0.25) 38.67 (0.37)TRE 5 10.19:1 1:7.92:0.78 51.01 (0.08) 42.98 (0.38)TRE 6 4.94:1 1:3.07:0.62 30.97 (0.14) 63.21 (0.13)

a Percent of added TRE actually entrapped into liposomes [mean (6SD) of six determinations, n 5 6].b Percent of added TRE left unentrapped [Mean (6SD) of six determinations, n 5 6].

Physical Analysis

All of the batches were examined under a polarizingmicroscope (model 580, Leitz Inc., Wetzlar, Germany)to determine the shape and lamellarity of liposomes. Ves-icle size distribution of the liposomes was determined bylight scattering based on laser diffraction using the Mas-tersizer (model S, ver. 2.15, Malvern, Worcestershire,U.K.).

Chemical Analysis

Drug entrapment efficiency of the liposomes, ex-pressed as percent of the added drug actually entrappedinto liposomes, was determined by estimation of TRE byHPLC (13). The liposomes were treated with methanol–triton X100 (95:5, v:v) and filtered on a 0.45-µm mem-brane filter. TRE was quantitatively determined by HPLCat 365 nm on a Hypersil ODS (5 µm) column using phos-phate buffer (pH 3.0)/tetrahydrofuran (46.5:53.5) as mo-bile phase (1 ml/min). The unentrapped drug was ana-lyzed in the supernatant after separating liposomes byultracentrifugation technique. The results are recorded inTable 1.

PC was estimated by the Stewart assay (14) andCHOL was estimated using the method of Zlatkis et al.(15) by measuring the absorption at 485 and 550 nm,respectively, against the reagent blank.

Drug Retention in Liposomes

The potential liposomal batch, TRE 3, and its liposo-mal gel were sealed in 30-ml amber glass vials and storedat refrigerated temperature (2–8°C), room temperature(25 6 2°C), and body temperature (37°C). The sample

from either batch at each temperature was taken at defi-nite time intervals, centrifuged at 3.3 3 106 3 g for 10min, then the supernatant was analyzed for free drug con-centration. The results calculated in terms of percent TREretained in liposomes are shown in Fig. 1(a, b, and c).

Diffusion Studies

A Franz static diffusion cell with a 2.271-cm2 diffu-sion area was used for the diffusion studies. Each studywas conducted using dermatomed and prepared HCSfrom the same donor. The gel (0.2 g plain TRE gel andliposomal TRE gel) was applied to the skin and spreadover the 2.271-cm2 test area with the tip of a pipet. Thedermal surface of the skin was perfused with receptorphase solution (pH 5.0 buffer with 20% PEG 400 as acosolvent) and the donor surface remained unoccluded.The test system was equilibrated with a water bath set at37°C for the specified duration of exposure, and the re-ceptor phase was agitated by magnetic stirrer.

Serial sampling of the dermal compartment was per-formed at specific times (1, 2, 4, 6, 24, 48, and 72 hr) byremoving the contents of the receptor and refilling it withfresh medium. The amount of TRE diffused was analyzed(13) at all sampling time points. The results are given inTable 2. The mean flux values of both the gels were alsocalculated and are shown in Table 2. At the end of thestudy (72 hr), the residual drug remaining on the surfacewas removed by washing the surface three times with themedium. The washings were analyzed for residual TRE(13). The skin was digested with 10 ml of methylenechloride overnight at 40°C and analyzed for the drug con-tent (13) in the skin. The results of skin retention studiesare given in Table 2.

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Figure 1. Mean percent TRE retention (n 5 6) in liposomes and liposomal TRE gel at different storage conditions. Key: (a)Refrigerated temperature (2–8°C); (b) room temperature (25 6 2°C); (c) body temperature (37°C). * 5 p , 0.05. Series 1: TREliposomes (batch TRE 3); series 2: liposomal TRE gel (batch TRE 3).

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Tretinoin Liposomal Gel for Acne 459

Table 2

Diffusion of TRE Across Human Cadaver Skin (HCS) and TRE Retention in HCS

Plain TRE Gel Liposomal TRE Gel

Time (hr) Time (sec1/2) Aa Bb Aa Bb

1 60.00 0.0088 0.04 0.00255 0.012(0.003) (0.016) (0.001) (0.005)

2 84.85 0.0176 0.08 0.00523 0.024(0.006) (0.027) (0.002) (0.01)

4 120.00 0.022 0.10 0.00643 0.029(0.006) (0.029) (0.003) (0.012)

6 146.94 0.044 0.20 0.0131 0.060(0.013) (0.057) (0.004) (0.020)

24 293.94 0.176 0.80 0.0521 0.24(0.020) (0.094) (0.017) (0.077)

48 415.69 0.207 0.94 0.0621 0.28(0.031) (0.145) (0.023) (0.102)

72 509.12 0.264 1.20 0.0781 0.35(0.047) (0.213) (0.024) (0.108)

Mean flux values (µg/min) (6SD) 2.269 3 1024 6.67 3 1025

(5.799 3 1025) (2.26 3 1025)

% TRE Retention in HCS After 72 hrc

Liposomal TRE GelPlain TRE Gel

Human skin 33.55 65.74Mean (6 SD) (1.74) (1.60)

Washing medium 61.33 29.95Mean (6SD) (1.01) (1.27)

a Mean (6SD) cumulative amount released (µg/cm2).b Mean (6SD) cumulative dose released (%).c Mean of six experiments conducted on six different days (n 5 6).

Clinical Studies

The study involved 30 human volunteers aged 16–28years (18 female and 12 male) with mild to moderateacne. The study was carried out in Skin-VD Departmentof S. S. G. Hospital, attached to Faculty of Medicine, M.S. University of Baroda, India, for a period of 3 months.All lesions located on the facial area were observed dur-ing the course of study. Few patients had been undergo-ing treatment with different drugs prior to entering thestudy and had been suffering from acne for 4 months to6 years before the period of study. All previous treat-ments were discontinued at least 30 days prior to thesestudies.

In a double-blind design, a dose of 0.5 g of both thegels was applied to the left (plain TRE gel) and right(liposomal TRE gel) side of the face, once daily at bed-time, to each patient after the face was cleaned with warm

water. The treatment was continued for 12 weeks and theprogress was observed on a weekly basis by a physician-in-charge in terms of percent reduction in separate typeand total number of skin lesions (Table 3). In addition,the overall improvement in skin condition was evaluatedon a monthly basis by an independent clinical physicianusing evaluation indices (16) (results are summarized inTable 4). The skin condition of patients was also evalu-ated in terms of adverse symptoms of therapy such aserythema, itching, burning, scaling, and irritation, and aresummarized in Fig. 2.

Data Analysis

Experiment Design and Comparison

Reproducibility of all the experiments performed wasconfirmed by repeating the experiment at least six timeson six different days, and the ANOVA technique of sta-

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Table 3

Mean Percent Reduction (SD) in Separate Type and Total Number of Skin Lesionsa Observed inAcne Patients During a Comparative Double-Blind Clinical Evaluation of Plain TRE Gel and

Liposomal TRE Gel

Comedones Papules Pustules TotalTime(Weeks) A B A B A B A B

04 18.57 34.17 11.00 18.49 5.00 11.33 13.62 23.44(10.43) (6.67) (15.98) (8.33) (10.17) (10.08) (8.44) (5.10)

06 27.00 53.33 14.89 30.95 06.67 16.00 19.17 36.91(7.03) (4.79) (16.02) (13.37) (11.24) (8.14) (8.31) (7.99)

08 40.97 75.83 25.56 41.27 10.56 23.33 29.82 51.68(8.67) (4.32) (10.66) (12.23) (13.22) (7.58) (7.26) (7.47)

10 50.95 83.75 35.55 52.94 21.67 36.00 39.31 62.03(4.08) (8.27) (8.23) (4.27) (13.77) (8.14) (4.18) (6.61)

12 62.36 94.17 49.33 69.76 36.39 55.33 53.50 76.64(5.39) (5.99) (8.28) (5.56) (8.04) (11.37) (3.30) (5.58)

a Study was conducted on 30 acne patients.A: Plain TRE gel.B: Liposomal TRE gel.

tistical analysis was used to compare the data of each setof experiments. The differences were considered signifi-cant at p , 0.05, unless otherwise specified.

The data of percent drug retained in liposomes werecalculated by subtracting the amount of drug leakagefrom the amount of drug initially present. The percentdrug retained in liposomes with respect to time (weeks)for a period of 3 months is shown in Fig. 1(a, b, and c).

The mean cumulative drug release, Q (µg/cm2) (n 5

Table 4

Improvement in Separate Type of Skin Lesions

Evaluation Indicesa

Comedones Mean (6SD) Papules Mean (6SD) Pustules Mean (6SD)

Formulation 4 weeks 8 weeks 12 weeks 4 weeks 8 weeks 12 weeks 4 weeks 8 weeks 12 weeks

Plain TRE gel 2.17 1.13 0.63 2.47 1.70 1.00 2.80 2.50 1.10(0.39) (0.35) (0.30) (0.73) (0.47) (0.00) (0.41) (0.63) (0.31)

Liposomal TRE gel 1.33 0.067 0.00 2.10 1.03 0.00 1.57 1.83 1.07(0.48) (0.25) (0.00) (0.31) (0.18) (0.00) (0.50) (0.38) (0.25)

a Initial index in all cases was approximately 3.0 5 Excellent (.66% improvement).1 5 Very good (33–66% improvement).2 5 Good (,33% improvement).3 5 No change.The sum of all individual patients’ indices was calculated and divided by the number of patients (n 5 30).

6) across the membrane was calculated at each samplingtime point (Table 2) and the data of Q vs. square root oftime (sec1/2) were subjected to linear regression analysisto determine the relationship between these two variables(r). The mean flux values were calculated from receptorfluids collected at the specified times using the equation(17), J 5 V (dc/dt), where J is flux of the drug acrossthe membrane, V is volume of the receptor compartment,and (dc/dt) is rate of change of concentration.

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Figure 2. Evaluation of skin condition for adverse symptoms of therapy. Key: *1 5 high, 2 5 less, 3 5 no change. (A) PlainTRE gel; (B) liposomal TRE gel (batch TRE 3).

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462 Patel, Misra, and Marfatia

The skin retention of the drug determined after thediffusion study was expressed as percent of the applieddose retained within the skin.

Clinical efficacy of plain TRE gel and liposomal TREgel was expressed in terms of percent reduction in sepa-rate type and total number of skin lesions. The skin le-sions consisted of different manifestations of acne, suchas comedones, papules, and pustules. The number of eachtype of lesion and total number of lesions were countedinitially and after application of either formulation atspecified time intervals on the same surface area (4 cm2)marked with a paper template by the physician-in-charge.The results, recorded as the percent reduction in separatetype and total number of skin lesions, are presented inTable 3. The therapeutic efficacy of both the gels wasgraded using evaluation indices. The initial index in allcases was approximately 3. The grading of therapeuticefficacy was expressed as excellent, .66 % improvement(0), very good, 33–66% improvement (1), good, ,33%improvement (2), and no change (3). The sum of all indi-vidual patients’ indices was calculated and divided by thenumber of patients (n 5 30). The results are recordedin Table 4. The adverse responses in terms of erythema,itching, burning, scaling, and irritation were graded ashigh (1), less (2), and no change (3), as shown in Fig. 2.The data shown in Fig. 2 are weighted means calculatedby multiplying the degree of reaction (1, 2, or 3) by thenumber of patients responding to that degree out of 30patients, then dividing the value obtained by total numberof patients.

RESULTS AND DISCUSSION

Preparation of Liposomes

The lipid film hydration technique of liposome prepa-ration used in this study resulted in MLV liposomes asconfirmed by microscopy and light-scattering laser dif-fraction of samples. Coevaporation of the lipid and TREfrom methylene chloride solvent was found to produceliposomes with maximum drug entrapment. This is con-sistent with reported work on liposome preparation, sug-gesting that coevaporation of lipid and lipophilic drugsfrom organic solvent generates liposomes with the high-est drug incorporation (18,19). Hence, the lipid film hy-dration technique was used for the preparation of TREliposomes. α-Toco was added to prolong the characteris-tic induction phase of auto-oxidation and thus the shelflife of the liposomes (11,20). The prepared MLVsshowed a normal size distribution pattern, the size rang-ing from 1.2 to 6.8 µm. The microscopic examination

of liposomes indicated that the liposomes prepared aremultilamellar and spherical in nature.

Six batches of liposomes were prepared with varyingTRE/PC/CHOL ratio, as shown in Table 1. The resultsof mean entrapment efficiency (n 5 6) were comparedstatistically using ANOVA technique. The differenceswere considered significant at p , 0.05.

Influence of Formulation Components

The composition of liposomes and the drug entrap-ment efficiencies in prepared liposomes are summarizedin Table 1. Increasing the PC concentration with the sameCHOL and TRE concentration (TRE 1, TRE 2, TRE 3)led to increased entrapment efficiency ranging from31.07 6 0.66% to 79.96 6 0.26% of dissolved TRE. Thisincrease in entrapment efficiency may be due to the in-crease in drug-specific amphiphilic properties of lipidmembrane. Increasing the CHOL concentration with theconstant PC and TRE concentration (TRE 3, TRE 4) re-sulted in a significant decrease in drug entrapment effi-ciency, which decreased from 79.96 6 0.26% to 55.046 0.25%, respectively. Increasing the CHOL beyond acertain concentration can disrupt the regular linear struc-ture of the liposomal membrane, thereby reducing the en-trapment efficiencies (21). Preparations with increasingTRE concentration were produced in order to investigatethe influence of solute concentration on the entrapmentefficiency of liposomes. As shown in Table 1, a markeddecrease in entrapment efficiency with increasing TREconcentration from 40.06 6 0.04% to 30.97 6 0.14%(TRE 2 to TRE 6) and from 79.96 6 0.26% to 51.01 60.08% (TRE 3 to TRE 5) was observed. This is in agree-ment with the findings of Schneider et al. (22). This maybe due to the disruption of the liposome membrane bythe higher drug concentration, resulting in poor drug en-trapment.

Drug Retention in Liposomes

The percent TRE retained in the liposomes was calcu-lated from the amount of drug leaked out from liposomesover a period of time under different storage conditions.The results are shown in Fig. 1(a, b, and c). For lipo-somes, maximum stability was observed at refrigerationtemperature, where leakage was minimal, because only2–4 % of the drug leaked out after 90 days. At roomtemperature, the liposomes were stable for 20 days, afterwhich about 17% leakage of drug was observed; after 90days, almost 28% of the drug had leaked out. At bodytemperature (37°C), about 75% drug leaked out within

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Tretinoin Liposomal Gel for Acne 463

20 days of storage, and all of the drug had leaked outafter 90 days.

For liposomal TRE gel, reduced leakage was observedas compared to that of liposomes at all the temperatureconditions. A statistically significant difference (p ,0.05) was observed at room temperature wherein theleakage of drug was decreased from about 28 to 5% after90 days.

In Vitro Diffusion Studies

The in vitro permeation of TRE was calculated interms of the mean cumulative amount diffused (Q, µg/cm2) at each sampling time point during the 72-hr study.The results are shown in Table 2. It is evident from theresults that cumulative permeation of TRE was signifi-cantly greater from plain TRE gel than from liposomalTRE gel (p , 0.05) at all time points. This agrees withthe findings of many investigators (23,24), i.e., liposomalentrapment of TRE significantly prolongs the drug re-lease across the HCS. The value of correlation coeffi-cient, r, 0.988 for both the gels, suggests a linear relation-ship between Q and t1/2, and suggests that the releasefollows the pattern delineated in Higuchi’s diffusion-con-trolled model (25). The mean flux values (Table 2) of2.269 3 1024 µg/min for plain TRE gel and 6.67 3 1025

µg/min for liposomal TRE gel show a significantly (p ,0.05) higher flux (almost 3.4 times) of TRE from plainTRE gel. This further confirms the reduction in the rateof diffusion of the drug across HCS after liposomal en-trapment.

Liposomal encapsulation of TRE has been reported toenhance the drug retention in the skin (8,23,26). Hence,the skin retention of TRE was studied. The procedureused for washing the skin surface was found to be effec-tive in removing .99% of the residual drug (27). Massbalance data from deposition studies account for high re-covery of the applied drug dose. In this investigation, li-posomal TRE gel produced significantly higher deposi-tion of TRE (p , 0.05) in the skin (65.74 6 1.60%) thandid plain TRE gel (33.55 6 1.74%).

Clinical Evaluation

A comparative double-blind clinical evaluation ofplain TRE gel and liposomal TRE gel was carried out for3 months on 30 patients suffering from mild to moderateacne. The percent reduction in separate types and totalnumber of skin lesions were observed on a weekly basis.The results, summarized in Table 3, reveal that liposomalTRE gel significantly improved the therapeutic response

of TRE at all evaluation time intervals, compared to theplain TRE gel (p , 0.05). A 1.5-fold improvement wasnoticed with liposomal TRE gel. The overall improve-ment in patients’ skin condition, observed on monthlybasis, is presented as evaluation indices (16) in Table 4.The results indicate that all of the manifestations of thedisease were not equally responsive to TRE therapy.Comedones were most effectively treated, followed bypapules and pustules. Better response to comedonesmight be attributed to the predominant comedolytic activ-ity of TRE, wherein TRE reserves follicular retention hy-perkeratosis and allows the keratin plugs of microcomed-ones to be expelled. However, the anti-inflammatoryactivity required to combat papules and pustules is notexhibited by TRE to a significant extent, which may ex-plain its reduced action on papules and pustules (6).

The skin condition of patients was examined for theadverse symptoms of therapy at the end of 2, 4, 6, 8, 10,and 12 weeks. The results recorded in Fig. 2 reveal thatall the adverse symptoms, except scaling, were remark-ably decreased with the use of liposomal TRE gel as com-pared to the plain TRE gel (p , 0.05). This significantdecrease in adverse symptoms may improve patient com-pliance.

The findings of this investigation have conclusivelydemonstrated an improvement in therapeutic responseand a marked reduction in adverse symptoms after treat-ment with liposome-entrapped TRE compared to theplain TRE in the treatment of patients with acne. How-ever, the role of liposomal TRE gel of this investigationcan only be confirmed after clinical evaluation of theproduct with a large number of patients, with special fo-cus on the adverse symptoms of therapy.

ACKNOWLEDGMENTS

Financial assistance provided by C.S.I.R., New Delhi,is gratefully acknowledged.

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