Formulation and Evaluation of a Poly Herbal Anti-acne gel

 

Chandrasekar R.¹*, G. Satheesh   Kumar²

¹Department of Pharmacognosy, Seven Hills College of Pharmacy, Tirupati, Chitoor AP India

²Department of Pharmaceutical Chemistry, Seven Hills College of Pharmacy, Tirupati, Chitoor AP India

 

ABSTRACT:

Aim: Acne a common skin disease in adolescence is a bacterial skin disorder found in many teenagers. Herbal remedies are used in ancient system of medicine they have natural healing properties, they are used in various acute and chronic skin diseases, which include eczema, contact dermatitis, psoriasis, leprosy, and other major skin diseases. Methods: The present research was undertaken to formulate a herbal gel containing Aloevera and turmeric extract. Aloevera and turmeric both these plants possess potential anti-inflammatory immunomodulatory and anti acne properties. Aloevera and turmeric contain antioxidants, scavenging of free radicals. Aloevera leaf was washed with water the gel was separated and collected from the leaf and stored in refrigerator for further studies. Turmeric (curcuma longa) was washed and dried in room temperature and size reduced to fine powder and stored in plastic container in desiccator for further studies. Gels were prepared with gelling agents and with incorporation of excipients and ingredients containing plant extracts. Results: Different gels were prepared with different concentrations of gelling agents. The gel was evaluated for different physicochemical properties which include colour, appearance, consistency, washability, pH, spreadibility, extrudability, viscosity, homogeneity, etc.  The prepared gel did not show any phase separation it was stable and did not show any irritation on skin. The prepared gels were free from microbial contamination. Conclusion: Both these plants have enhanced efficacy and potency and can be safe and effective against many diseases. These two plants have fewer side effects and toxicity and they have enhanced acne healing and other properties. Results shows prepared gel has promising effect on the anti acne process

 

 

 

INTRODUCTION:

Skin diseases are common nowadays and 80% of people rely on herbal remedies. Herbal cosmetics are gaining importance in this current scenario. Recent trends in the development of new formulations in the treatment of skin diseases are gaining importance in curing chronic skin diseases.  Acne is a common inflammatory skin disorder affecting adolescence 85% of teenagers. It is most commonly found in women than in men. Acne mostly affects the puberty stage it is most common in the age of 18 to 25. This disorder is frequently less in adults in the age range of 35-40 years. Acne vulgaris is an inflammatory disorder of sebaceous glands and is prevalent in adolescence. This condition may be painful with redness and inflammation and sometimes pus may be formed. This condition may be due to exposure of the skin to environmental hazards like dust and pollution, consumption of more oily foods, increased production of sebum in the sebaceous glands, due to food habits, etc. acne can be characteristic lesions, inflammatory papules, pustules, nodules and cysts, which may lead to scarring and pigmentary change. Acne may occur also due to hormonal changes in the body. Acne vulgaris is a common inflammatory skin condition. Nearly 90% of teenagers have acne, and half of them continue to experience symptoms as adults. By age 40 years, 1% of men and 5% of women still have lesions. Recent analyses show an increasing prevalence of acne in children, perhaps because of pubertal onset.[1]

 

The objective of the study was to prepare a poly herbal gel containing aloevera and turmeric with incorporation of excipients. Aloevera belonging to the (Liliaceae) family is a perennial succulent plant. This plant has been known as “the healing plant”. Aloevera has been used in traditional medicine since ancient times in several centuries for millennia it has been demonstrated that aloevera has immunomodulatory and growth promoting activities. Aloevera is a plant which has more efficacy and potency against many skin diseases with less side effects and toxicity. The gel present in the plant can be directly applied on the skin to cure folklore claims. 

 

Turmeric Curcuma longa belonging to Zingiberaceae family is a spice that has received much attention from both the medical/scientific fields as well as from the culinary use. Turmeric is a rhizomatous herbaceous perennial plant (Curcuma longa) of the ginger family [2]. The medicinal properties of turmeric have been known for thousands of years. However, the ability to determine the exact mechanism(s) of action and to determine the bioactive components present in the plant. [3] Curcumin is the main natural polyphenol found in the rhizome of Curcuma longa (turmeric) and in others Curcuma spp. [4]. Curcuma longa has been traditionally used in Asian countries as a medical herb due to its antioxidant, anti-inflammatory [5], antimutagenic, antimicrobial [6], wound healing and anticancer properties [7].

 

The aim of the study is to formulate a herbal gel of aloevera and turmeric extract containing different concentrations of gelling agents and to investigate the effects of topical application of gel containing aloevera and turmeric extract on the healing of acne. Various physicochemical parameters of the gel that influences the properties of gel are also studied.

 

MATERIALS AND METHODS:

Collection of plant material:

Aloevera plant was obtained from Herbal Garden, Seven Hills College of Pharmacy, Tirupati. Turmeric Curcuma longa was procured from departmental stores, Tirupati.

 

Chemicals and Reagents:

PEG 400 and Carbopol 940 were procured from (Sigma Aldrich Pvt Ltd., Mumbai, India), calcium acetate, triethanol amine, glycerin, methyl paraben, Propyl Paraben, was procured from SD Fine chemicals, ltd., Mumbai, India.

 

Instruments and Equipments:

Double Beam UV Spectrophotometer-Analytical technologies Limited, Model 212R RI, Centrifuge-R-8C REMI Instruments, Digital pH meter Systronics, Mumbai, Brookfield Viscometer Servewell Pvt. Ltd. Model Number. LVDVE, SUPERFIT ROTAVAP, Model-PBU-6. Servewell Instruments Pvt. Ltd. Stability Chamber REMI SC-19PLUS.

 

Extraction of plant material:

The leaves of Aloevera were collected, washed with water and were cut transversely into pieces. The Aloevera gel was obtained from the centre of the parenchyma part of the plant leaf of Aloevera. The epidermis was removed and the inner gel-like pulp in the center of the leaf was separated with a spoon, minced, and homogenized in a mixer. Turmeric rhizome (Curcuma longa) was washed and dried and size reduced to fine powder. The fine powder was passed through sieve no 80. Turmeric powder was subjected to maceration with alcohol in a flask for 3 days with intermittent shaking, the extract was filtered. The collected extract was concentrated using a rotary evaporator to get a semisolid mass, the extract was stored in a desiccator for further studies.

 

Preparation of gel:

For the preparation of gel accurately weighed quantity of gelling agents were dissolved in water, and then triethanolamine was added drop by drop with constant stirring till pH was neutralized and gel was formed. Triethanolamine was added slowly to the dispersion with continuous stirring until a stiff gel was formed. Then the measured quantity of aqueous extract of Aloevera and turmeric were added to gelling agent and mixed with continuous stirring for 30 mins. Glycerine (humectants) was added as moistening agent, Propyl paraben and methyl paraben (preservatives) were added in required quantities dissolved in water and added slowly with continuous stirring until a homogenous gel was formed, until a semisolid consistency was obtained. The consistency was checked every time to improve the viscosity of the preparation. Volume was made with water and stirred continuously till a uniform gel was obtained. [8]:

Table 1 Formulation of anti acne gel

 

Table 2 Physicochemical Parameters

Gels	Clarity	pH	Spreadibility (cm)	Extrudability (gm/cm2)	Viscosity (cps)
G 1	+	6.64	6.3	18.1	5826
G 2	++	6.82	6.5	18.5	5874
G 3	+++	7.22	8.6	20.3	5754
G 4	++	7.34	8.8	22.3	5516

Turbid: +, clear: ++, very clear (glassy): +++.

 

Evaluation study of Gels:

Clarity:

The clarity of formulation was determined by visual inspection under black and white background and it is graded as follows; turbid: +, clear: ++, very clear (glassy): +++.

 

Measurement of pH:

The pH of various gel formulations was determined by using digital pH meter. One gram of gel was dissolved in 100 ml distilled water and stored for two hours. The measurement of pH of each formulation was done in triplicate and average values were calculated.

 

Drug content:

1 g of the prepared gel was mixed with 100ml of suitable solvent. Aliquots of different concentration were prepared by suitable dilutions after filtering the stock solution and absorbance was measured. Drug content was calculated using the equation, which was obtained by linear regression analysis of calibration curve.

 

Viscosity study:

The measurement of viscosity of the prepared gel was done with a Brookfield Viscometer. The gels were rotated at 0.3, 0.6 and 1.5 rotations per minute. At each speed, the corresponding dial reading was noted. The viscosity of the gel was obtained by multiplication of the dial reading with factor given in the Brookefield Viscometer catalogues.

 

Spreadability:

It indicates the extent of area to which gel readily spreads on application to skin or affected part. The therapeutic potency of a formulation also depends upon its spreading value. Spreadability is expressed in terms of time in seconds taken by two slides to slip off from gel which is placed in between the slides under the direction of certain load. Lesser the time taken for the separation of two slides, better the spreadability. It is calculated by using the formula:

 

S = M. L / T

Where,

S = Spreadability.

M = Weight tide to upper slide.

L = Length moved on the glass slide.

T = Time taken to separate the slide completely from each other

 

Extrudability study:

After the gels were set in the container, the formulations were filled in the collapsible tubes. The extrudability of the formulation was determined in terms of weight in grams required to extrude a 0.5 cm. ribbon of gel in 10 second.

 

Homogeneity:

After the gels have been set in the container, all developed gels were tested for homogeneity by visual inspection. They were tested for their appearance and presence of any aggregates.

 

Grittiness:

All the formulations were evaluated microscopically for the presence of any appreciable particulate matter which was seen under light microscope. Hence obviously the gel preparation fulfils the requirement of freedom from particular matter and from grittiness as desired for any topical preparation. [9, 10]

 

Pharmacological studies:

Animals:

Wistar rats of either sex with an average weight of 150-200g were obtained from Sri Venkateswara Enterprises Registration No.: 237/99/CPCSEA Bangalore. The animals were housed in clean cages placed in a well ventilated house. They were acclimatized to the animal house condition for seven days during which they were allowed free access to commercial pelleted rat chow. All experimental procedures were performed in compliance with international policies governing the Institutional Animal Ethical Committee for the treatment of experimental animals.

Experimental animals:

The animals were acclimatized to standard laboratory conditions (temperature: 25 ± 5°C), humidity (55 ± 5%) and maintained on a 12-h light: 12-h dark cycle. They were provided with regular rat chow and drinking water and libitum. The experimental protocols were approved by the Institutional Animal Ethics Committee CPCSEA Reg. No. (1995/PO/RE/S/17/CPCSEA)

 

In Vivo skin irritation study:

Animal:

 

Required   :     Albino rats of either sex (18)

Weight      :     150-200 gm

Groups      :     Three

Study        :     Primary irritancy level DRAIZE             scoring method

 

Table 3 Draize Evaluation of Dermal Reactions

Type of Skin Irritation		Scores
No erythema	No edema	0
Very slight erythema	Very slight edema	1
Well-defined erythema	Slight edema	2
Moderate to severe erythema	Moderate edema	3
Severe erythema	Severe edema	4

 

Table 4 Evaluation of primary irritation index

Index	Evaluation
0.00	No Irritation
1.00-1.99	Slight Irritation
2.00-2.99	Mild Irritation
3.00-5.99	Moderate Irritation
6.00-8.00	Severe Irritation

 

Table 5 Animal grouping

Groups	Treatment	Score
		Day 1	Day 3
Group I	Control	0	0
Group II	Gel without drug	0	0
Group III	Drug loaded gel	0	0.5

 

Skin irritation study:

Albino rats (150-200 g) of either sex were used for testing of skin irritation. The animals were divided into 3 groups containing 6 animals in each group. Group 1 was (control), group 2 (gel without drug) and group 3 (drug loaded gel). The animals were maintained on standard animal feed and had free access to water. The animals were kept under standard conditions. Hair was shaved from back of Albino rats and area of 2 cm² mark was done on both the sides, one side served as control while the other side was test. Gel was applied (50 mg / Albino rats) twice a day for 3 days and the site was observed for any sensitivity, edema, and erythema. [11]

 

Stability:

The stability studies were carried out for all the gel formulations. Here, by subjecting the product to a temperature of 4° C, 25°C and 40°C for 4 weeks, syneresis was also observed. After this, the gel was exposed to ambient room temperature and liquid exudate separation was noted. [11]

 

Microbial enumeration test and absence of specified microorganism

The microbial stability of the cosmetic formulations was evaluated for microbial contamination test. [12, 13]:

Bacterial Count:

The culture media was prepared and autoclaved at 125°C for 20 minutes then 20 ml of the culture medium was poured into a sterile petridish. 0.2g of the formulation was placed in the center of each petridish, and the plates were incubated at 37°C or at 25°C for 3 days according to the inoculated microorganisms. After the incubation period, plates were taken out and checked for microbial growth, which is an indication of contamination.

 

Yeast and Mould count:

Sample was dissolved in phosphate buffer pH 7.2, shaken well and the volume was made up to 100ml. 1 ml of dilution was pipetted out into a sterile petridish, and 30ml of the soyabean casein digest agar was poured, and mixed well. The agar was solidified and incubated at 30^o C to 35^o C for 48 to 72 hrs for Bacterial count. To the specimen of the sample fluid lactose medium was added and volume was made up to 100 ml, incubated for 24 hrs at 37 ^o C for E. coli. Incubate at 30^o to 35^o and observed for 48 to 72 hrs for presence of bacteria.

 

Figure No 1 Bacterial Count, Yeast and Mould count

 

 

Test for E. coli

Similarly 1 ml of dilution was pipetted out into a sterile petridish and 30 ml of sabour and dextrose agar was poured and mixed solidified inverted and incubated at 20 ^o C to 25 ^o C for 5 days for yeast. The contamination test was performed using a variety of microorganisms namely Bacteria and Yeast. After the incubation period, the plates were taken out and checked for microbial growth by comparing it with the control. No microbial growth was observed in the formulation. The obtained results confirmed that there was no contamination in the formulation.

 

Figure No 2 Test for E. coli, Control for E. Coli

 

RESULTS AND DISCUSSION:

 

Figure 3 pH of the gel formulation

 

 

Figure 4 Viscosity of the formulation

 

Figure 5 Spreadability of the gel formulation

 

Table No. 6 Spectrophotometric test

Concentration	Absorbance
0.01µg/ml	0.060
0.02µg/ml	0.090
0.03µg/ml	0.135
0.04µg/ml	0.169
0.05µg/ml	0.233

 

Figure 6 Linearity curve for gel formulation

 

Figure 7 Ultraviolet Spectroscopy of gel Formulation

 

 

 

 

Table 7 Stability Studies

Turbid: +, clear: ++, very clear (glassy): +++.

 

DISCUSSION:

Different gels were prepared in order to select the best gelling agent. Various gels were prepared containing extracts and excipients containing polymers, viscosity enhancers, etc. The clarity, consistency and viscosity of the gels were enumerated by using various instruments like Brookfield viscometer, pH Meter, and different methods to confirm the stability of the preparations like clarity, pH, spreadability, extrudability, spectrophotometric studies, skin irritation studies and microbial contamination tests. These gels were subjected to stability studies in a stability chamber to ensure the stability of the gels for 4 weeks. [14]

 

The gels with normal pH, viscosity, consistency, clarity, spreadability, extrudability, were selected based on the stability of the preparation.  Gel 1 showed phase separation and was very thin with low consistency resulted in liquefaction it was rejected. Gel 2 showed low viscosity problem with handling of gel and non uniform distribution of gel. Gel 3 showed uniform thickness smooth and clear with normal viscosity and good consistency and considered best. The spreadibility and extrudability values were good for gel 3. Gel 4 was very thick in consistency high viscosity, was more sticky that could not be properly spread out and Syneresis was observed. Thus, Gel 3 was selected as the optimized concentration of gelling agent. [15]

 

The pH of the formulation was determined to ensure that the formulation can be used without the risk of irritancy to the skin. The pH was found to be 6.6 for gel which was very near to the neutral pH, thus the formulation can be used without the risk of irritancy on the skin. This also indicated that the selected ingredients of the formulation did not alter the pH of the formulation.

 

The Spreadability of formulations was found to decrease with increase in the concentration of gelling agent. The value of Spreadability for optimized gel was found to be 8.6 cm indicating that the gel is easily spreadable by small amount of shear. The results indicated that the formulation can be applied easily without being runoff. This assures that the formulation maintain a good wet contact time when applied to the site of application.

 

The microbial contamination studies confirmed that no microbial growth was observed in the formulation. The obtained results confirmed that there was no contamination in the formulation. The skin irritation test was performed on albino wistar rats since there was no irritation erythema or edema after 4 days the studies confirmed that the gel was safe for use on skin.

 

CONCLUSION:

This study aimed at developing a poly herbal gel for anti-acne treatment using extracts of Aloevera and Curcuma longa in an aqueous based gel system. Four formulations of the gel were prepared by varying the proportions of gelling agents and evaluated for their physicochemical properties, like pH, spreadability, viscosity, and skin irritation study and microbial contamination tests. Based on these tests, formulation G3 was selected as the best formulation. The microbial enumeration studies of all the formulations demonstrated better microbial activity against various microorganisms stood competitive to the standard formulation. It was concluded that the present research might hopefully bring advancement in the treatment of acnes using herbs as well as in developing poly herbal formulations for safe and effective management of diseases.

 

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Received on 26.12.2019            Accepted on 31.01.2020           

Accepted on 21.02.2020              ©A&V Publications all right reserved