IN VITRO ANTIBACTERIAL AND ANTIOXIDANT POTENTIAL OF MEDICINAL PLANTS USED IN THE TREATMENT OF ACNE Research Article PRATIBHA NAND 1 , SUSHMA DRABU 1 , RAJINDER K GUPTA 2* 1 Maharaja Surajmal Institute of Pharmacy, Janakpuri, 2 University School of Biotechnology, Guru Gobind Singh Indraprastha University, New Delhi, India. Email: rkg67ap@yahoo.com Received: 12 Feb 2011, Revised and Accepted: 18 May 2011 ABSTRACT In the present study, petroleum ether, dichloromethane and methanolic extracts of three medicinal plants namely Camellia sinensis, Glycyrrhiza glabra and Calendula officinalis were analyzed for their antibacterial and antioxidant activity in different test systems. Highest zone of inhibition (≥15) mm was observed with methanolic extract of Camellia sinensis using disc diffusion method. Phytochemical analysis indicated that amongst nine test extracts, methanolic extract of Camellia sinensis had the highest total phenolic content (104.93 ± 1.630 mg GAE /g). Flavonoid (115.503 ± 2.984 mg RuE /g dry extract) and flavonol content (574.446 ± 7.94mg RuE /g dry extract) of methanolic extract of Glycyrrhiza glabra was also found to be superior among all the extracts. Antioxidant assays revealed highest DPPH scavenging (IC50=44.03 ± 1.784 µg/ml) and metal chelating (IC50=234.64 ± 5.467 µg/ml) effect in methanolic extracts of Camellia sinensis. Similarly, methanolic extract of Glycyrrhiza glabra exhibited highest radical scavenging activity (IC50=21.37 ± 1.422 µg/ml) when reacted with the ABTS . + . Keywords: Acne, Free radicals, Medicinal plants, Total phenolic, Total flavonoid, Total flavonol, DPPH, ABTS, Metal chelating. INTRODUCTION Acne vulgaris is the most common skin disorder of pilosebaceous unit, generally characterized by formation of seborrhea, comedone, inflammatory lesions and presence of bacteria namely Staphylococcus aureus, Staphylococcus epidermidis and Propionibacterium acnes in the follicular canal 1 . Propionibacterium acnes evokes mild local inflammation by producing neutrophil chemotactic factors. Consequently, neutrophils get attracted to the acne lesions and constantly release inflammatory mediators such as reactive oxygen species (ROS) 2 . Although oxygen is an important component for human beings, yet it can produce various ROS such as superoxide anion, hydrogen peroxide and hydroxyl radicals etc. Furthermore, ROS play a critical role in irritation and disruption of the integrity of the follicular epithelium and are responsible for the progression of inflammatory acne 3 . These toxic ROS can also act as messengers in the induction of several biological responses such as NF-kB, AP-1 and the generation of cytokines. These radicals are formed with the reduction of oxygen to water. Normally, the production of these radicals is slow and they are removed naturally by antioxidant enzymes like superoxide dismutase (SOD), catalase (CAT) and glucose-6-phosphate dehydrogenase (G6PD) existing in the cell, but due to the depletion of immune system and natural antioxidants in different ailments, it becomes necessary to use antioxidants as free radical scavengers for the removal of ROS to reduce cell damage that occurs during acne inflammation 4 . Drugs like tetracycline, erythromycin, minocycline and metronidazole are gaining more importance and are preferred over antibiotics 5 because of their antioxidant effect. Moreover, antibiotic resistant P. acnes has now become a critical problem worldwide 6 . Hence Antioxidants are extensively studied for their capacity to protect organism and cell from damage that are induced by oxidative stress 7 . There are a number of synthetic antioxidants like butylated hydroxy anisole, butylated hydroxy toluene, propyl gallate and gallic acid esters which are available but are suspected to cause negative health effects and are also unstable at elevated temperatures 8 . Hence, the objective of our research work was to investigate antibacterial and antioxidant potential of the following three medicinal plants viz: Camellia sinensis 9 , Glycyrrhiza glabra 10, 11 and Calendula officinalis 12, 13. MATERIALS AND METHODS Plant material, chemicals and reagents Fresh and dried plant materials were collected from medicinal gardens and authorized herbal stores in Delhi. Their botanical identities were determined and authenticated at the National Institute of Science Communication and Information Resources, New Delhi, India vide voucher specimen (NISCAIR/RHM/consult/2007- 08/936/120) and (NISCAIR/RHM/consult/2008-09/978/09). Clindamycin phosphate was procured from Sri Ram Institute of Industrial Research, New Delhi. All other chemicals used were of analytical grade and obtained from either Sigma-Aldrich or Merck. Preparation of extracts and phytochemical screening The air dried leaves, flowers, stolons, roots and seeds were pulverized and passed through sieve #10 and were used for extraction in soxhlet apparatus at room temperature. Sequential extraction of 200 g was carried out with solvents of increasing polarity i.e. petroleum ether (PE), dichloromethane (DCM) and methanol (ME) and the extracts were abbreviated as: Camellia sinensis (CSPE, CSDCM, CSME), Glycyrrhiza glabra (GGPE, GGDCM, GGME) and Calendula officinalis (COPE, CODCM, COME). The extracts were evaporated under vacuum conditions using a rotary evaporator and stored at 4 ˚C in air tight containers for further studies. The percentage yield was recorded. Preliminary phytochemical screening of the test extracts was carried out 14 . Antimicrobial screening of extracts Microorganisms and media Aerobic bacteria: Staphylococcus aureus (MTCC 96), Staphylococcus epidermidis (MTCC 2639) and anaerobic bacteria: Propionibacterium acnes (MTCC *1951) were obtained from the Microbial Type Culture Collection Centre, Institute of Microbial Technology, Chandigarh. Fresh cultures of the isolates of aerobic and anaerobic bacteria were suspended in nutrient broth and reinforced clostridium medium respectively. S. aureus and S. epidermidis cultures were incubated for 24 h at 37°C and 30°C, respectively. P. acnes culture was incubated in an anaerobic chamber at 37°C consisting of 10% CO2, 10% H2 and 80% N2 for 48 h. Disc diffusion method Antibacterial activity of extracts was tested using agar disc diffusion method 15. 100 µl of fresh culture suspension of test bacteria was evenly spread on nutrient agar and reinforced clostridial agar plates. The concentration of cultures was 5x10 5 CFU/ml. For screening, 6 mm diameter filter paper disc, impregnated with 20 µl of extract solution equivalent to 0.2 mg of extract, was placed on the surface of inoculated media agar plates. Incubation was done at 37°C or 30°C for 24 h and 48 h depending upon the type of bacteria under optimum conditions. Clear zones of inhibition were measured in mm and Clindamycin (10 µg/disc) was used as positive control. International Journal of Pharmacy and Pharmaceutical Sciences ISSN- 0975-1491 Vol 4, Issue 1, 2012 A A c c a a d d e e m mi i c c S Sc c i i e e n nc c e e s s