Chandra Subhash et al. IRJP 2011, 2 (12), 213-216

INTERNATIONAL RESEARCH JOURNAL OF PHARMACY, 2(12), 2011

INTERNATIONAL RESEARCH JOURNAL OF PHARMACY ISSN 2230 – 8407 Available online www.irjponline.com Research Article

BERBERIS ASIATICA FUTURE BASED EXCELLENT FRUIT IN NUTRITIONAL PROFILE,

ANTIMICROBIAL AND ANTIOXIDANT INGREDIENTS Saklani Sarla1, Chandra Subhash1*, Kandari Singh Alok2

1Department of pharmaceutical Sciences, H.N.B. Garhwal University (A Central University) Srinagar Garhwal, 246174, Uttarakhand, India 2Department of university Science instrumentation centre (USIC), H.N.B. Garhwal University (A Central University) Srinagar Garhwal,

246174, Uttarakhand, India

Article Received on: 17/10/11 Revised on: 20/11/11 Approved for publication: 09/12/11 *E-mail: subhashkothiyal@gmail.com ABSTRACT The aim of the present study was to calculate in vitro antimicrobial activity of Berberis asiatica fruit extracts against 13 Gram-positive and Gram-negative bacteria and fungus strains. The ethanolic fruit extracts of Berberis asiatica showed significant activity 15±1mm, 14±1mm and 13±1mm against Streptococcus pyogenes, Streptococcus aureus and Bacillus cereus against food poisoning bacteria and fungus, Development of a product with nutritional profile and secondary metabolites as a model, a wild edible fruit of Himalaya (Berberis asiatica) was screened. The fruits have been found to rich in nutrients such as crude protein1.3%, carbohydrates17.39%, crude fiber3.4%, ash content1.25% and minerals as calcium, magnesium, potassium and phosphorus (1.0, 8.4, 1.98 and 0.24 mg/100g) respectively, vit. A Vit. C and secondary metabolites were carried out by using standard analytical techniques. These analyses revealed that, the fruits contained higher value of fat, protein, fiber and minerals as compared to the cultivated fruits. 200 gm fruits contain sufficient amount of nutrients, required per day by a person. Key words: Antimicrobial, Nutritional profile, Antioxidants, Phenolics, and Flavonoids, INTRODUCTION A large portion of the world population, especially in. developing countries depends on the traditional system of medicine for a variety of disease. Several hundred genera are used medicinally and plants are vital sources for potent and powerful drugs1. Plants are rich in a wide variety of secondary metabolites such as tannins, alkaloids and flavonoids, of the phytochemical constituents found In vitro to have antimicrobial properties2. Many of the spices and herbs used today Kanyakumari coast, South West Coast of India have been valued for their antimicrobial effects and medicinal powers in addition their flavor and fragrance qualities3, 4&5. Man needs an appreciable amount of nutrients in his diets to perform various body functions and to lead a healthy life. The nutrients include protein, fat, carbohydrates, fiber, vitamins, and minerals. An average Indian of 60 kg body weight, doing moderate physical and mental work requires 60 gm protein, 20 gm fat, 28 mg iron, 40 mg vit. C 2875 k cal energy etc. in the daily diet6. The small berries including seeds are sweet, with a blend of acid. They are slightly bitter and the bitterness is due to the seeds. The taste and flavour of the fruits is amusing. Wild edible fruits, besides being important sources of minerals, fibre, and vitamins, provide essential nutrients for maintaining good health. The aim of the present study is to find out the health promoting potential of such fruits by determining their nutritional as well as medicinal value, to utilize the traditional knowledge for the palatability overall acceptability, and availability of the fruits. In addition, to utilize the native expertise for the collection, screening, identification, vernacular names, distribution, documentation, methods of harvesting and preparation of various useful indigenous products from the fruits. MATERIALS AND METHODS Material The fresh parts of fruit, of Berberis asiatica, was collected from adjoining area of Karanprayag city (Dist- Chamoli, Uttarakhand) in the month of May-June. The plant was authenticated by botanist Dr. R. D. Guar, Department of Botany; H. N. B. G. U. Srinagar Garhwal. Preparation of plant Extract The plant material was separated into its selected part fruit air dried ground to moderately fine powder and Soxhlet extracted with increasing polarity solvent (Petroleum ether, chloroform, ethyl

acetate, acetone, methanolic, and water7. Each extract was evaporated to dryness under reduce pressure using rotary evaporator. The coarse powder of fruit bark and root was subjected to successive hot continuous extraction with various solvent each time before extracting with next solvent the powdered material will be air dried (weight of crude extract 100gm). The various concentrated extracts were stored in air tight container for further studies. Antibacterial assay The disc diffusion assay methods were used to determine the growth inhibition of bacteria by plant extracts8, 9. Diluted bacterial culture (100μl) was spread over nutrient agar plates with a sterile glass L-rod. 10mg/ml and 50mg/ml of the each extracts were applied to each filter paper disc (Whatman No. 1, 5 mm diam.) and allowed to dry before being placed on the agar plate. Each extract was tested in triplicate (3 discs/ plate) and the plates were inoculated at 37°C for 24 h. After incubation, the diameter of inhibition zones was measured with a caliper. Antifungal assay The antifungal activity was tested by disc diffusion method10, 11. The Sabouraud dextrose agar plates were each similarly seeded with each fungal strain The 24 hrs. broth culture of each bacterium and 7 days inoculated fungus culture were used to seed sterile Sabouraud dextrose agar at 45°C respectively, and fungal plates were incubated at 25-28°C for 7 days after which diameter of zones of inhibition were measured. Each disc filled with extract. Nutritional & Mineral assay The number of water molecule is contain % of moisture, Pt. ether and hexane soluble part is called crude fat and the non soluble part of acid- base medium is called crude fibre (cellulose and lignin), and mineral estimated by flame photometry12-14. Quantification of Total Phenolics Content Total phenolics content in the methanolic extract of fruit part of selected species were estimated calorimetrically using the Folin-Ciocalteu calorimetric method15. Quantification of Total Flavonoids Content Flavonoids content in the methanolic extract of plant was determined by Aluminum chloride calorimetric method16. Methanolic extract of sample (0.5 ml) was diluted in distilled water (1.5 ml) and mixed with 10% Aluminum chloride (0.5 ml). In this mixture 1 M potassium acetate (0.1 ml) and distilled water (2.8 ml)

Chandra Subhash et al. IRJP 2011, 2 (12), 213-216

INTERNATIONAL RESEARCH JOURNAL OF PHARMACY, 2(12), 2011

was added and incubated at room temperature for 20 minutes. The absorbance of resulting reaction mixture was measured at 415 nm using UV-VIS spectrophotometer. Quantification of total flavonoids content was done on the basis of standard curve of quercetin prepared in 80% (v/v) methanol. Results were expressed in mg quercetin equivalent (QE) to per gram of fresh weight. Assay of Antioxidant Activity DPPH Traditional DPPH (1, 1-diphenyl-2- picrylhydrazyl) assay was carried out with minor modification. 100 ìM DPPH was dissolved in pure 80% ethanol. DPPH cation (3.0 ml) was mixed with sample extract (1ml) and kept in dark at room temperature for 20 minutes. Reduction in the absorbance at 520 nm was recorded by UV-VIS spectrophotometer. Results were expressed in mm ascorbic acid equivalent (AAE) per 100 g few of plant material RESULTS AND DISCUSSION Plants are important source of potentially useful structures for the development of new chemotherapeutic agents. The first step towards this goal is the in vitro antimicrobial activity assay. The results of antibacterial, antifungal, nutritional value and antioxidant activity, table 1, 2, 3, and 4, reveals that antibacterial, antifungal, nutritional, (antioxidant, phenolics, flavonoids) activity of fruit explants of was evaluated against ten bacterial and three fungal pathogenic strains Antibacterial and antifungal activity Berberis asiatica ethanolic fruit extract significant activity 15±1mm, 14±1mm and 13±1mm against Streptococcus pyogenes, Streptococcus aureus and Bacillus cereus against food poisoning bacteria and fungus, the order of the species based on total antibacterial activity is as follows: Streptococcus pyogenes ˃Streptococcus aureus ˃ Bacillus cereus. Nutritional value The level of nutrients such as crude protein1.3%, carbohydrates17.39%, crude fiber3.4%, ash content1.25% and minerals as calcium, magnesium, potassium and phosphorus (1.0, 8.4 & 1.98 and 0.24 mg/100g) respectively. Moisture (%): 65.20 ± 0.15 Total nitrogen (%): 0.52 ± 0.05 Crude fat (%): 0.80± 0.05 Soluble carbohydrates: 24.98± 0.16 Vit A mg/100g: 0.09± 0.02 Berberine mg/100g: 1.08± 0.10 Tannins: 0.64± 0.05 Acidity: 1.07± 0.02 Mg mg/100gm: 0.061± 0.08 P mg/100gm: 0.079 ± 0.04 Fe mg/100gm: 0.012± 0.02 Ash (%): 2.60 ± 0.05 Total protein (%): 3.30 ± 0.03 Crude fibre (%): 3.12 ± 0.15 Organic matter (%): 97.40± 0.16 Energy value /100 gm: 109± 0.15 Soluble solids: 18.90± 0.20 Pectin: 0.37± 0.04 Ca mg/100gm: 0.065 ± 0.05 K mg/100gm: 0.44± 0.20 N mg/100gm: 0.528± 0.04 Vit. C mg/100g: 6.90± 0.17 Phenolics mg/100g: 670± 0.12 Flavonoids mg/100g: 190.40± 0.52

Antioxidant activity: 03.20± 0.12 Phytochemical screening The phytochemical screening for the presence of glycosides, flavonoids, phenols, resin and tannins. However, alkaloids were absent. This analysis revealed that, the fruits contained higher value of fat, protein, fiber and minerals as compared to the cultivated fruits with apple and 200 gm fruits contain sufficient amount of nutrients, required per day by a person. CONCLUSION The in vitro antimicrobial studies present Berberis asiatica to have considerable efficacy against various pathogenic bacteria. The study provides a scientific basis for the use of the plant as folk medicine. The fruit of the plant is a good source of essential nutrients including minerals, carbohydrates, proteins and lipids. However, more advanced pharmacological and clinical studies would be required to investigate in vivo mechanism of nutraceuticals effects of this important wild plant. ACKNOWLEDGMENT We sincerely acknowledge the financial support granted by UCS&T/R&D/CHEM-16/09/10/6539/1, 06/01/2010 (UCOST) Dehra Dun to work on a project. The present research paper is the outcome of the same. REFERENCES

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4. De, A.K., 2004. Spices: Traditional Uses and Medicinal Properties. Daryaganj: Asian Books Pvt. Ltd., pp. vii-xvii.

5. Davidson, P.M., J. N. Sofos and A. L. Branen, 2005. Antimicrobials in food. 3 ed. CRC Press, Taylor and Francis Group. Boca Raton FL 33431, USA.

6. Gopalan, C., Rama Shastri, B.V., and Balasubramanian, S.C., (1996), Nutritive value of Indian Foods, National Institute of Nutrion Hyderabad. India.

7. Lin J, Opak War, and Geheeb-Keller M. 1999. Preliminary screening of some traditional Zulu medicinal plants for anti-inflammatory and antimicrobial activities. Journal of Ethnopharmacology, 68: 267–274.

8. Iennette E.H. 1985. Manual of clinical microbiology, 978–987. 4th edition. American Association for Microbiology, Washington.

9. Rosoanaivo and Ratsimanaga Urverg 1993. Biological evaluation of plants with reference to the Malagasy flora. Monograph for the IFs. NAPRECA Workshop on Bioassays.

10. Taylor, R.S.L., N.P. Manandhar, J.B. Hudson and G.H.N. Towers, 1995. Screening of selected medicinal plants of Nepal for antimicrobial activities. J. Ethnopharmacology., 546: 153-159.

11. Espinel Ingroff, A., A. Fothergill, J. Peter, M .G. Rinaldi, and T.J. Walsh. 2002. Testing conditions for determination of minimum fungicidal concentrations of new and established antifungal agents for Aspergillus spp.: NCCLS Collaborative Study. Journal of Clinical Microbiology 40:3204-3208.

12. Gaur, R. D., Flora of the district Garhwal North West Himalaya, 1st Ed. Transmedia Srinagar Garhwal, July-1999,

13. Dutta NK and Iyer SN. Anti-amoebic value of Berberine and kurchi alkaloids. Journal of the Indian Medical Association 1968, 50(8), 349-354.

14. Badoni S. Rawat M.S.M., Negi Y.S. Nutritional Composition of some Berberis species, J. Ind. J. of Hort. Pub.-1994,

15. Singleton, V. L., Rossi, J. A, 1965. Colorimetry of total phenolies with phosphomolybolic acid phosphotunguntic and reagent. American Journal of Enololgy and Viticulture, 16: 144-158.

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Table 1, Antibacterial activity of ten bacterial strains against Berberis asiatica plant extract. Disc size, 5 Mm, Inhibitory zone size ±1 Mm, Mm means (millimetres) and – indicate (NIZ) No inhibitory zone.

Bacterial Name

Petroleum ether Extract

Chloroform Extract

Ethyl acetate Extract

Acetone Extract

Methanol Extract

Water Extract

Genus /Species/Subspe. MTCC (Code)

10 Mg/ Ml

50 Mg/ ml

10 Mg /ml

50 Mg/ ml

10 Mg/ ml

50 Mg/ml

10 Mg/ ml

50 Mg/ ml

10 Mg/ ml

50 Mg/ ml

10 Mg/ ml

50 Mg/ ml

Bacillus cereus 1272 - - 6 7 7 6 7 8 13 - 9

Escherichia coli 729 - - 7 8 - 8 6 9 10 12 7 11

Enterobacter gergoviae 621 - - 6 7 - 7 7 8 - - 7 9

Klebsiella pneumonia 432 - - 8 9 7 8 - 9 11 14 - 11

Salmonella entericatyphim 98 - - 6 7 - 6 - 8 - 9 6 8

Shigella flexneri 1457 - 6 6 7 - 7 6 8 - 10 - 9

Staphyloccus aureus 902 - 7 7 8 6 8 7 9 11 14 - 10

Staphyloccus epidermidis 435 - - 6 7 - 9 7 9 - 10 - 10

Streptococcus pyogenes 1925 - - - 6 7 8 - 8 - 11 - 9

Escherichia coli 443 - 6 7

8 - 8 8 9 7 10 - 11

Table 2, Fungal activity of three fungal strains against Berberis asiatica plant extract. Disc size, 5 Mm, Inhibitory zone size ±1 Mm, Mm means (millimetres) and –

indicate (NIZ) No inhibitory zone.

Fungal Name

Petroleum ether Extract

Chloroform Extract

Ethyl acetate Extract

Acetone Extract

Methanol Extract

Water Extract

Genus /Species/Subspe.

MTCC (Code)

10 Mg/ml

50 Mg /ml

10 Mg /ml

50 Mg /ml

10 Mg /ml

50 Mg/ ml

10 Mg /ml

50 Mg /ml

10 Mg/ml

50 Mg/ml

10 Mg/ml

50 Mg/ ml

Candida albicans 3017 - - - 6 - -

- - 7 9 - 8

Aspergillus flavus 2798 - - - 6 - -

- - 6 8 - 7

Aspergillus parasiticus

2796 - 7 - 7 - -

- - 7 10 - 9

Figure 1.1 and 2.1, Antimicrobial activity of ten bacterial strains & three fungal strains against Berberis asiatica plant extract

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Fig. 1.1; Comparison of Berberis asiatica fruit and Apple for Nutrients value.

Fig.1.2; Comparison of per day intake of nutrients by Adults with the nutrients present in the fruits of Berberis asiatica.

.

Fig. 1.3; Comparison of per day intake of minerals by Adults with the mineral present in the fruits of Berberis asiatica

Source of support: Nil, Conflict of interest: None Declared