Journal of Life Sciences and Technologies Vol. 2, No. 2, December 2014

Optimization of Physico-Chemical Parameters for the Extraction of Quercetin from Medicinal Herbs

D. V. Surya Prakash and Meena Vangalapati Centre for Biotechnology, Department of Chemical Engineering, Andhra University, Visakhapatnam-530003, Andhra Pradesh, India. Email: [email protected]; [email protected]

Abstract—Quercetin is a widely distributed in inflammatory activity [5], antimicrobial activity, nature. It found in the fruits of cytoprotective activity, antioxidant activity etc. (Haritaki), Phyllanthus emblica (Amla) and seeds of Dimocarpus longan, commonly known as the Longan Dimocarpus longan (Longan) species etc. It showed many and belongs to the family Sapindaceae. It is a tropical tree pharmacological activities anti-cancer activity, anti-oxidant that produces edible fruit. This fruit used as a traditional activity amd anti-inflammatory activities etc. The present studies on optimization of physico-chemical parameters like Chinese medicine for different treatments, such as effects of different solvents, soaking time, extraction time soothing nerves, relieving insomnia and promoting blood with hexane, particle size, different solvent percentages, metabolism. Longan pulps are tasty and rich in nutritious different volumes of hexane with ethanol and methanol as ingredients. The dried longan pulps are used as a tonic in solvents, pH and different weight dosages for the extraction traditional Chinese medicine to nurture heart and spleen, of Quercetin from Medicinal herbs. The highest Quercetin nourish blood, calm mind, add luster and beauty to the concentration for optimized conditions of Terminalia skin, and have therapeutic effects on heart palpitation, chebula fruit, Amla fruit and Longan seeds were 33.0µg/ml, 26.0µg/ml and 28.0µg/ml. insomnia, amnesia, and anxiety. Longan seeds have long

 been used as a folk medicine in China for treatment of

Index Terms—, Terminalia chebula fruit, pains, hernia, and skin diseases, acariasis, hernia, wound amla fruit, longan seed, aluminum chloride method. hemorrhages, eczema and scrofula etc. This seeds have been found to be a rich source of poly phenolic compounds such as , , chebulinic acid, I. INTRODUCTION , like quercetin [6] etc. Longan seeds extract showed strong scavenging Terminalia chebula species commonly called as Black activities of free radicals [7], inhibition on the myrobalan, Ink tree or Chebulic myrobalan. It is belongs proliferation of human colorectal carcinoma cells, to the family combretaceae. It is used for the treatment of antifatigue effect, hypoglycemic effects etc. number of diseases like cardio vascular diseases, arthritis, Quercetin is a flavonoid widely distributed in nature. paralysis, cancer, gout, ulcers, epilepsy etc. contains Foods rich in quercetin include black and green tea, tannins up to 30%, 3-5% [1], chebulinic apples, onion, red grapes, citrus fruit, tomato, broccoli acid 30%, tannic acid 20-40%, ellagic acid, 2,4-chebulyi– and other leafy green vegetables etc. It is water soluble β-D-gluco pyranose, gallic acid, ethyl gallate, flavonoids polyphenolic compound, which is extremely common and like rutins, quercetin [2] and luteolin etc. It shows wide spread in the plant kingdom as their glycosides. several pharmacological activities like anti-oxidant [3], Quercetin is the aglycone form of a number of other anti-cancar, anti-diabetic, anti-microbial etc. flavonoid glycosides, such as rutin and quercitrin, found Phyllanthus emblica, also known as Emblic, Emblic in citrus fruit, buckwheat and onions. It shows myrobalan or Indian gooseberry (Amla). It is belongs to pharmacological activities like anti-oxidant [8], anti- the family Phyllanthaceae. These fruits fruits are reputed cancer [9], anti-viral and anti-inflammatory activities [10] to contain high amounts of ascorbic acid (vitamin C), etc. Quercetin can be measured using Aluminum chloride such as emblicanin A (37%), emblicanin B colorimetric assay method. The present paper (33%), (12%) and (14%). It Optimization of Physico-Chemical Parameters for the also contains punicafolin, phyllanemblinin, gallic acid, Extraction of Quercetin from fruits of Terminalia chebula, ellagic acid, chebulinic acid, flavonoids like quercetin [4] Phyllanthus emblica (Amla) and seeds of Dimocarpus and kaempferol etc. It used as a rasayana (rejuvenative) longan species. to promote longevity, and traditionally to enhance digestion, treatconstipation, reduce fever, purify the blood, reduce cough, alleviate asthma, strengthen the heart, II. MATERIAL AND METHODS benefit the eyes, stimulate hair growth, enliven the body, and enhance intellect. It shows antitumor activity, anti- A. Chemicals and Reagents Aluminium chloride, Potassium acetate, Methanol,

Ethanol, Ethyl acetate, Hexane, Distilled water. Manuscript received May 10, 201 4; revised December 18, 2014.

2014 Engineering and Technology Publishing 90 doi: 10.12720/jolst.2.2.90-93 Journal of Life Sciences and Technologies Vol. 2, No. 2, December 2014

B. Collection of Plant Material The solvent, ethanol of Longan seed was shows best The dry fruits of Terminalia chebula, Amla and seeds results of Quercetin and it concentration was 3.0µg/ml. of Longan collected from local market in Visakhapatnam, The results were shown in Fig. 1. Andhra Pradesh, India. B. Effect of Soaking Time for Extraction of Quercetin C. Processing of the Plant Material The samples were incubated under proper conditions at These fruits and seeds were cut into small pieces and different time intervals viz., 1, 2, 3 and 4 days to powdered. The total powder done in to different mesh investigate the influence on extraction of Quercetin. It sizes from 44 to 120. The different size powders were was observed that Terminalia chebula, Amla fruits and stored in the air tight small covers. Longan seeds were shows second day was the best soaking time for the extraction of Quercetin and the D. Extract Preparation concentrations were 11.5µg/ml, 9.0µg/ml and 8.5µg/ml Weigh the different amounts of 7.4g of Terminalia respectively. The results were shown in Fig. 2. chebula fruit, 7.4g of Amla fruit and 7.0g of Longan seeds of powders and add methanol (60%) and ethanol (80%) in different flasks and makeup this solution up to 50 ml. Soak the solution for 2 days. After the soaking time filtrate the solution by using Whatman No.1 filter paper and heat the filtrate solution at 780c and 650c respectively. So that the solvent which is taken in the glass wear is evaporated and make up this solution up to 25 ml with distilled water to this solution add 25 ml of hexane solvent [11], mix the solution thoroughly. Pour the entire mixture in the separating funnel by using glass funnel. Incubate the solutions of ethanolic and methanolic extract for 1hr.

E. Determination of Quercetin by Colorimeter Figure. 2. Effect of soaking time for extraction of quercetin Aluminum chloride colorimetric assay method: 0.5ml of ethanolic and methanolic extracts taken in each test C. Effect of Extraction Time with Hexane for Extraction tube and 1.5 ml of methanol, 0.1 ml of 10% aluminium of Quercetin chloride, 0.1 ml of 1M potassium acetate and 2.8 ml of To investigate the influence of hexane on extraction of distilled water were added. The mixture was allowed to Quercetin different time intervals were taken viz., 1, 2, 3 stand for 30 min at room temperature. The absorance of and 4 hr. Solvent-Solvent extraction was done with the reaction mixture was measured at 415 nm using hexane as one of the solvent. The fruits of Terminalia colorimeter. The Quercetin was determined by using chebula, Amla and Longan seeds were shows that calibration curve. optimum concentrations were observed at first hour extraction time with hexane for extraction of Quercetin. III. RESULTS AND DISCUSSION The concentrations were 13.0µg/ml, 10.5µg/ml and 9.0µg/ml respectively. The results were shown in Fig. 3. A. Effect of Different Solvents for Extraction Different organic solvents such as methanol, ethanol, ethyl acetate and water were used to extract the optimumyield of Quercetin from these plants. The solvent, methanol of Terminalia chebula and Amla fruit were shows best results of Quercetin and its concentrations were 8.5µg/ml and 5.5µg/ml respectively.

Figure. 3. Effect of extraction time with hexane for extraction of quercetin

D. Effect of Different Particle size for the Extraction of Quercetin Different particle size viz., 354, 328, 250, 205, 149 and Figure. 1. Effect of different solvents for extraction of quercetin 125 microns were used to find out the optimum

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concentrations of Quercetin from these plant species. The achieved at 1:1 with methanol as a solvent of Terminalia present investigation suggests that the extraction of chebula fruit and Amla fruit and the optimum Quercetin at different particle sizes indicates that the concentrations were 19.5µg/ml, 16.5µg/ml. The optimum optimum particle size [12] was 125 microns of extraction was also recorded at 1:1 with ethanol of Terminalia chebula, Amla fruits and Longan seeds. The Longan seeds and the concentration was 17.5µg/ml. The optimum concentrations were 15.5µg/ml, 13.5µg/ml and observed results were shown Fig. 6. 15.0µg/ml. The results were shown in Fig. 4.

Figure. 6. Effect of different volumes of hexane for the extraction of Figure. 4. Effect of different particle size for the extraction of quercetin quercetin

Figure. 7. Effect of pH for the Extraction of quercetin Figure 5. Effect of different solvent percentages for the extraction of quercetin G. Effect of pH for the Extraction of Quercetin pH places a major role for the extraction of Quercetin. E. Effect of Different Solvent Percentages for the To optimize the pH for this process different pH extract Extraction of Quercetin samples viz., 4, 5, 6, 7 and 8. It was observed that the Percentage of the solvent is also plays a vital role for extraction of Quercetin of Terminalia chebula was found the extraction of components. The study on different to be optimum pH at 5.0 and optimum concentration was solvent (Ethanol, Methanol) percentages like 0%, 20%, 20.0µg/ml. The Amla fruit and Longan seeds shows 40%, 50%, 60%, 80% and 100% shows significant optimum pH at 6.0 and optimum concentrations were variations. Fig. 5 shows that optimum solvent percentages found to be 17.0µg/ml and 18.5µg/ml. The results were were found to be at 60% methanol of both Terminalia shown in Fig. 7. chebula & Amla fruit and 80% ethanol of Longan seeds H. Effect of Weight Dosages for the Extraction of for the extraction of Quercetin. The optimum Quercetin concentrations of 60% methanol were 18.5µg/ml & 15.5µg/ml and 80% ethanol was 16.5µg/ml. The results Different weight dosages were used to find out the were shown in Fig. 5. optimum concentrations of Quercetin of these plants. The present investigation suggests that the extraction of F. Effect of Different Volumes of Hexane for the Quercetin at different weight dosages indicates that the Extraction of Quercetin optimum weight dosages were 7.4 gm of Terminalia To determine the volume of hexane for the extraction chebula fruit, 7.4 gm of Amla fruit and 7.0 gm of Longan of Quercetin at the different volumes of hexane with seeds. The optimum concentrations were 33.0µg/ml, solvent (Ethanol, Methanol) were considered such as 1:1, 26.0µg/ml and 28.0µg/ml respectively. The results were 1:2 and 2:1. The optimum extraction of Quercetin was shown in Fig. 8.

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sustainer,” Journal of Applied Pharmaceutical Science, vol. 2, no. 1, pp. 246-249, December 2011. [5] M. Z. Asmawi, H. Kankaanranta, E. Moilanen, and H. Vapaatalo, “Anti-inflammatory activities of emblica officinalis gaertn leaf extracts,” Journal of Pharmacy and Pharmacology, vol. 45, no. 6, pp. 581-4, June 1993. [6] Y. F. Chen and L. Zhi, “Optimization of extraction conditions of flavonoids from longan seed,” Journal of Guangdong Chemical Industry, vol. 89, no. 2, 2009. [7] A. Panyathepa, T. Chewonarina, K. Taneyhillb, and U. Vinitketkumnuen, “Antioxidant and anti-matrix metalloproteinases activities of dried longan (Euphoria longana) seed extract,” Journal of Science Asia, vol. 39, pp. 12–18, January 2013. [8] H. Liu, L. Zhang, and S. Lu, “Evaluation of antioxidant and immunity activities of quercetin in isoproterenol-treated rats,” Journal of Molecules, vol. 17, no. 4, pp. 4281-4291, April 2012. [9] E. Sugantha Priya, K. Selvakumar, S. Bavithra, P. Elumalai, R. Figure 8. Effect of weight dosages for the extraction of quercetin Arunkumar, et al., “Anti-cancer activity of quercetin in neuroblastoma: An in vitro approach,” Journal of the Neurological Sciences, vol. 35, no. 2, pp. 163-70, February 2014. IV. CONCLUSION [10] U. J. Joshi, A. S. Gadge, P. D. Mello, R. Sinha, S. Srivastava, and G. Govil, “Anti-inflammatory, antioxidant and anticancer activity Experiments were performed for the optimization of of quercetin and its analogues,” International Journal of Research extraction of Quercetin from medicinal herbs like dry in Pharmaceutical and Biomedical Sciences, vol. 2, no. 4, pp. 1756-66, October 2011. fruits of Terminalia chebula, Amla and seeds of Longan [11] H. Walia, S. Kumar, and Saroj Arora, “Comparative antioxidant fruit. Quercetin estimation and optimize the physico- analysis of hexane extracts of terminalia chebula retz. prepared by chemical parameters by using Aluminum chloride maceration and sequential extraction method,” Journal of colorimetric assay method. The parameters like effects of Medicinal Plants Research, vol. 5, no. 13, pp. 2608-2616, July 2011. different solvents, soaking time, extraction time with [12] D. M. Anil and R. P. Nandini, “Development and validation of hexane, particle size, different solvent percentages, HPLC method for quantification of phytoconstituents in haritaki different volumes of hexane with ethanol and methanol as churna,” International journal of ChemTech research, vol. 3, no. 1, solvents, pH and different weight dosages for the pp. 329-336, March 2011. extraction of Quercetin concentrations were observed from the experimental work. The highest Quercetin Mr. D. V. Surya Prakash, Research scholar, Department of Chemical Engineering, Andhra concentration for optimized conditions of Terminalia chebula fruit, Amla fruit and Longan seeds were University College of Engineering (A), Andhra University, Visakhapatnam, Andhra 33.0µg/ml, 26.0µg/ml and 28.0µg/ml respectively. Pradesh, India. He has attended 10 more National and International conferences and

REFRRENCES published 15 more Papers published in National and International Reputed Journals, [1] D. V. Surya Prakash and V. Meena, “A review on chebulinic acid published 2monographs/Books. E-mail id: from medicinal herbs,” World Journal of Pharmaceutical [email protected]. Research, vol. 3, no. 6, pp. 2127-2139, August 2014. [2] A. Kumar, K. Lakshman, K. N. Jayaveera, S. N. and K. V. Satish, “Estimation of gallic acid, rutin and quercetin in Dr. Meena Vangalapati M.Tech., Ph.D, terminalia chebula by HPTLC,” Jordan Journal of Associate Professor, Department of Chemical Pharmaceutical Sciences, vol. 3, no. 1, pp. 63-68, January 2010. Engineering, Andhra University College of [3] S. Suchalatha and C. S. Devi, “Antioxidant activity of ethanolic Engineering (A), Andhra University, extract of terminalia chebula fruit against isoproterenol – induced Visakhapatnam, Andhra Pradesh, India. She oxidative stress in rats,” Indian Journal of Biochemistry and has 13 years of teaching experience and Biophysics, vol. 42, no. 4, pp. 246-249, August 2005. published 60 more Papers published in [4] E. Singh, S. Sharma, A. Pareek, J. Dwivedi, S. Yadav, and S. National and International Reputed Journals, Sharma, “Phytochemistry, traditional uses and cancer published 8 monographs/Books. chemopreventive activity of Amla (Phyllanthus emblica): The E-mail id: [email protected].

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