Biotransformation based Pharmacophores from Plant Extract of Picrorhiza kurroa Royle Ex Benth

Brajeshwar Singh; Somal Gupta

doi:10.55544/jrasb.1.1.7

Authors

Brajeshwar Singh Department of Microbiology, Faculty of Basic Sciences, SKUAST-Jammu, INDIA.
Somal Gupta Department of Microbiology, Faculty of Basic Sciences, SKUAST-Jammu, INDIA.

DOI:

https://doi.org/10.55544/jrasb.1.1.7

Keywords:

Fermentation, Picrohiza kurroa, total Phenolic content, Biotransformation, total flavonoid content

Abstract

An investigation entitled “Optimization of fermentation parameters to enhance biotransformation based pharmacophores from plant extract of Picrorhiza kurroa Royle ex Benth” was conducted in the Division of Microbiology, Sher-e-Kashmir University of Agricultural Sciences to optimize different fermentation parameters to enhance the polyphenols in the plant Picrorhiza kurroa Royle ex Benth collected from the Galhar region of District Kishtwar of UT Jammu & Kashmir. Four fermentation parameters were optimized during the experiment i.e. temperature, pH, inoculum load and incubation time with Saccharomyces cerevisiae and Lactobacillus acidophilus. The experimental results revealed that among Saccharomyces cerevisiae and Lactobacillus acidophilus, Saccharomyces performed better in all the conditions. Saccharomyces showed the maximum increase in phenolics and flavonoids. It showed the max increase at 30⁰C (92.10 to 127.04mg GAE/g dw) temperature and at pH of 5.5(92.43 to 128.96mg GAE/g dw) When incubated for 48h. Also, it was observed that increasing incubation time has an increasing effect on total phenolic content and flavonoid content but prolonged fermentation results in the decrease of both. In case of Saccharomyces, decrease in phenolics was observed after 48h (136.77 to 100.05mg GAE/g dw) but in case of Lactobacillus, decrease was seen after 72h, which could be due to the oxidation of phenols by bacteria. Further LCMS analysis also revealed an increase in the quantity of compounds as there is increase in the peak area. Thus, this study suggests optimum fermentation parameters and microorganism for enhancing bioactive compounds in Picrorhiza kurroa Royle ex Benth. The results suggest that Saccharomyces cerevisiae is the best microorganism for fermentation of Picrorhiza kurroa Royle ex Benth and the optimum conditions for incubation is 30⁰C temperature at pH5.5 for 48h; that gives improved nutraceutical values and enhance the bioavailability of these bioactive compounds.

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References

Ali, M., Sultana, S. and Mir, S.R. 2017. Chemical constituents from the roots of Picrorhiza kurroa royle ex benth. International journal of pharmacy and pharmaceutical research. 9:25-35.

Beatriz, A., Acosta-Estrada., Janet , A., Gutierrez ,U., Sergio ,O. and Serna, S. 2014. Bound phenolics in foods. Food Chemistry.152:46-55.

Devi, K., Samant , S.S., Puri, S., Paul, S. and Dutt, S. 2019. Diversity distribution pattern and indigenous uses of medicinal plants in kanawar wildlife sanctuary of Himachal Pradesh ,north western Himalayas India .Journal of conservation biology ,photon .117:172-219.

Feng, Y., Zhang, M., Fan, K. and Mujumdar, A. S. 2017.Effects of drying method on quality of fermented plant extract powder. Drying Technology, 36:1913-1919.

Hossain, M.A., Weli, A.M. and Ahmed.S.H.I. 2019. Comparison of total phenols, flavonoids and antioxidant activity of various crude extracts of Hyoscymaus gallagheri traditionally used for the treatment of epilepsy .Clinical Phytoscience .5:20.

Joshi, R.K., Satyal, P. and Setzer, W.N. 2016. Himalayan aromatic medicinal plants, their ethnopharmacology, volatile Phytochemistry and biological activities .Medicines .3(6).

Krupashree, K., Kumar, K.H., Rachitha, P., Jayashree, G.V. and Khanum, F. 2014. Chemical composition, antioxidant and macromolecule damage protective effects of Picrorhiza kurroa Royle ex Benth. S Afr J Bot. 94:249–254.

Lone, P.A., Bhardwaj, A.K. and Bahar, F.A. 2013. Traditional knowledge on healing properties of plants in bandipora district of jammu and Kashmir .International journal of recent scientific research .4: 1755-1756.

Malik, Z.A., Bhat, J.A., Ballabha, R.M., Bussmann, R.W. and Bhatt, A.B. 2015. Ethno medicinal plants traditionally used in health care practices by inhabitants of western Himalaya’s .Journal of ethnopharmacology .172:133-144.

Masood, M., Muhammad, A., Qureshi, R., Sidra, S., Muhammad, S., Amjad., Qureshi, H. and Tahir, Z. 2015. Picrorhiza kurroa: An enthopharmacologically important plant species of Himalayan region. Pure and Applied Biology.4 (3):407-417.

Myers, N., Mittermeier, R., Fonseca, G. and Kent, J. 2000. Biodiversity hotspot for conservative priorities. Nature, 403:853-858.

Prachyakij, P., Charernjiratrakul, W., and Kantachote, D., 2008. Improvement in the quality of a fermented seaweed beverage using an antiyeast starter of Lactobacillus plantarum DW3 and partial sterilization. World Journal of Microbiology and Biotechnology, 1713-1720.

Randhir, R., Lin, Y. T., and Shetty, K. 2014. Stimulation of phenolics, antioxidant and antimicrobial activities in dark germinated mung bean sprouts in response to peptide and phytochemical elicitors. Process Biochemistry.39: 637–646.

Rupali, S., Raha, S.B. and Santosh, S. 2016. Evaluation of analgesic activity of roots of Picrorhiza kurroa. J Drug Deliv Ther 3:99-104.

Salma, U., Kundu, S., and Gantait, S. 2017. Phytochemistry and Pharmaceutical significance of Picrorhiza kurroa Royle ex Benth. Phytochemistry and pharmacology of medicinal herbs. 3:26-37.

Samant, S.S., Pant .S.,Singh. M.,Lal. M., Singh.A., Sharma.A. and Bhandari . S. 2007. Medicinal plants in Himachal Pradesh, north western Himalayas India. The international journal of biodiversirty science and management.3:234-251.

Singh, V., Haque, S., Niwas, R., Srivastava., Pasupuleti , M. and Tripathi ,C.K.M. 2017. Strategies for fermentation medium optimization. Frontiers in Microbiology .7:2087.

Rasool, S. Sultan, P., and Hassan, Q.P. 2017. Picrorhiza kurroa Royle ex Benth, a plant of diverse pharmacological potential. Annals of phytomedicine, 6:63-67.

Tali, B.A., Khuroo, A.A., Nawchoo, I.A. and Ganie, A.H. 2018. Prioritizing conservation of medicinal flora in the Himalayan biodiversity hotspot: an integrated ecological and sub economic approach. Environmental conservation .1-8.

Wang, Y., Fang, X., An, F., Wang, G., Zhang, X. 2011. Improvement o antibiotic activity of Xenorhabdus bovienii by medium optimization using response surface methodology. Microbial cell factories, 10:1-15.

Siddiqi, A., Alam, S.S., Begum, S., Nazneen, Z. and Aaqil, B. 2015. Evaluation of

a. therapeutic potential of Picrorhiza kurroa glycosidal extract against nimesulide nephrotoxicity: A pilot study. Journal of Ayub medical college Abbottabad. 27:312 313.

Sehgal, R., Chauhan, A., Gilhotra, U.K. and Gilhotra, A. 2013. In-vitro and in-vivo evaluation of antiasthmatic activity of Picrorhiza kurroa plant. International journal of pharmaceutical sciences and research. 4:3440-3443.

Husain, G.M., Singh, P.N. and Kumar, V. 2009. Antidiabetic activity of standardized extract of Picrorhiza kurroa in rat model of NIDDM. Drug Discov Ther. 3:88-92.

Kumar, R., Gupta, Y.K., Singh, S. and Arunraja, S. 2016. Picrorhiza kurroa inhibits experimental arthritis through inhibition of Pro-inflammatory Cytokines, Angiogenesis and MMPs. Phytother Res 30:112–119.

Gupta, A., Khajuria, A., Singh, J., Bedi, K.L., Satti, N.K., Dutt, P., Suri, K.A., Suri, O.P. and Qazi, G.N. 2006. Immunomodulatory activity of bio polymeric fraction RLJ-NE om Picrorhiza kurroa. International Immunopharmacology. 6:1543–1549.

Mulliken, T.A. 2000. Implementing cites for Himalayan Medicinal Plants Nardostachys grandiflora and Picrorhiza kurroa. In: TRAFFIC B 18:63-72.

Salma, U., Kundu, S. and Gantait, S. 2017. Phytochemistry and Pharmaceutical Significance of Picrorhiza kurroa Royle ex Benth. Phytochemistry and Pharmacology of medicinal plants

Mulay, S.and Khale, A. 2011. Asavarishtas through improved fermentation technology. International journal of pharmaceutical sciences and research.2:1421-1425.

Valiathan, M.S. 2009.The legacy of vagbhata. Hyderabad, India: universities press Pvt. ltd.

Huynh, N.T.,Vancamp, J., Smagghe, G. and Raes, K. 2014.Improved release and metabolism of flavonoids by stirred fermentation processes. International journal of molecular sciences.15:19369-19388.

Hansen, H.B., Andreasen, M., Nielsen., Larsen, L., Kundsen, B.K., and Meyer, A. 2002. Changes in the dietary fibre ,phenolic acids and activity of endogenous enzymes during rye bread making. European food and research and technology.214:33-42.

Tyrakowska, B., Lemanska, K., Szymusiak, H., Borkowski, T. and Rietjens,I. 2003. Modified TEAC test for determination of the antioxidant properties of dietary polyphenolic compounds over a wide pH range. Polish journal of food and nutrition sciences.12:141-148.

Calinoiu, L.F., Catoi, A.F. and Vodnar, D.C. 2019. Solid-state fermented wheat and oat bran as a route for delivery of antioxidants. Antioxidants.8:372.

Nisa, K., Rosyida, V.T., Nurhayati, S., Indrianingsih, A.W., Darsih, C. and Apriyana, W. 2018. Total phenolic content and antioxidant activity of rice bran fermented with lactic acid bacteria. Earth and environmental science.251:012020.

Coghe, S., Benoot, K., Delvaux, F.,Vanderhaegen, B. and Delvaux, F.R. 2004. Ferulic acid release and 4-vinylguaicol formation during brewing and fermentation ,indication for feruloyl esterase activity in Saccharomyces cerevisiae. Journal of agricultural and food chemistry.52:602-608.

Nazarni, R., Purnama, D., Umar, S. and Eni, H. 2016. The effect of fermentation on the total phenolic, flavonoid and tannin content and its relation to antibacterial activity in Jaruk tigarum. International food research journal.23:309-315.

Ambwani, S., Tandon, R., Ambwani, T.K., Malik, Y.S. 2018. Current knowledge on nano delivery system and their beneficial applications in enhancing the efficacy of herbal drugs. Journal of experimental biology and agricultural sciences. 6:87-107

Govindrajan, R., Vijayakumar, M., and Pushpangadan, P. 2005. Antioxidant approach to disease management and role of ‘Rasayana’ herbs of ayurveda. Journal of ethnopharmacology.99:165-178.

Amri, E. and Kisangau, D.P. 2012. Ethno medicinal study of plants used in villages around Kimboza forest reserve in Morogoro, Tanzania. Journal of ethno biology and ethno medicine. 8:1.

Samant, S.S. and Pant, S. 2006. Diversity, distribution pattern and conservation status of plants used in liver diseases/ailments in Indian Himalayan region. Journal of mountain science. 3: 28-47.

Deshpande, N., Das, R.K., Muddeshwar, M., Das, V.O., Kandi, S. and Ramana, K.V. 2015. Antioxidants effects of Picrorhiza kurroa extracts in alcoholic cirrhosis of liver. American journal of

Rathee, D., Rathee, P., Rathee, S. and Rathee, D. 2012. Phytochemical screening and antimicrobial activity of Picrorhiza kurroa, an Indian traditional plant used to treat chronic diarrhea. Arabian journal of chemistry. 9: 1307-1313.

Sanlier, N., Gokcen, B.B. and Swzgin, A.C. 2017. Health benefits of fermented foods. Critical reviews in food science and nutrition. 1-22.

Barthlott, W., Mukte, J., Rafiqpoor, M.D., Kier, G. and Kreft, H. 2005. Global centers of vascular plant diversiry. Nova Acta Leopoldina. 92: 61-83.

Touret, T., Oliveira, M. and Semedo-lemsaddek, T. 2018. Putative probiotic lactic acid bacteria isolated from sauerkraut fermentations. Plos one.13.

Chandra, S., Saklani, S., Mishra, A.P. and Agrawal, R.K. 2016. In vitro antioxidant activity and Phytochemical screening of Garhwal Himalaya medicinal plants. International journal of medical research and health sciences. 8: 35-43.

Madhumitha, G. And Sara, A.M. 2009. Free radical scavenging assay of Thevetia neriifolia leaf extracts. Asian journal of chemistry. 21: 2468-2470.

Kulkarni, S.D., Tilak, J.C., Acharya, R., Rajurkar, N.S., Devasagayam, T.P.A. and Reddy, A.V.R. 2006. Evaluation of the antioxidant activity of wheatgrass (Triticum aestivum L.) as a function of growth under different conditions. Phytotherapy research. 20: 218-227.

Dixit, P., Devasagayam, T.P.A. and Ghaskadbi, S.S. 2005. Antioxidant effects of germinated fenugreek. Phytotherapy research. 19: 977-983.

Lone, P.A., Bhardwaj, A.K., Shah, K.W. and Tabasum, S. 2014. Ethno botanical survey of some threatened medicinal plants of Kashmir Himalaya, India. Journal of medicinal plant research.8: 1362-1373.

Kumar, R., Bhandari, P., Singh, B. and Ahuja, P.S. 2012. Evaluation of Picrorhiza kurroa accessions fpr growth and quality in north western Himalayas. Journal of medicinal plant research. 6: 2660-2665.

Bantwa, P., Saharoy, O., Gosh, S.K., and Mondal, T.K. 2011. In vitro regeneration of an endangered medicinal plant Picrorhiza scrophulariiflora. Biologia plantarum. 55: 169-172.

Uniyal, A., Uniyal, S.K. and Rawat, G.S. 2011. Commercial extraction of Picrorhiza kurroa Royle ex Benth in the western Himalaya. Mountain research and Development. 31: 201-208.

Chettri, N., Sharma, E., and Lama, S.D. 2005. Non-timber forest produces utilization, distribution and status in a trekking corridor of Sikkim, India. Lyonia. 8: 89-101.

Kumar, N., Kumar, T. and Sharma, S.K. 2013. Phyto pharmacological review on genus Picrorhiza. International journal of universal pharmacy and bio sciences. 2: 334-347.

Elizabeth, W.M. 2002. Major herbs of ayurveda. The Dabur research foundation and Dabur Ayurvet limited. 220-222.

Chin, Y.W., Balunas, M.J., Chai, H.B. and Kinghron, A.D. 2006. Drug discovery from natural sources. APPS Journal. 28: 132.

Anonymous, 2007. The ayurvedic pharmacopoeia of India. Part-I, government of India, ministry of health and family welfare department of Ayush. 2: 91-93.

Rasool, S., Khan, M.H., Hamid, S., Sultan, P., Qazi, P.H. and Butt, T. 2016. An overview and economical importance of few selected endangered medicinal plants grown in jammu and Kashmir region of India. Ann. Phytomed. 5: 27-37.

Liu, X.L., You, C., Yang, Y.W., Zhang, L. and Qian, Z.G. 2010. Anatomy and adaptation tp environment study of endangered alpine medical plant, Neopicrtorhiza scrophulariiflora zhong. Yao. Cai. 33: 507-510.

Smit, H.F. 2000. Picrorhiza scrophulariiflora, from traditional use to immunomodulatory activity. Utrecht, university of Utrecht, faculty of pharmacy.

Kumar, D., Kumar, R. and Ahuja, P. 2016. Comprehensive chemical profiling of Picrorhiza kurroa royle ex benth using NMR, HPTLC and LC-MS/MS techniques. Conbinatorial chemistry & high throughput screening. 19.

Bhattacharjee, S., Bhattaharya, S., Jana, S. and Bhagel, D.S. 2013. A review on medicinally important species of Picrorhiza. International journal of pharmacy research and bio sciences. 2: 1-16.

Upadhyay, D., Dash, R.P., Anandjiwala, S. and Nivsarkar, M. 2013. Comparative pharmacokinetic profiles of picrosides I and II from kutkin, Picrorhiza kurroa extract and its formulation in rats. Fitoterapia. 85: 76-83.

Rajkumar, V., Guha, G. and Kumar, R.A. 2011. Antioxidant and antineoplastic activities of Picrorhiza kurroa extract. Food and chemical toxicology. 49:363-369.

Philippens, I.H., Wubben, J.A., Finsen, B. and Hart, B.A. 2013. Oral treatment with the NADPH oxidase antagonist apocyanin mitigates clinical and pathological features pf Parkinsonism in the MPTP marmoset model. Journal of neuroimmune pharmacology. 8: 715-726.

Basu, K., Dasgupta, B., Bhattacharya, S.K. and Debnath, P.K. 1971. Chemistry and pharmacology of apocyanin, isolated from Picrorhiza kurroa Royle ex Benth. Current science.22: 603-604.

Park, S.B., Han, B.K., Oh, H.J., Lee, S.J., Cha, S.K., Park, Y.S. and Choi, H.J. 2012. Bioconversion of green tea extract using lactic acid bacteria. Food Engineering Progress. 16: 26-32.

Suthanthangjai, W., Kilmartin, P.A., Phillips, A.R.J., Davies, K. and Ansell, J. 2014. Bioconversion of Pinot Noir anthocyanin into bioactive phenolic compounds by lactic acid bacteria. The journal of nutrition, health and aging. 2: 145-149.

Lorenz, P., Bunse, M., Sauer, S., Conrad, J., Florian, C.S. and Kammerer, D.R. 2019. Conversion of Plant Secondary Metabolites upon Fermentation of Mercurialis perennis L. Extracts with two Lacto bacteria Strains. Fermentation. 5: 42.

Hussain, A., Bose, S., Wang, J.H., Yadav, M.K., Mahajan, G.B. and Kim, H. Fermentation, a feasible strategy for enhancing bioactivity of herbal medicine. 2016. Food research international. 81:1-16.

Pandit, S., Shitiz, K., Sood, H. and Chauhan, R.S. 2013. Differential biosynthesis and accumulation of picrosides in an endangered medicinal herb Picrorhiza kurroa. Journal of plant biochemistry and biotechnology. 22: 335-342.

Singh, M., Tiwari, V., Jain, A. and Ghoshal, S. 2006. Protective activity od picroliv on hepatic amoebiasis associated with carbon tetrachloride toxicity. Indian journal of medical research. 121: 676-682.

Stuppner, H., Wagner, H. 1989. Minor iridoids and phenol glycosides of Picrorhiza kurroa. Plant medicine. 55: 467-469.

Zhang, Y., Dewitt, D., Murugesan, S. and Nair, M. 2005. Cyclooxygenase-2 enzyme inhibitory triterpenoids from Picrorhiza kurroa seeds. Life sciences. 77: 3222-3230.

Wang, J.P., Xiao, Y., Rui, X., Xu, X., Guan, Y., Zhang, Q.Q. and Dong, M.S. 2016. Fu brick tea extract supplementation enhanced probiotic viability and antioxidant activity of tofu under simulated gastrointestinal digestion condition. RSC Advances. 6: 103668–103682.

Huang, Q., Zhang, H. and Xue, D. 2017. Enhancement of antioxidant activity of Radix Puerariae and red yeast rice by mixed fermentation with Monascus purpureus. Food Chemistry. 226: 89–94.

Zhu, F., Sakulnak, R. and Wang, S.A. 2016. Effect of black tea on antioxidant, textural, and sensory properties of Chinese steamed bread. Food Chemistry. 194: 1217–1223.

Xu, E.B., Wu, Z.Z., Long, J., Wang, F., Xu, X.M., Jin, Z.Y. and Jiao, A.Q. 2015. Improved bio accessibility of phenolic and antioxidant activity of glutinous rice and its fermented Chinese rice wine by simultaneous extrusion and enzymatic hydrolysis. Journal of functional foods. 17: 214–226.

Deshpande, N., Das, R.K., Muddeshwar, M., Das, V., Kandi, S. and Ramana, K.V. 2015. Antioxidant effects of picrorhiza kurroa rhizome extracts in alcoholic cirrhosis of liver. American journal of pharmacological sciences. 3: 49-51.

Katina, K., Laitila, A., Juvonen, R., Liukkonen, K. H., Kariluoto, S. and Piironen, V. 2007. Bran fermentation as a means to enhance technological properties and bioactivity of rye. Food Microbiology. 24 : 175–186.

Martins, S.,Mussatto, S. I., Martínez-Avila, G., Montañez-Saenz, J., Aguilar, C. N. and Teixeira, J. A. 2011. Bioactive phenolic compounds: production and extraction by solid-state fermentation. A review. Biotechnology Advances. 29: 365–373.

Oh, Y. C., Cho,W. K., Oh, J. H., Im, G. Y., Jeong, Y. H. and Yang, M. C. 2012. Fermentation by Lactobacillus enhances anti-inflammatory effect of Oyaksungisan on LPS-stimulated RAW 264.7 mouse macrophage cells. BMC Complementary and Alternative Medicine, 12: 17.

Parvez, S., Malik, K., Ah Kang, S. and Kim, H. Y. 2006. Probiotics and their fermented food products are beneficial for health. Journal of Applied Microbiology. 100: 1171–1185.

Wu, T., Wang, N., Zhang, Y. and Xu, X. 2013. Advances in the study on microbial fermentation and transformation of traditional Chinese medicine. African Journal of Microbiology Research. 7: 1644–1650.

Kim, A., Im, M., Hwang, Y., Yang, H. and Ma, J. 2014. Jaeumganghwa-tang induces apoptosis via the mitochondrial pathway and Lactobacillus fermentation enhances its anti-cancer activity in HT1080 human fibrosarcoma cells. PloS One. 10: 0127898.

Ajila, C., Brar, S., Verma, M., Tyagi, R. and Valéro, J. 2011. Solid-state fermentation of apple pomace using Phanerocheate chrysosporium—liberation and extraction of phenolic antioxidants. Food Chemistry. 126: 1071–1080.

Choi, J. H., Hwang, S. J., Jeong, S. N., Lee, Y. K., Jin,M. H., Park, S. G., & Lee, C. K. (2009). Ant aging effect on skin with 9 repetitive steaming and fermenting process herbal composition extract. The Korea Journal of Herbology, 24(4), 101–106.

Hur, S. J., Lee, S. Y., Kim, Y. C., Choi, I. and Kim, G. B. 2014. Effects of fermentation on the antioxidant activity in plant-based foods. Food Chemistry. 160: 346–356.

Lee, I. H., Hung, Y. H. and Chou, C. C. 2008. Solid-state fermentation with fungi to enhance the antioxidative activity, total phenolic and anthocyanin contents of black bean. International Journal of Food Microbiology. 121 : 150–156.

Ryu, J. S., Lee, H. J., Bae, S. H., Kim, S. Y., Park, Y., Suh, H. J. And Jeong, Y. H. 2013. The bioavailability of red ginseng extract fermented by Phellinus linteus. Journal of Ginseng Research. 37: 108.

Heiniö, R. L., Katina, K., Wilhelmson, A., Myllymäki, O., Rajamäki, T. and Latva-Kala, K. 2003. Relationship between sensory perception and flavour-active volatile compounds of germinated, sourdough fermented and native rye following the extrusion process. LWT- Food Science and Technology. 36: 533–545.

Jung, H.J., Choi, H., Lim, H.W., Shin, D., Kim, H., Kwon, B., Lee, J.E., Park, E.H. and Lim, C.J. 2012. Enhancement of anti inflammatory actions of red ginseng extract by fermentation. Journal of pharmacy and pharmacology. 64: 756-762.

Prabhu, A.A., Mrudula, C.M. and Rajesh, J. 2014. Effect of yeast fermentation on nutraceutical and antioxidant properties of rice bran. International journal of agriculture and food sciences. 4: 59-65.

Manwar, J., Mahadik, K., Sathiyanarayanan, L. and Paradkar, A. 2013. Comparative antioxidant potential of Withiania somnifera based herbal formulation prepared by traditional and non-traditional fermentation process. Integrative medicine research. 2: 56-61.

Haile, M. and Kang, W.H. 2019 The role of microbes in coffee fermentation and their impact on coffee quality. Journal of food quality. 12.

Wang, D.Q., He, Z.D., Feng, B.S. and Yang, C.R. 1993. Chemical constituents from Picrorhiza scrophulariiflora. Acta botanica Yunnanica. 15: 169-174.

Ansari, R.A., Aswal, B.S., Chander, R., Dhawan, B.N., Garg, N.K. and Kapoor, N.K. 1998. Hepatoprotective activity of kutkin- the iridoids glycoside mixture of Picrorhiza kurroa. Indian journal of med research. 87: 401-407.