Indian Journal of Microbiology

, Volume 56, Issue 1, pp 28–34 | Cite as

Microbiological Analyses of Traditional Alcoholic Beverage (Chhang) and its Starter (Balma) Prepared by Bhotiya Tribe of Uttarakhand, India

  • Kailash N. Bhardwaj
  • Kavish Kumar Jain
  • Sandeep Kumar
  • Ramesh Chander KuhadEmail author
Original Article


Present article depicts microbiology of starter (Balma) used in traditional solid-state fermentation of alcoholic beverage (Chhang) by Bhotiya tribe of Uttarakhand. It also highlights the importance of herbs in Balma preparation and kinetics of lactic acid and ethanol fermentation under Chhang preparation using Balma. Balma contains 214 × 106 cfu/g yeasts, 2.54 × 106 cfu/g lactic acid bacteria (LAB) and 1.4 × 106 cfu/g other mesophilic bacteria. ITS sequence analysis revealed a rich diversity of yeast comprising of Saccharomyces cerevisiae, Saccharomycopsis fibuligera and Saccharomycopsis malanga in Balma. 16S rDNA sequence analysis revealed Lactobacillus pentosus and Pediococcus pentosaceus among LAB, while amylolytic Bacillus subtilis and B. aerophilus among other bacteria in Balma. Based on the results, it is speculated that herbs such as Inula cuspidata, Micromeria biflora, Origanum vulgare, Rubus sp. and Thymus linearis used earlier by Bhotiya in Balma preparation contribute as a source of yeasts, LAB and amylolytic bacilli. Study also demonstrates that Bhotiya tribe is rational in preparation of starter as they have circumvented the need of plants by using previous year Balma as inoculum and possibility of deficient quality of Balma due to weak colonization of phyllosphere and rhizosphere microbiota. Results suggest that simultaneous saccharification and lactic acid–ethanol fermentation take place in traditional cereal based Chhang fermentation system of Bhotiya.


Traditional fermentation system Balma Keem Bhotiya Jaunsari 



KNB gratefully acknowledge the financial assistance from UGC-Dr DS Kothari Postdoc Fellowship [F.4-2/2006(BSR)/13-374/2010(BSR)], KKJ acknowledge CSIR (Council of Scientific and Industrial Research) for Senior research fellowship, SK thanks to the ICMR (Indian Council of Medical Research) for Senior research fellowship and University of Delhi South Campus, New Delhi for providing necessary laboratory facility. We wish to thank Dr Rishi Gupta (DST-fast track young scientist, Lignocellulose biotechnology laboratory, UDSC) for his help in analysis of ethanol and manuscript writing. Authors also wish to thank Dr Arvind Kumar (UGC-Dr DS Kothari fellow, Lignocellulose biotechnology laboratory, UDSC) for his help in molecular taxonomic analysis.


  1. 1.
    Scott R, Sullivan WC (2008) Ecology of fermented foods. Hum Ecol Rev 15:25–31Google Scholar
  2. 2.
    Roy B, Kala CP, Farooquee NA, Majila BS (2004) Indigenous fermented food and beverages: a potential for economic development of the high altitude societies in Uttaranchal. J Hum Ecol 15:45–49Google Scholar
  3. 3.
    Savitri Bhalla TC (2007) Traditional foods and beverages of Himachal Pradesh. Indian J Tradit Knowl 6:17–24Google Scholar
  4. 4.
    Dhal NK, Pattanaik C, Raddy CS (2010) Bakhar starch fermentation: a common tribal practice in Orissa. Indian J Tradit Knowl 9:279–281Google Scholar
  5. 5.
    Das AJ, Deka SC, Miyaji T (2012) Methodology of rice beer preparation and various plant materials used in starter culture preparation by some tribal communities of North-East India: a survey. Int Food Res J 19:101–107Google Scholar
  6. 6.
    Das AJ, Deka SC (2012) Fermented foods and beverages of the North-East India. Int Food Res J 19:377–392Google Scholar
  7. 7.
    Rana TS, Dutt B, Rao RR (2004) Soor: a traditional alcoholic beverage in Tons valley, Garhwal Himalaya. Indian J Tradit Knowl 3:59–65Google Scholar
  8. 8.
    Bhatt KC, Silas RA (1990) Traditional beverages and their importance in the folk life of Bhotiyas in U.P. Himalaya. J Himal Stud Reg Dev 13–14:56–62Google Scholar
  9. 9.
    Das CP, Pandey A (2007) Fermentation of traditional beverages prepared by Bhotiya community of Uttaranchal Himalaya. Indian J Tradit Knowl 6:136–140Google Scholar
  10. 10.
    Miller GL (1959) Use of dinitrosalicylic acid reagent for determination of reducing sugars. Anal Chem 31:426–428CrossRefGoogle Scholar
  11. 11.
    Singleton VL, Orthofer R, Lamuela-Raventos RM (1999) Analysis of total phenols and other oxidation substrates and antioxidants by means of Folin–Ciocalteu reagent. Meth Enzymol 299:152–178. doi: 10.1016/S0076-6879(99)99017-1 CrossRefGoogle Scholar
  12. 12.
    Kumar A, Mukherjee S, Chakraborty R (2010) Characterization of a novel trimethoprim resistance gene, dfrA28, in Class 1 Integron of an oligotrophic Acinetobacter johnsonii strain, MB52, isolated from river Mahananda, India. Microb Drug Resist 16:29–37. doi: 10.1089/mdr.2009.0111 CrossRefPubMedGoogle Scholar
  13. 13.
    White TJ, Bruns T, Lee S, Taylor J (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenitics. In: Innis N, Gelfand D, Sninsky J, White T (eds) PCR protocol: a guide to methods and applications. Academic Press, London, pp 315–322Google Scholar
  14. 14.
    Perveen K, Alabdulkarim B, Arzoo S (2011) Effect of temperature on shelf life, chemical and microbial properties of cream cheese. Afr J Biotechnol 10:16929–16936. doi: 10.5897/AJB11.1695 CrossRefGoogle Scholar
  15. 15.
    Thakur N, Savitri Saris PerEJ, Bhalla TC (2015) Microorganisms associated with amylolytic starters and traditional fermented alcoholic beverages of North Western Himalayas in India. Food Biosci 11:92–96. doi: 10.1016/j.fbio.2015.05.002 CrossRefGoogle Scholar
  16. 16.
    Tamang JP, Dewan S, Tamang B, Rai A, Schillinger U, Holzapfel WH (2007) Lactic acid bacteria in Hamei and Marcha of North East India. Indian J Microbiol 47:119–125. doi: 10.1007/s12088-007-0024-8 PubMedCentralCrossRefPubMedGoogle Scholar
  17. 17.
    Thokchom S, Joshi SR (2014) Physicochemical analysis of ethnically fermented soybean products of north-east India and molecular characterization of associated lactic acid bacteria. Proc Natl Acad Sci India Sect B 85:527–533. doi: 10.1007/s40011-013-0199-1 CrossRefGoogle Scholar
  18. 18.
    Yu JJ, Park HJ, Kim SG, Oh SH (2009) Isolation, identification and characterization of Weissella strains with high ornithine producing capacity from Kimchi. Korean J Microbiol 45:339–345Google Scholar
  19. 19.
    Tanti B, Gurung L, Sarma HK, Baragohain AK (2010) Ethnobotany of starter cultures used in alcohol fermentation by a few ethnic tribes of Northeast India. Indian J Tradit Knowl 9:463–466Google Scholar
  20. 20.
    Kanwar SS, Gupta MK, Katoch C, Kanwar P (2011) Cereal based traditional alcoholic beverages of Lahaul and Spiti area of Himachal Pradesh. Indian J Tradit Knowl 10:251–257Google Scholar
  21. 21.
    Daeschel MA, Anderson RE, Fleming HP (1987) Microbial ecology of fermenting plant materials. FEMS Microbiol Rev 46:357–367. doi: 10.1111/j.1574-6968.1987.tb02472.x CrossRefGoogle Scholar
  22. 22.
    Sati B, Thapliyal S, Sati H, Saklani S, Kumar P, Bhatt PC (2011) Preliminary phytochemical, physicochemical and antimicrobial studies of Inula cuspidata leaves. J Global Trends Pharm Sci 2:290–295Google Scholar
  23. 23.
    Kumar A, Gupta R, Mishra RK, Shukla AC, Dikshit A (2012) Pharmaco-phylogenetic investigation of Micromeria biflora Benth and Citrus reticulata Blanco. Natl Acad Sci Lett 35:253–257. doi: 10.1007/s40009-012-0029-7 CrossRefGoogle Scholar
  24. 24.
    Thiem B, Goslinska O (2004) Antimicrobial activity of Rubus chamaemorus leaves. Fitoterapia 75:93–95. doi: 10.1016/j.fitote.2003.08.014 CrossRefPubMedGoogle Scholar
  25. 25.
    De Falco E, Mancini E, Roscigno G, Mignola E, Taglialatela-Scafati O, Senatore F (2013) Chemical composition and biological activity of essential oils of Origanum vulgare L. subsp. vulgare L. under different growth conditions. Molecules 18:14948–14960. doi: 10.3390/molecules181214948 CrossRefPubMedGoogle Scholar
  26. 26.
    Hussain AI, Anwar F, Shahzad ASC, Latif S, Sherazi STH, Ahmad A, Worthington J, Sarker SD (2013) Chemical composition and bioactivity studies of the essential oils from two Thymus species from the Pakistani flora. Food Sci Technol 50:185–192. doi: 10.1016/j.lwt.2012.06.003 Google Scholar
  27. 27.
    Greeshma AG, Srivastava B, Srivastava K (2006) Plants used as antimicrobials in the preparation of traditional starter cultures of fermentation by certain tribes of Arunachal Pradesh. Bull Arunachal Forest Res 22:52–57Google Scholar
  28. 28.
    Nout MJR (1992) Upgrading traditional biotechnological processes. In: Gaden EL, Bokanga M, Harlander S, Hesseltine CW, Steinkraus KH (eds) Applications of biotechnology in traditional fermented foods. National Academy Press, Washington. ISBN: 0-309-56575-8, pp 11–19. Accessed 29 Sept 2015
  29. 29.
    Zheng X-W, Tabrizi MR, Nout MJR, Han B-Z (2011) Daqu—a traditional Chinese liquor fermentation starter. J Inst Brew 117:82–90. doi: 10.1002/j.2050-0416.2011.tb00447.x CrossRefGoogle Scholar
  30. 30.
    Dung NTP (2013) Vietnamese rice-based alcoholic beverages. Int Food Res J 20:1035–1041Google Scholar
  31. 31.
    Sekar S, Mariappan S (2005) Usage of traditional fermented products by Indian rural folks and IPR. Indian J Tradit Knowl 6:111–120Google Scholar
  32. 32.
    Aidoo KE, Nout MJR, Sarkar PK (2006) Occurrence and function of yeasts in Asian indigenous fermented foods. FEMS Yeast Res 6:30–39. doi: 10.1111/j.1567-1364.2005.00015.x CrossRefPubMedGoogle Scholar
  33. 33.
    Tamang JP, Sarkar PK (1995) Microflora of Murcha: an amylolytic fermentation starter. Microbios 81:115–122PubMedGoogle Scholar
  34. 34.
    Tsuyoshi N, Fudou R, Yamanaka S, Kozaki M, Tamang N, Thapa S, Tamang JP (2005) Identification of yeast strains isolated from Marcha in Sikkim, a microbial starter for amylolytic fermentation. Int J Food Microbiol 99:135–146. doi: 10.1016/j.ijfoodmicro.2004.08.011 CrossRefPubMedGoogle Scholar
  35. 35.
    Tamang JP, Thapa S (2006) Fermentation dynamics during production of BhaatiJaanr, a traditional fermented rice beverage of the Eastern Himalayas. Food Biotechnol 20:251–261. doi: 10.1080/08905430600904476 CrossRefGoogle Scholar

Copyright information

© Association of Microbiologists of India 2015

Authors and Affiliations

  • Kailash N. Bhardwaj
    • 1
    • 2
  • Kavish Kumar Jain
    • 1
  • Sandeep Kumar
    • 1
  • Ramesh Chander Kuhad
    • 1
    • 3
    Email author
  1. 1.Lignocellulose Biotechnology Laboratory, Department of MicrobiologyUniversity of Delhi South CampusNew DelhiIndia
  2. 2.Uttarakhand State Council for Science and Technology (UCOST)DehradunIndia
  3. 3.Central University of HaryanaPaliIndia

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