Isolation and Characterization of Thermophilic Microbial Enzymes Protease and Amylase from Hot Spring Madkot, Pithoragarh in Uttarakhand

Abstract

Thermophilic environments such as hot springs are invaluable natural reservoirs of extremophilic microorganisms that produce thermostable enzymes with significant industrial utility. This study explored the thermophilic microbial community of the Madkot geothermal hotspot (Pithoragarh), Uttarakhand. Samples collected from high-temperature niches were subjected to enrichment and isolation under thermophilic conditions, followed by comprehensive biochemical profiling including indole, methyl red, Voges–Proskauer, phosphate solubilization, catalase, oxidase, urease and nitrate reduction tests to elucidate metabolic capabilities relevant to industrial processes. Selected thermophilic strains were molecularly identified by 16S rRNA gene sequencing and affiliated with the genera Bacillus and Pseudomonas. Both taxa demonstrated the capacity to produce industrially relevant enzymes under elevated temperature regimes: a Bacillus sp. isolate exhibited protease activity with a maximum specific activity of 23.05 U/ml, while a Pseudomonas sp. isolate synthesized amylase with a peak specific activity of 31.02 U/ml. The thermostability and activity profiles of these enzymes suggest suitability for applications that demand resilience to heat and harsh chemical conditions, including biofuel generation, high-temperature food processing, pharmaceutical enzyme formulations and bioremediation of thermally stressed or contaminated environments. Overall, the results underscore the rich, yet underexplored, thermophilic microbial diversity of Uttarakhand hot springs and emphasize their importance as a source of sustainable, heat-tolerant biocatalysts for industrial biotechnology. Future work will focus on enzyme purification, detailed thermostability profiling and cost effective pilot-scale production in relevant process conditions.