The study aims to isolate the yeast strains that could be used effectively as baker's yeast and compare them with the commercial baker's yeast available in the market of Nepal. A total of 10 samples including locally available sources like fruits, Murcha, and a local tree "Dar" were collected from different localities of Bhaktapur, Kavre, and Syangja districts of Nepal, respectively. Following enrichment and fermentation of the samples, 26 yeast strains were isolated using selective medium Wallerstein Laboratory Nutrient Agar. From the differential tests which included morphological and microscopic observation and physiological and biochemical characterization such as nitrate reduction and lactose utilization tests, 8 strains were selected as possible Saccharomyces strain. The selected strains were further assessed for their efficient leavening ability by tests such as ethanol tolerance, osmotolerance, invertase test, and stress exclusion test. The three most potent strains ENG, MUR3B, and SUG1 isolated from grape, Murcha, and sugarcane, respectively, were used in the fermentation and baking of dough. These strains also carried a possibility of being used as industrial baker's yeast.

A thermophilic strain, Aeribacillus pallidus BTPS-2 was isolated from Bhurung geothermal spring of Nepal. The 16 s rRNA sequence showed 99.8 % similarity with the type strain Aeribacillus pallidus DSM 3670. The morphological, physiological and biochemical properties were similar to the type strain. Alpha-amylase from A. pallidus BTPS-2 was purified to 19-fold purification by DEAE-Cellulose ion exchange chromatography. The K m value of amylase on starch was 0.51 ± 0.05 mg/mL. The optimum pH and temperature were 7.0 and 70 °C. SDS-PAGE analysis showed a single band at 100 kDa. The half-life of the enzyme at 80 °C was 2.81 h. The enzyme showed an inhibitory effect in the presence of Fe 2+ , Pb 2+ , Sn 2+ and Hg 2+ at 10 mM concentrations. TLC analysis showed that the enzyme is a liquifying alpha-amylase. The enzyme reduced the viscosity of algal biomass suspension up to 74.2 ± 0.17 % which was more efficient than Bacillus amyloliquefaciens alpha-amylase (80.5 ± 0.2 %).

Alkaline protease is one of the most important industrial enzymes which are excessively used in the detergent industry, food industry, feed industry, pharmaceutical industry, leather industry, etc. 60% of the produced alkaline protease is consumed by the detergent industry alone. In the present study, bacterial isolates that can produce alkaline protease for purpose of bio-detergent were screened among the isolates isolated from kinema (an alkaline fermented food of eastern Nepal). Selected bacterial isolates were further screened for hemolysis activity and the production of other hydrolytic enzymes. Four bacterial isolates selected were tested for their capacity to produce alkaline protease in five different fermentation mediums. Isolate BKHE produces a high amount of alkaline protease (0.4705 ± 0.035 U/mL/min) in fermentation medium M2 (sucrose, 11 g/L; yeast extract, 5 g/L; and KNO3, 5.2 g/l, pH 9). The selected isolate was identified as Bacillus amyloliquefaciens BKHE based on 16S rRNA sequencing and phenotypic features. This bacterial strain was also found to be thermotolerant (confluent growth at 50°C) and salt tolerant up to 10% NaCl concentration. With its versatile ability, bacterial isolate or purified enzymes have potential applications in the food and detergent industry.

A thermophilic amylolytic strain, Anoxybacillus tengchongensis RA1-2-1 was isolated from geothermal spring of Rasuwagadi district of Nepal. The BLAST alignment of the 16s rRNA sequence revealed 99.3% similarity with the type strain Anoxybacillus tengchongensis T-11. The morphological, physiological and biochemical properties were similar to the type strain. The enzyme from the strain was purified to 40-fold purification by DEAE-cellulose ion exchange chromatography. The Km value of the enzyme was 0.68±0.05 mg/ml. The optimum pH and temperature were 7.0 and 70 °C. SDS-PAGE analysis showed a single band at 69 kDa. The half-life of the enzyme at 70°C and 80°C were 85.01min and 51.96 min respectively. TLC analysis of the hydrolysis product showed that the enzyme is maltogenic amylase. The calcium independent enzyme was completely inhibited by Hg2+ but showed inhibitory effect in the range of 100 %-30 % in the presence of other salts at 1-10 mM concentrations.