Ravi Kant Bhatia

Biosynthesis of polyesters and polyamide building blocks using microbial fermentation and biotransformation

Biopolymers can be a green alternative to fossil-based polymers and can contribute to environmental protection because they are produced using renewable raw materials. Biopolymers are composed of various small subunits (building blocks) that are the intermediates or end products of major metabolic pathways. Most building blocks are secreted directly outside of cells, making downstream processes easier and more economic. These molecules can be extracted from fermentation broth and polymerized to produce a variety of biopolymers, e.g., polybutylene terephthalate, polyethylene terephthalate, polytrimethylene terephthalate, nylon-5,4 and nylon-4,6, with applications in medicine, pharmaceuticals, and textiles. Microbes are unable to naturally produce these types of polymers; thus, the production of building blocks and their polymerization is a fascinating approach for the production of these polymers. In comparison to naturally occurring biopolymers, synthesized polymers have improved and controlled structures and higher purity. The production of monomer units provides a new direction for polymer science because new classes of polymers with unique properties that were not previously possible can be prepared. Furthermore, the engineering of microbes for building-block production is an easy process compared to engineering an entire biopolymer synthesis pathway in a single microbe. Polyesters and polyamide polymers have become an important part of human life, and their demand is increasing daily. In this review, recent approaches and technology are discussed for the production of polyester/polyamide building blocks, i.e., 2-hydroxyisobutyric acid, 3-hydroxypropionic acid, mandelic acid, itaconic acid, adipic acid, terephthalic acid, succinic acid, 1,3-propanediol, 2,3-butanediol, 1,4-butanediol, 1,3-butanediol, cadaverine, and putrescine.

Purification and characterization of arylacetonitrile‐specific nitrilase of Alcaligenes sp. MTCC 10675

Arylacetonitrile‐hydrolyzing nitrilase (E.C. 3.5.5.5) of Alcaligenes sp. MTCC 10675 has been purified by up to 46‐fold to homogeneity and 32% yield using ammonium sulfate fractionation, Sephacryl S‐300 gel permeation, and anion exchange chromatography. The molecular weight of the native enzyme was estimated to be 520 ± 60 kDa. The subunit has a molecular weight of 60 ± 14 kDa in sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS‐PAGE). The optimum pH and temperature of the purified enzyme were 6.5 and 50 °C, respectively. The purified arylacetonitrilase has a half‐life of 3 H 20 Min at its optimum temperature. The value for Vmax, Km, kcat, and ki of enzyme for mandelonitrile as a substrate was 50 ± 05 µmol/Min/mg, 13 ± 02 mM, 26 ± 03 Sec−, and 32.4 ± 03 mM, respectively. Alcaligenes sp. MTCC 10675 arylacetonitrilase amino acid sequence has variations from other reported arylacetonitrilase, namely, A11G, N21H, D149N, S170T, P171R, S179A, Q180N, and S191A, and it has a high thermal stability and catalytic rate as compared with the already purified arylacetonitrilase.

Biotechnological potential of microbial consortia and future perspectives

Abstract Design of a microbial consortium is a newly emerging field that enables researchers to extend the frontiers of biotechnology from a pure culture to mixed cultures. A microbial consortium enables microbes to use a broad range of carbon sources. It provides microbes with robustness in response to environmental stress factors. Microbes in a consortium can perform complex functions that are impossible for a single organism. With advancement of technology, it is now possible to understand microbial interaction mechanism and construct consortia. Microbial consortia can be classified in terms of their construction, modes of interaction, and functions. Here we discuss different trends in the study of microbial functions and interactions, including single-cell genomics (SCG), microfluidics, fluorescent imaging, and membrane separation. Community profile studies using polymerase chain-reaction denaturing gradient gel electrophoresis (PCR-DGGE), amplified ribosomal DNA restriction analysis (ARDRA), and terminal restriction fragment-length polymorphism (T-RFLP) are also reviewed. We also provide a few examples of their possible applications in areas of biopolymers, bioenergy, biochemicals, and bioremediation.

Microbial Degradation of Cyanides and Nitriles

Cyanide and nitriles are produced by a wide range of microorganisms and plants as part of their normal metabolism. These are ubiquitous at low levels in soil and water including surface and ground water. Cyanide is used in metallurgical operations in mining of gold and chemical synthesis and nitriles are also extensively used in organic synthesis and as agrochemicals. The sources of cyanide and nitrile contamination of environment include emissions from iron and steel production, coal combustion, petroleum refineries, solid waste incinerators, combustion of nitriles, use of agrochemicals, chemical industries, vehicle exhausts and cigarette smoke. The industrial and anthropological activities have resulted in contamination of soil, water and air with toxic levels of nitriles and cyanide in the environment. In some situations nitrile and cyanide pollution becomes a threat to animal and human beings. A large number of microorganisms have been reported to the degrade nitriles and cyanide to corresponding non-toxic acids or amides. Some microbes have cyanide hydratase or dihydratase enzymes to convert cyanide to formamide or formic acid while others are endowed with nitrilase, nitrile hydratase and amidase systems which transform nitriles to acids and amides. In this chapter we will discuss the sources and extent of cyanide and nitrile contamination of soil, water and air and the potential application of nitrile or cyanide metabolizing organisms in the bioremediation of contaminated habitats will be discussed.

Fuel from Waste: A Review on Scientific Solution for Waste Management and Environment Conservation

Millions of tons of solid wastes are produced as a result of various household, agricultural, and industrial activities around the globe every year, which if not managed and disposed properly can create serious health and environmental issues. At the same time, huge amounts of coal, oil, and natural gas are burnt daily to generate electricity and power to run domestic, workplace, and industrial appliances. The excessive and un-managed use of fossil fuels has not only put pressure on already limited resources, but has also resulted as major contributor of environment pollution. The scientific community has been continuously searching for the renewable and alternate sources of fuel on one hand and an amicable solution to manage the waste on the other hand. These two biggest challenges before the world presently, if diagnosed and managed scientifically, can provide solution to each other by providing clean renewable energy from solid and liquid waste materials. Waste-to-energy technologies physically convert waste matter into more useful forms like bioethanol, biobutanol, biogas, biohythane, CNG, and syngas through various processes such as combustion, pyrolysis, gasification, or biological treatments. The processes like anaerobic digestion and fermentation and combinations of various technologies can be used to tackle the rising demand of energy. Utilizing these wastes would not only provide supply of fuels on sustainable basis but would also be helpful in conserving our environment. The selection of appropriate raw material and efficient technology for biofuel production is of immense importance in order to produce high-quality product with reduced environmental impact. Various aspects of waste utilization through clean renewable source of energy for sustainable development of society are of paramount significance in today’s context and need immediate attention.

Bio-statistical enhancement of acyl transfer activity of amidase for biotransformation of N-substituted aromatic amides.

Acyl transfer activity (ATA) of amidase transfers an acyl group of different amides to hydroxylamine to form the corresponding hydroxamic acid. Bacterial isolate BR-1 was isolated from cyanogenic plant Cirsium vulgare rhizosphere and identified as Pseudomonas putida BR-1 by 16S rDNA sequencing. This organism exhibited high ATA for the biotransformation of N-substituted aromatic amide to the corresponding hydroxamic acid. Optimization of media, tryptone (0.6%), inducer, pH 8.5, and a growth temperature 25°C for 56 h, resulted in a 7-fold increase in ATA. Further, Response Surface Methodology (RSM) and multiple feeding approach (20 mM after 14 h) of inducer led to a 29% enhancement of ATA from this organism. The half life (t1/2) of this enzyme at 50°C and 60°C was 3 h and 1 h, respectively. The ATA of amidase of Pseudomonas putida BR-1 makes it a potential candidate for the production of a variety of N-substituted aromatic hydroxamic acid.