Production and Purification of Pectinase from Bacillus subtilis 15A-B92 and Its Biotechnological Applications.

Enzymes that degrade pectin are called pectinases. Pectinases of microbial origin are used in juice clarification as the process is cost-effective. This study screened a pectinase-producing bacterium isolated from soil and identified as Bacillus subtilis 15A B-92 based on the 16S rRNA molecular technique. The purified pectinase from the isolate showed 99.6 U/mg specific activity and 11.6-fold purity. The molecular weight of the purified bacterial pectinase was 14.41 ± 1 kD. Optimum pectinase activity was found at pH 4.5 and 50 °C, and the enzyme was 100% stable for 3.5 h in these conditions. No enzymatic inhibition or activation effect was seen with Fe2+, Ca2+, or Mg2+. However, a slight inhibition was seen with Cu2+, Mn2+, and Zn2+. Tween 20 and 80 slightly inhibited the pectinase, whereas iodoacetic acid (IAA), ethylenediaminetetraacetate (EDTA), urea, and sodium dodecyl sulfate (SDS) showed potent inhibition. The bacterial pectinase degraded citrus pectin (100%); however, it was inactive in the presence of galactose. With citrus pectin as the substrate, the Km and Vmax were calculated as 1.72 mg/mL and 1609 U/g, respectively. The high affinity of pectinase for its substrate makes the process cost-effective when utilized in food industries. The obtained pectinase was able to clarify orange and apple juices, justifying its application in the food industry.

Purification and biochemical characterization of extracellular glucoamylase from Paenibacillus amylolyticus strain.

In the present study, glucoamylase produced from a soil bacterium Paenibacillus amylolyticus NEO03 was cultured under submerged fermentation conditions. The extracellular enzyme was purified by starch adsorption chromatography and further by gel filtration, with 2.73-fold and recovery of 40.02%. The protein exhibited molecular mass of ∼66,000 Da as estimated by SDS-PAGE and depicted to be a monomer. The enzyme demonstrated optimum activity at pH range 6.0-7.0 and temperature range 30-40 °C. Glucoamylase was mostly activated by Mn2+ metal ions and depicted no dependency on Ca2+ ions. The enzyme preferentially hydrolyzed all the starch substrates. High substrate specificity was demonstrated towards soluble starch and kinetic values Km and Vmax were 2.84 mg/ml and 239.2 U/ml, respectively. The products of hydrolysis of soluble starch were detected by thin layer chromatography which showed only D -glucose, indicating a true glucoamylase. The secreted glucoamylase from P. amylolyticus strain possesses properties suitable for saccharification processes such as biofuel production.

Large-scale production of enzymes for biotechnology uses.

Enzymes are biocatalysts that speed up the chemical reaction to obtain the final valuable product/s. Biotechnology has revolutionized the use of traditional enzymes to be applicable in industries such as food, beverage, personal and household care, agriculture, bioenergy, pharmaceutical, and various other segments. With respect to the exponential growth of enzymes in biotech industries, it becomes important to highlight the advancements and impact of enzyme technology over recent years. In this review article, we discuss the existing and emerging production approaches, applications, developments, and global need for enzymes. Special emphasis is given to the predominantly utilized hydrolytic microbial enzymes in industrial bioprocesses.

Potential of herbal cocktail of medicinal plant extracts against 'big four' snake venoms from India.

BACKGROUND: Venomous snake bites cause acute medical emergencies and are fatal. India accounts for large proportion of snake-bite deaths globally. Medically important 'BIG FOUR' snakes of India are Bungarus caeruleus (krait), Naja naja (cobra), Echis carinatus (saw-scaled viper) and Daboia russelii (Russell's viper). Polyherbal formulations have been proved to be effective in treatment of diseases than a single formulation. OBJECTIVE(S): To evaluate aqueous ethanolic extract cocktail of Azadirachata indica, Butea monosperma, Citrus limon, Clerodendrum serratum and Areca catechu for antidote potential against BIG FOUR venoms in ex vivo and in vivo model. MATERIALS AND METHODS: Anti-hemorrhagic and venom neutralization studies were performed in seven-day old chick embryo model for ex vivo studies. In vivo studies were performed using male Swiss albino mice for antivenom potential of herbal cocktail by performing anti-edematic, anti-hemorrhagic, anti-myotoxic activity, and venom neutralization. RESULTS: Herbal cocktail exhibited differential venom inhibition potential against four venoms tested. Hemorrhagic activity was completely neutralized by the herbal cocktail; myotoxic activities of krait and Russell's viper venom were neutralized; while anti-edematic activity was observed for krait and cobra venom. Herbal cocktail completely neutralized venom lethality (3∗LD50) of krait and saw-scaled viper venom. CONCLUSION: Inhibitions of various venom components of all four venoms suggests presence of phytochemicals in herbal cocktail with therapeutic properties. Further studies would help in the development of a formulation as a first-aid towards treatment of snake bite victims.

Bungarus caeruleus venom neutralization activity of Azima tetracantha Lam. Extract.

Azima tetracantha Lam. is native to Africa and India. The plant and its parts are used for treating various ailments including snake bites. The different concentrations of ethyl acetate leaf extract of A. tetracantha were used to neutralize the toxic effect of venom through dose dependent enzyme studies and in vivo studies. The extract was able to neutralize the 5' nucleotidase, phospholipase A2, Phosphodiesterae, phosphomonoesterase, acetylcholinesterase and hyaluronidase in a dose dependent manner with concentrations ranging from 43.98 -340.1 µg/mL of extract. The extract could retain the lysis of fibrinogen at the concentration of 1:10 (venom: extract, w/w) and also the lysis of lecithin was reduced at a concentration of 1:25 (venom: extract, w/w). The extract was able to neutralize the LD50 of venom in both mice and embryo and also reduce the myotoxic and edema properties of the venom in mice models. The venom did not show any procoagulant and hemorrhagic effect. The leaf extract possess adequate compounds/phytochemicals that could neutralize the toxic properties/activity of the B. caeruleus venom.

Scorpion Toxin Polyptides as Therapeutic Agents: An Overview.

Scorpions are distributed throughout the world and numerous biological molecules are found in their venom most importantly peptide toxins. These toxins modulate the ion channels either by blocking the pore of the channel or by altering the voltage gating. Molecules which block the pores have been useful in deciphering the structure of the ion channels. Many scorpion toxins have already been used for probing the voltage gated sodium channels and studying their activation and inactivation processes. The specialty of scorpion toxins is to discriminate between vertebrate and invertebrate channels which have led them to applications as pharmacological tools. Most of the scorpion toxin polypeptides were isolated, characterized and were shown to possess vital properties useful in the field of medicine. For instance, they show therapeutic properties such as antimicrobial activity, anticancer activity, used to treat autoimmune diseases and cardiovascular effects. Although the scorpion toxins exhibited good therapeutic effects in vitro and in vivo, no one has reached the market with success up to date. In this mini-review, the scorpion polypeptides, their interactions with ion channels and their uses as therapeutic agents are discussed.

Enhanced degradation of pendimethalin by immobilized cells of Bacillus lehensis XJU.

A bacterium capable of degrading pendimethalin was isolated from the contaminated soil samples and identified as Bacillus lehensis XJU based on 16S rRNA gene sequence analysis. 6-Aminopendimethalin and 3,4-dimethyl 2,6-dinitroaniline were identified as the metabolites of pendimethalin degradation by the bacterium. The biodegradation of pendimethalin by freely suspended and the immobilized cells of B. lehensis on various matrices namely agar, alginate, polyacrylamide, and polyurethane foam was also investigated. The batch degradation rate was nearly the same for both free and immobilized cells in agar and alginate, whereas polyacrylamide- and PUF-immobilized cells degraded 93 and 100 of 0.1 % pendimethalin after 96 and 72 h, respectively. At higher concentration, the degradation rate of freely suspended cells decreased; whereas the same immobilized cells on polyurethane foam completely degraded 0.2 % pendimethalin within 96 h. The repeated batch degradation with the polyurethane foam-immobilized cells was reused for 35 cycles without losing the 0.1 % pendimethalin degrading ability. In contrast, agar-, alginate- and polyacrylamide-immobilized cells could be reused for 15, 18, and 25 cycles, respectively. When the pendimethalin concentration was increased to 0.2 %, the immobilized cells could be reused but the pendimethalin degradation rate was decreased. Polyurethane foam-immobilized cells exhibited better tolerance to pH and temperature alterations than freely suspended cells and could be stored for more than 3 months without losing pendimethalin degrading ability. The immobilization of cells capable of degrading pendimethalin may serve as an ideal technique for the complete degradation of the herbicide in the environment.