Current Biotechnology - Volume 9, Issue 2, 2020

The diversity of indole concerning its production and functional role has increased in both prokaryotic and eukaryotic systems. The bacterial species produce indole and use it as a signaling molecule at interspecies, intraspecies, and even at an interkingdom level for controlling the capability of drug resistance, level of virulence, and biofilm formation. Numerous indole derivatives have been found to play an important role in the different systems and are reported to occur in various bacteria, plants, human, and plant pathogens. Indole and its derivatives have been recognized for a defensive role against pests and insects in the plant kingdom. These indole derivatives are produced as a result of the breakdown of glucosinolate products at the time of insect attack or physical damages. Apart from the defensive role of these products, in plants, they also exhibit several other secondary responses that may contribute directly or indirectly to the growth and development. The present review summarized recent signs of progress on the functional properties of indole and its derivatives in different plant systems. The molecular mechanism involved in the defensive role played by indole as well as its’ derivative in the plants has also been explained. Furthermore, the perspectives of indole and its derivatives (natural or synthetic) in understanding the involvement of these compounds in diverse plants have also been discussed.

Introduction: Selinum vaginatum is an endemic Indian plant present on high altitude and is known as “Bhootkeshi,” by the locals. The root of plant Selinum vaginatum commonly known as “Bhootkeshi” from the family Umbellifercae is an aromatic herbal drug. Selinum vaginatum has been used traditionally for curing seizures, epilepsy, mental illness, and hysteria. This plant has shown therapeutic effects as used by the locals. There is no data available regarding standardization of the roots of Selinum vaginatum. It is a novel work as the plant is rarely found and shows medicinal values but no quality control work has been performed. Objective: The aim of this paper is to study quality control and carry out physicochemical analysis and standardization of the roots of Selinum vaginatum for determining its quality and purity. As no work has been performed on the roots of this plant, so standardization will help in keeping a record of the parameters exhibited during the evaluation of the roots of Selinum vaginatum. Methods: The organoleptic characteristics were studied and physicochemical tests, phytochemical analysis, fluorescence analysis, TLC, UV, FTIR, NMR, and Electrospray Ionisation - Mass spectroscopy, HPLC and other parameters were carried out for the standardization of the test drug. The procedures were carried out as per the WHO standards. The total ash values, swelling index, foaming index, moisture content, alcohol, and water-soluble extractive values were calculated. The TLC profile of Selinum vaginatum was observed and the UV, FTIR, HPLC, NMR and Mass spectroscopies of the plant were also performed. Results: The microscopic results showed the presence of schizogenous canals in abundance, cork cells, oil and globules, while medullary rays and pith were absent and around 10-17 layers of cells were seen. The presence of active compounds like phenols, terpenoids, alkaloids, coumarins, essential oils, tannins, saponins, carbohydrates, and acids was reported by phytochemical tests, TLC, UV, NMR, HPLC, Mass, and FTIR spectroscopy methods. These compounds can be studied in detail for understanding the potential pharmacological benefits and decoding the traditional uses. The spectroscopic analysis and physicochemical tests have shown the presence of phenols, esters, organic halogens, carbohydrate, carboxylic acids, alcohols, ethers, and glycosides which must be responsible for the medicinal properties of the plant. Conclusion: This study has provided a set of information that will be useful for the identification of fresh batches of the roots of Selinum vaginatum and this can be used as a reference set of information for accessing the purity, quality and identification, and analysis of the authenticity of the plant roots. Spectroscopic techniques have shown peaks, which on interpretation have reported the presence of active compounds - (coumarins like selenidin, angelicin, vaginidine, furocoumarin - oroselol and coumarin derivative- lomatin, isovaleric acid, Valeranone, Tricyclene, p-Cymene, α-Pinene, β-Pinene, Camphene, Borneol, p- Camphene, cholorgenic acid, β-Myrcene, Limonene and α-Pineneoxide which can be studied for understanding and identifying the pharmacological actions of the rhizomatous roots of S. vaginatum. All the information collected by research on the roots of S. vaginatum will help in the identification of crude drugs and also in the development of a standard for the drug Selinum vaginatum.

Background: The focus of this study was to prepare and characterize the single-chain variable fragment antibody (scFv)-coupled immunoaffinity column for purification of subtilisin BRC. Methods: The scFv against subtilisin BRC was immobilized onto CNBr-activated Sepharose 4B. Adsorption isotherm for subtilisin BRC on scFv-BRC-coupled Sepharose 4B was obtained and calculated the maximum binding capacity. The extraction conditions, including eluting solution, the concentration of eluting solution and flow rate, were optimized. Under the optimized eluting conditions, the dynamic binding capacity of the immunoaffinity column was determined. Results: The scFv-BRC-coupled Sepharose 4B for immunoaffinity purification of subtilisin BRC was prepared. The coupling efficiency was about 78.4%, e.g. about 8 mg of scFv-BRC was covalently coupled to 1 g CNBr-activated Sepharose 4B. The maximum equilibrium binding capacity (qm) and dissociation constant (Kd) of the immunoaffinity column for subtilisin BRC were 3.01 mg/mL and 0.465 mg/mL, respectively. The immunoaffinity chromatography conditions were optimized and the subtilisin BRC was purified 3.29-fold with 55.6%. Conclusion: The subtilisin BRC was effectively purified with high purity using scFv-BRC-coupled Sepharose 4B and the dynamic binding capacity of the column was determined. These results suggested that scFv-BRC can be used as a ligand for affinity purification of subtilisin BRC.

Background: Due to the last two decades of drought disaster, which resulted in the loss of the main part of Urmia Lake water and changed the natural conditions of an environment, especially ionic strength. Objective: We aimed to isolate and characterize halophilic and halotolerant bacteria in Urmia Lake, Iran, 2015. Urmia Lake is a permanent and salty inland lake located in the Azerbaijan region in northwestern Iran. Methods: Sampling was carried out in multiple water-filled locations of the lake. Liquid basal media for the enrichment of bacteria was successively applied and colonies were isolated by the plating method. Isolates were then distinguished based on differences in colony, Gram staining, microscopic shape, and biochemical properties. Results: One chemolithotrophic isolate belonging to Thiobacillus thioparus and 41 heterotrophic isolates were obtained. The 16S rRNA gene sequences showed that all 42 isolates belong to the genera Kocuria (21.42%), Marinobacter (11.90%), Micrococcus (11.90%), Thalassobacillus (11.90%), Bacillus (11.90%), Halomonas (7.14%) and Thiobacillus (2.38%). Conclusion: Based on 16S rRNA similarity, 5 of 41 isolates showed the potential to be introduced as new species. The dominant genera with abounded frequency were found to be Kocuria, Bacillus and Thalassobacillus genera.

Background: Centella asiatica (L.) is an herbaceous, frost-tender perennial plant belonging to the Apiaceae family. It is rich in important secondary metabolites such as terpenoids, flavonoids and saponins. It is used as a traditional medicine for skin infection, cough and asthma. Methods: The present study was intended at the quantitative and qualitative analysis of its secondary metabolites. Thin Layer Chromatography (TLC) was performed to separate the compounds according to their Rf (Retention factor) values. Results: The total phenolic content of ethanol leaf extract was 45.72 ± 6.05 mg GAE/g determined by Folin Ciocalteu’s assay. The maximum content of total flavonoids was found in aqueous leaf extract, 44.62 ± 2.45 mg QE/g. Aqueous extract of leaf and stem showed the highest antioxidant activity with DPPH (2,2- diphenyl-1-picrylhydrazyl) scavenging assay (45.66 ± 1.29 & 45.20 ± 3.0, respectively). Conclusion: The present study revealed the antioxidant properties in Centella asiatica.

Background: Azo dyes are widely used recalcitrant chemicals and may promote environmental hazards. Amongst the known azo dyes is CI Direct Red 28 (C32H22N6Na2O6S2) aka Congo Red (CR), which is a known mutagen and carcinogen. In this regard, this work aimed at the biodegradation of a CR-containing synthetic effluent, and the evaluation of the ecotoxicity of post-treatment residues. Methods: Lentinus sp. Laccase (Lac)-mediated bioremediation of CR was optimized upon added concentrations of sucrose and CuSO4, moreover a standard ecotoxicity assay was performed. Results: Results showed that the addition of 5% sucrose and 2 mM CuSO4 increased CR degradation, with Lac activity at 48 h of 30.2 U mL-1, and at 72 h of 31.9 U mL-1. Moreover, the ecotoxicity assay showcased that CR degradation by Lentinus sp. Lac seemingly generated low ecotoxic byproducts. Conclusion: Given that CR bioremediation byproducts were known to exhibit high toxicity, our results shed light on the use of Lentinus sp. catalytic arsenal to promote proper remediation of azo dyepolluted effluents.

Background: Proteases play a crucial role in the pharmacological properties of latex producing plants. Some of them exhibited intervention with fibrinogenolysis and/or fibrinolysis, two crucial wound healing events. Objective: To evaluate wound healing potential of crude and partially purified enzyme from Tabernaemontana divaricata (stem and latex). Materials and Methods: Proteolytic activity, clot inducing/dissolving potential, fibrinogen polymerization, recalcification time, blood clot lysis and Tricine-SDS PAGE for enzyme treated fibrinogen and human plasma clot were performed. Results: Latex PPE exhibited significant proteolytic activity (115.8 ± 0.3 U/ml) compared to that of the stem (28.78 ± 0.2 U/ml). Enzyme preparations exhibited temporally spaced clot inducing and subsequent dissolving properties favoring hemostatic effect, procoagulant effect being dominant and the first event. Significant reduction in fibrinogen absorbance at 540 nm with time, recalcification time and human fibrinogenolytic product analysis on Tricine PAGE substantiated procoagulant effect. Disappearance of Aα and Bβ fibrinopeptides by both stem and latex PPEs in the PAGE was observed. γ subunits were completely hydrolysed by latex PPE, however, it showed comparative resistance to stem PPE. Reduction in blood clot weight and fibrin subunit intensity supported thrombolytic property. Conclusion: The study provides evidence of the procoagulant and thrombolytic activity associated with T. divaricata proteases.

Background: Polyhydroxyalkanoates (PHA) are polyesters produced by various microorganisms. The major drawback of PHA, which is restricting its large-scale production in industries, is its high cost of raw materials and lower PHA concentration. Objective: The present study aimed to enhance the production of PHA from Wickerhamomyces anomalus VIT-NN01 by optimizing various nutrient and environmental stress conditions and its characterization. Methods: Effect of various stress conditions viz. nitrogen limitation, salinity, chemical mutagens (acridine orange, sodium azide), and physical stresses (UV, Low Electric Current (LEC), Magnetic Field Intensity (MFI), sound waves) were optimized to screen the best strategic growth conditions for the maximum accumulation of PHA in W. anomalus VIT-NN01. Instrumental analysis was done to evaluate the various changes that occurred in the treated cells and extracted PHA. Results: The maximum PHA content was observed on the effect of sound waves (88.74%), followed by LEC (87.8%) and MFI (85.75%). The morphological changes in length, shape, and size of the treated W. anomalus cells were observed by Transmission Electron Microscopy (TEM). Smooth, porous matrix, and pseudo spherical microstructure of the extracted PHA were observed by scanning electron microscopy and TEM analysis. The extracted polymer was identified as poly(3-hydroxybutyrate-co-3- hydroxyvalerate) [P(3HB-co-3HV)] co-polymer comprised of 51.66% 3HB and 48.33% 3HV monomer units based on gas chromatography-mass spectrophotometer and nuclear magnetic resonance spectroscopic analysis. X-ray diffraction analysis revealed the crystalline nature of the extracted P(3HB-co- 3HV). The degradation and melting temperatures were found to be 275.9 and 113.8°C, respectively, through thermogravimetric and differential scanning calorimetry analysis. Conclusion: These results supported the potentiality of W. anomalus, which tolerated the stress conditions and enhanced P(3HB-co-3HV) production from 60 to 88.74% and showed the novelty of present work. This is the first report elucidating the importance of physical stress conditions viz. low electric current, magnetic field intensity, and sound waves for the significant enhancement of PHA production in yeast.