Evaluation of antimicrobial activity of extracts of in vivo and in vitro grown Vinca rosea L. (Catharanthus roseus) against pathogens.

The antimicrobial activity of Vinca rosea was evaluated against pathogenic bacterial strains (Bacillus subtilis, B. licheniformis and Azotobacter sp.) and fungal strains (Asprgillus niger, Alternaria solani and Rhizopus oryzae) using agar well diffusion method. Methanolic extracts of in vivo leaf, in vitro leaf, in vitro calluses of leaf, nodal and fruit explants were used and exhibited antimicrobial activity as indicated by minimum inhibitory concentration (MIC). In vitro extracts showed better results as compared to the in vivo extracts for both the antibacterial as well as the antifungal activity. Among all the extracts, maximum zone of inhibition (30.3 mm ± 0.58(a)) was formed by in vitro leaf callus extract concentration of 2.0mg/ml against B. licheniformis. Similarly in case of antifungal activity, maximum zone of inhibition (34.6mm ± 0.57(a)) was formed by in vitro leaf callus extract and MIC value is 6.0mg/ml against A. niger. Hence these results clearly depicts that V. rosea possess a great strength to fight against the microbial activity and can be used against various infections.

Photoremovable protecting groups as controlled-release device for sex pheromone.

Photoremovable protecting groups (PRPGs) were demonstrated as a delivery device for controlled release of pheromone under both UV light (≥350 nm) and direct sunlight irradiation. In the present work, (Z)-11-hexadecen-1-ol (sex pheromone of Chilo infuscatellus Snellen) was chemically caged by four different photoremovable protecting groups (7-hydroxy-4-hydroxymethylcoumarin, 1-pyrenemethanol, 9-anthracenemethanol and 2-(hydroxymethyl)anthraquinone) individually. Photophysical studies showed that the caged pheromone with coumarin, pyrene and anthracene derivatives exhibited strong fluorescence. Controlled release of (Z)-11-hexadecen-1-ol was achieved by irradiating the caged compounds in aqueous ethanol both under UV and sunlight. Further, to mimic the environmental conditions, controlled release of (Z)-11-hexadecen-1-ol was also studied in soil medium under direct sunlight. Thermogravimetric analysis showed that caging of (Z)-11-hexadecen-1-ol by PRPGs significantly reduced its volatility. Bioassay experiments indicated that PRPGs are harmless to soil bacteria (Azotobacter sp. and Pseudomonas aeruginosa) and in vitro cytotoxicity studies on eukaryotic L929 cells showed that PRPGs are also non-toxic. Field bioassays were performed using caged pheromone against maize stalk borer (Chilo partellus) and the results showed that the caged alcohol is effective in a number of moths catches instead of free alcohol in a blend for a longer period of time. Our studies indicated that use of PRPGs as delivery device for controlled release of pheromone by sunlight holds great interest for field applications.

Effects of culture conditions on the production of polyhydroxyalkanoates by Azotobacter chroococcum H23 in media containing a high concentration of alpechín (wastewater from olive oil mills) as primary carbon source.

Large amounts of homopolymers containing beta-hydroxybutyrate (PHB) and copolymers containing beta-hydroxyvalerate (P[HB-co-HV]) are produced by Azotobacter chroococcum strain H23 when growing in culture media amended with alpechín (wastewater from olive oil mills) as the sole carbon source. Copolymer was formed when valerate (pentanoate) was added as a precursor to the alpechín medium, but it was not formed with the addition of propionate as a precursor. A. chroococcum formed homo- and copolymers of polyhydroxyalkanoates (PHAs) up to 80% of the cell dry weight, when grown on NH(4)(+)-medium supplemented with 60% (v/v) alpechín, after 48 h of incubation at 100 rev min(-1) and 30 degrees C. Production of PHAs by strain H23 using alpechín looks promising, as the use of a cheap substrate for the production of these materials is essential if bioplastics are to become competitive products.

Analogue-resistant mutants of Azotobacter chroococcum derepressed for nitrogenase activity and early ammonia excretion having potential as inoculants for cereal crops.

Spontaneous mutants resistant to methionine sulfoximine (Msx), methyl alanine (Mal) and methyl ammonium chloride (Mac) were derived from A. chroococcum strain A103. Msx and Mal-resistant mutants expressed 1.73 to 10.98% of the fully derepressed nitrogenase activity when grown in Burk's medium containing ammonium acetate. Mac-resistant mutants did not express nitrogenase activity in ammonium acetate supplemented medium. The mutants excreted ammonia even after 2 days of growth and some mutants excreted more ammonia as compared to the parent. Selected mutants were inoculated on wheat (Triticum aestivum) and barley (Hordeum vulgare) under field conditions. Majority of the derepressed mutants increased grain yield of wheat and barley varying from 1.2 to 33.3%. However, host-dependent effects on grain yield were observed with different mutants. Two mutants, Mal 27 and Mac 19 showed significant increase in grain yields of both the crops. The results suggest that metabolic analogue-resistant mutants of Azotobacter have potential for use as a biofertilizer for cereal crops.

Characteristics of N2 fixation in Mo-limited batch and continuous cultures of Azotobacter vinelandii.

Steady-state chemostat cultures of Azotobacter vinelandii were established in a simple defined medium that had been chemically purified to minimize Mo and that contained no utilizable combined N source. Growth was dependent on N2 fixation, the limiting nutrient being the Mo contaminating the system. The Mo content of the organisms was at least 100-fold lower than that of Mo-sufficient cultures, and they lacked the characteristic g = 3.7 e.p.r. feature of the MoFe-protein of nitrogenase. A characteristic of nitrogenase activity in vivo in Mo-limited populations was a disproportionately low activity for acetylene reduction, which was 0.3 to 0.1 of that expected from the rate of N2 reduction. Acetylene was also a poor substrate in comparison with protons as a substrate for nitrogenase, and did not markedly inhibit H2 evolution, in contrast with Mo-sufficient populations. In batch cultures in similar medium or 'spent' chemostat medium inoculated with Mo-limited organisms, the addition of Mo elicited a biphasic increased growth response at concentrations as low as 2.5 nM, provided that sufficient Fe was supplied. In this system V did not substitute for Mo, and Mo-deficient cultures ceased growth at a 25-fold lower population density compared with cultures supplemented with Mo. Nitrogenase component proteins could not be unequivocally detected by visual inspection of fractionated crude extracts of Mo-limited organisms. 35SO42-pulse-labelling studies also showed that the rate of synthesis of the MoFe-protein component of nitrogenase was too low to be quantified. However, the Fe-protein of nitrogenase was apparently synthesized at high rates. The discussion includes an evaluation of the possibility that A. vinelandii possesses an Mo-independent N2-fixation system.

Steady-state enzyme kinetics with high-affinity substrates or inhibitors. A statistical treatment of dose-response curves.

A statistical treatment of steady-state enzyme kinetic measurements is described that allows for depletion of free substrate or free inhibitor concentrations owing to significant binding to the enzyme. V(max.), K(m) or K(i), enzyme concentration, the concentration of substrate or inhibitor required for a half-maximal effect and standard errors of these parameters can be calculated from dose-response measurements; the concentration of each component of the system may be estimated also. The statistically best values of the parameters are used to convert dose-response curves into convenient linear forms. The method is applied to dose-response measurements of hydroxyquinoline N-oxide inhibition of bacterial respiration and aminopterin inhibition of dihydrofolate reductase. Two FORTRAN programs for this method have been deposited as Supplementary Publication no. SUP 50019 at the National Lending Library for Science and Technology, Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies may be obtained on the terms indicated in Biochem. J. (1973) 131, 5.

Chemical composition of Azotobacter vinelandii cysts.

Cysts of Azotobacter vinelandii ATCC 12837 were germinated by exposure to 3.0 mm ethylenediaminetetraacetic acid (EDTA)-tris(hydroxymethyl)aminomethane buffer at pH 7.8, and their outer coats (exines) were purified by differential and isopycnic centrifugation. Electron micrographs of exine showed it to consist of multilayers of a three-membered sheet structure whose thickness was 7.0 to 7.5 nm. The inner, less electron-dense layer (intine) was also prepared from cysts by EDTA treatment, centrifugation, concentration, and dialysis. The exine consisted of 32% carbohydrate, 28% protein, 30% lipid, and 3.2% ash, with the ash comprised of 1.62% calcium, 0.02% magnesium, and 0.34% phosphorus. The amino acid composition of exine was similar to that of gram-negative bacterial cell walls. The intine consisted of 44% carbohydrate, 9.1% protein, 37% lipid, and 4.1% ash, with the ash comprised of 2.45% calcium, 0.02% magnesium, and 0.38% phosphorus. The carbohydrates of both exine and intine contained glucose, mannose, xylose, and rhamnose. Glucosamine and galactosamine were found only in the exines. The fatty acids consisted of normal, iso, and anteiso saturated fatty acids with 10 to 18 carbon atoms and mono-unsaturated C(11), C(16), and C(18) fatty acids. The exines contained mostly bound lipid, but intines contained primarily free lipid.