Inhibitors of ester hydrolysis as synergists for biological activity of Cecropia juvenile hormone.

In Manduca sexta pupae the sensitivity to exogenous Cecropia C18 juvenile hormone (JH) is governed in large part by the rate at which the hormone is metabolized. By analysing the distribution of labelled JH and its metabolites, when dispensed by injection in light mineral oil, I have shown that the half-life of JH at concentrations ranging from 0·02 to 1 μg/g live weight of insect is approximately 24 hr.When the same doses of JH are dispensed in mineral oil containing endocrinologically inactive carboxylesterase inhibitors, organophosphate, and carbamate insecticides, the hormone's half-life in certain cases is prolonged significantly. The formulations containing those materials which most effectively prolong the hormone's half-life give higher assay scores than equivalent doses of hormone in mineral oil alone.From these results it is inferred that the chemicals tested synergize the Cecropia juvenile hormone's biological activity by serving presumably as inhibitors of the principal enzyme responsible for the rapid breakdown of exogenous C18-JH in haemolymph—a carboxylesterase. Preliminary in vitro experiments with carboxylesterase-rich Manduca pupal blood further support this inference.     

Tomato (Solanum lycopersicum) growth and fruit quality affected by organic fertilization and ozonated water

Protoplasma - The use of modern and safe techniques to increase plant growth and yield is of significance. There is little data, to our knowledge, on the use of organic fertilization and ozonated...     

Overexpression and optimization of glutamate decarboxylase in Lactobacillus plantarum Taj‐Apis362 for high gamma‐aminobutyric acid production

Gamma‐aminobutyric acid (GABA) is an important bioactive compound biosynthesized by microorganisms through decarboxylation of glutamate by glutamate decarboxylase (GAD). In this study, a full‐length GAD gene was obtained by cloning the template deoxyribonucleic acid to pTZ57R/T vector. The open reading frame of the GAD gene showed the cloned gene was composed of 1410 nucleotides and encoded a 469 amino acids protein. To improve the GABA‐production, the GAD gene was cloned into pMG36e‐LbGAD, and then expressed in Lactobacillus plantarum Taj‐Apis362 cells. The overexpression was confirmed by SDS‐PAGE and GAD activity, showing a 53 KDa protein with the enzyme activity increased by sevenfold compared with the original GAD activity. The optimal fermentation conditions for GABA production established using response surface methodology were at glutamic acid concentration of 497.973 mM, temperature 36°C, pH 5.31 and time 60 h. Under the conditions, maximum GABA concentration obtained (11.09 mM) was comparable with the predicted value by the model at 11.23 mM. To our knowledge, this is the first report of successful cloning (clone‐back) and overexpression of the LbGAD gene from L. plantarum to L. plantarum cells. The recombinant Lactobacillus could be used as a starter culture for direct incorporation into a food system during fermentation for production of GABA‐rich products.     

Time-course changes in fungal elicitor-induced lignan synthesis and expression of the relevant genes in cell cultures of Linum album

Linum album has been shown to accumulate anti-tumor podophyllotoxin (PTOX) and its related lignans. In the present study, we examined the effects of five fungal extracts on the production of lignans in L. album cell cultures. Fusarium graminearum extract induced the highest increase of PTOX [140μgg(-1) dry weight (DW) of the L. album cell culture] which is seven-fold greater than the untreated control, while Rhizopus stolonifer extract enhanced the accumulation of lariciresinol, instead of PTOX, up to 365μgg(-1) DW, which was 8.8-fold greater than the control. Quantitative PCR analyses showed that expression of the enzyme genes responsible for the PTOX biosynthesis cascade, such as pinoresinol-lariciresinol reductase (PLR), phenylalanine ammonia-lyase (PAL), cinnamoyl-CoA reductase (CCR) and cinnamyl-alcohol dehydrogenase (CAD) genes, were also up-regulated in a fungal extract-selective fashion. These results provide evidence that the fungal extracts used in this study differentially increase the production of PTOX or larisiresinol via the up-regulation of the genes in lignan biosynthesis in L. album cell cultures, and suggest that such selective actions of fungal elicitors on the lignan synthesis will lead to more efficient metabolic engineering-based production of PTOX and other beneficial lignans using L. album cell cultures.     

Cloning and Characterization of a Plasmid Encoded ACC Deaminase from an Indigenous Pseudomonas fluorescens FY32

In addition to the characterized mechanisms responsible for many direct effects of plant growth promoting bacteria (PGPB) on plants, it has been suggested that a number of PGPB contain the enzyme ACC deaminase that catalyzes degradation of 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor of ethylene, into α-ketobutyrate and ammonia. As part of an effort to obtain an ACC deaminase encoding gene from a collection of soil samples, only one bacterial isolate, Pseudomonas fluorescens FY32 was capable of growing on ACC as a sole source of nitrogen. The ACC deaminase gene was amplified from the above isolate by polymerase chain reaction (PCR) giving an expected DNA fragment, 1017 bp. Sequence analysis of the fragment showed that it was highly homologous (94% and 98% identities at nucleotide and amino acid levels, respectively) to the previously characterized acdS gene from Pseudomonas sp. 6G5. Furthermore, fusion of the ACC deaminase ORF with lacZ gene resulted in the expression of active enzyme in Escherichia coli. In addition, further analyses revealed that the acdS gene was plasmid-encoded so that a large plasmid (pFY32) with almost 50 kb in size was identified from this bacterium. Furthermore, transfer of pFY32 into E. coli DH5α proved its ACC deaminase activity. This result was in accordance with previous reports suggesting horizontal transfer of the acdS gene. However, it needs more investigation to identify whether this pFY32 plasmid has undergone lateral gene transfer during the evolutionary process.     

Colonization with arbuscular mycorrhizal fungi improves salinity tolerance of tomato (Solanum lycopersicum L.) plants

The purpose of this study was to investigate the mechanisms underlying alleviation of salt stress by mycorrhization. Solanum lycopersicum L. cultivars Behta and Piazar with different salinity tolerance were cultivated in soil without salt (EC?=?0.63 dSm^?1), with low (EC?=?5 dSm^?1), or high (EC?=?10 dSm^?1) salinity. Plants inoculated with the arbuscular mycorrhizal fungi Glomus intraradices (+AMF) were compared to non-inoculated plants (?AMF). Under salinity, AMF-mediated growth stimulation was higher in more salt tolerant Piazar than in sensitive Behta. Mycorrhization alleviated salt-induced reduction of P, Ca, and K uptake. Ca/Na and K/Na ratios were also better in +AMF. However, growth improvement by AMF was independent from plant P nutrition under high salinity. Mycorrhization improved the net assimilation rates through both elevating stomatal conductance and protecting photochemical processes of PSII against salinity. Higher activity of ROS scavenging enzymes was concomitant with lowering of H₂O_2, less lipid peroxidation, and higher proline in +AMF. Cultivar differences in growth responses to salinity and mycorrhization could be well explained by differences in ion balance, photochemistry, and gas exchange of leaves. Function of antioxidant defenses seemed responsible for different AMF-responsiveness of cultivars under salinity. In conclusion, AMF may protect plants against salinity by alleviating the salt-induced oxidative stress.     

Quantifying the HIV-1 integrase inhibitor raltegravir in female genital tract secretions using high-performance liquid chromatography with ultraviolet detection

Understanding the pharmacokinetics of drugs in peripheral body compartments, such as the genital tract, is particularly important in the infectious diseases arena. However, extracting drugs from small volumes of viscous, proteinacious substances like cervicovaginal fluid is particularly challenging. The goal of this study was to develop a method to quantify raltegravir, an HIV-1 integrase inhibitor, in the female genital tract. The method included sample preparation with perchloric acid followed by solid-phase extraction, separation with reverse-phase high-performance liquid chromatography, and detection with an ultraviolet wavelength of 218 nm. The method was linear from 0.05 to 10.0 mg/L, with minimal endogenous interference. The method was accurate (1.2–11.0% deviation) and precise (1.1–12.6% CV) for both within and between-day analyses. The ability to detect raltegravir in the female genital tract is essential for future investigations of raltegravir as an agent for prevention of HIV acquisition, and this method will be used for clinical studies further evaluating pharmacokinetic–pharmacodynamic relationships in this body compartment.     

Use of hydrophilic natural gums in formulation of sustained-release matrix tablets of tramadol hydrochloride

The objective of this work was to develop matrix sustained-release tablets of highly water-soluble tramadol HCl using natural gums (xanthan [X gum] and guar [G gum]) as cost-effective, nontoxic, easily available, and suitable hydrophilic matrix systems compared with the extensively investigated hydrophilic matrices (ie, hydroxypropyl methylcellulose [HPMC]/carboxymethyl cellulose [CMC] with respect to in vitro drug release rate) and hydration rate of the polymers. Matrix tablets of tramadol (dose 100 mg) were produced by direct compression method. Different ratios, of 100∶0, 80∶20, 60∶40, 20∶80, 0∶100 of G gum (or X):HPMC, X gum:G gum, and triple mixture of these polymers (G gum, X gum, HPMC) were applied. After evaluation of physical characteristics of tablets, the dissolution test was, performed in the phosphate buffer media (pH 7.4) up to 8 hours. Tablets with only X had the highest mean dissolution time (MDT), the least dissolution efficiency (DE₈%), and released the drug following a zero-order model via swelling, diffusion, and erosion mechanisms. Guar gum alone could not efficiently control the drug release, while X and all combinations of natural gums with HPMC could retard tramadol HCl release. However, according to the similarity factor (f ₂), pure HPMC and H₈G₂ were the most similar formulations to Topalgic-LP as the reference standard. Published: March 17, 2006     

Adriamycin release from poly(lactide-coglycolide)-polyethylene glycol nanoparticles: synthesis, and in vitro characterization

The preparation, properties, and application in adriamycin delivery ofbiocompatible and biodegradable poly(lactide-co-glycolide)-polyethylene glycol (PLGA-PEG) nanoparticles are discussed. PLGA-PEG copolymers were synthesized by ring opening polymerization of the dl-lactide and glycolide in the presence of PEG1000. 1H-NMR and FT-IR spectrum were consistent with the structure of PLGA-PEG copolymers. The adriamycin-loaded nanoparticles could be prepared using a precipitation-solvent evaporation technique. The nanoparticles have been produced by a precipitation-solvent evaporation technique. The physical characteristics and drug loading efficiency of the PLGA-PEG nanoparticles were influenced by the composition of the PLGA-PEG copolymers used to prepare the nanoparticles. Particle sizes were between 65 and 100 nm for different compositions of PLGA-PEG copolymers. PLGA-PEG nanoparticles prepared from copolymers having relatively high PLGA/PEG ratios were smaller. Entrapment efficiency was 25%-33%. Adriamycin release from the nanoparticles at pH 7.4 showed an initial burst release and then sustained release phase. These results showed that PLGA-PEG nanoparticles could be an effective carrier for cancer therapy.