Indirect Measurement of Variables in a Heterogeneous Reaction for Biodiesel Production.

This research focuses on the development of a state observer for performing indirect measurements of the main variables involved in the soybean oil transesterification reaction with a guishe biochar-based heterogeneous catalyst; the studied reaction takes place in a batch reactor. The mathematical model required for the observer design includes the triglycerides' conversion rate, and the reaction temperature. Since these variables are represented by nonlinear differential equations, the model is linearized around an operation point; after that, the pole placement and linear quadratic regulator (LQR) methods are considered for calculating the observer gain vector L(x). Then, the estimation of the conversion rate and the reaction temperature provided by the observer are used to indirectly measure other variables such as esters, alcohol, and byproducts. The observer performance is evaluated with three error indexes considering initial condition variations up to 30%. With both methods, a fast convergence (less than 3 h in the worst case) of the observer is remarked.

Gum resin of Boswellia serrata inhibited human monocytic (THP-1) cell activation and platelet aggregation.

ETHNOPHARMACOLOGICAL RELEVANCE: Stem bark gum resin extract of Boswellia serrata is traditionally used in India for its hemostatic, antiinflammatory and cardiovascular health effects and it is named as Sallaki in Ayurvedic medicine. AIM OF THE STUDY: This study was conducted to evaluate the antioxidative and antithrombotic properties of stem bark gum resin extracts of Boswellia serrata (BS). MATERIALS AND METHODS: The inhibitory activity of the BSWE and BSAE on FeCl(3) induced lipid peroxidation (in vitro) in rat liver and heart homogenates was measured spectrophotometrically. Their effect on H(2)O(2) induced reactive oxygen species (ROS) generation in human monocytic (THP-1) cells was investigated by tracking intensity of a cell permeable fluorescent dye, H(2)DCFDA and subjecting the cell samples to confocal microscopy. Further, the effect of BSAE and BSWE on ADP-induced platelet aggregation was assessed using a multimode detection plate reader, plasma coagulation times using an automated blood coagulation analyzer and on human blood clotting factors Xa and XIa using chromogenic substrate. Phytomarker analysis of the water (BSWE) and hydroalcoholic (BSAE) extracts of BS-gum resin was done through HPLC using a standard compound AKßBA. RESULTS: BSAE and BSWE inhibited, to varied extents, the lipid peroxidation in liver (80%) and heart (50%) tissue homogenates of male Wistar rats. Further, BSAE (30 µg dwt/mL) and BSWE (300 µg dwt/mL) attenuated ≥ 60% of H(2)O(2) mediated ROS generation in THP-1 cells. In case of standard compounds, ascorbate (20 µg dwt/mL) and butylated hydroxytoluene (BHT) (10 µg dwt/mL) completely scavenged ROS in the cells. BSAE and BSWE at 3 mg dwt/mL completely inhibited ADP induced platelet aggregation and activities were comparable to 20 µg/mL of heparin. The extracts also showed very high activity in prolonging coagulation time periods. Both types of extracts extended prothrombin time (PT) from ∼13 to >60s and activated partial thromboplastin time (APTT) from ∼32s to >90s. BSAE inhibited clotting factors Xa and XIa remarkably at 6 µg of dwt where as BSWE did not show much effect on FXa and showed 30% inhibition on FXIa at 120 µg. 10 µg of heparin was required to inhibit about 30% activity of the above factors. HPLC analyses suggested that BSAE and BSWE had AKßBA of 9% (w/w) and 7.8% (w/w) respectively. CONCLUSION: Present study demonstrated antioxidant and antithrombotic anticoagulant activities of water and hydroalcoholic extracts of Boswellia serrata's gum resin. We suggest that BS-gum resin as a good source for lead/therapeutic compounds possessing antioxidant, antiplatelet and anticoagulant activities.

Bowman-Birk proteinase inhibitor from Cajanus cajan seeds: purification, characterization, and insecticidal properties.

A red gram proteinase inhibitor (RgPI) was purified from red gram ( Cajanus cajan ) seeds by using ammonium sulfate precipitation and ion-exchange, affinity, and gel filtration chromatography. SDS-PAGE under nonreducing condition revealed two protein bands with molecular masses of approximately 8.5 and approximately 16.5 kDa corresponding to monomeric and dimeric forms of RgPI, respectively. Similarly, matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry also confirmed the presence of dimer as well as other oligomeric forms: trimer, tetramer, and pentamer. Reduction of RgPI with dithiothreitol (DTT) led to the dissociation of the dimeric and oligomeric forms. Native-PAGE and two-dimensional gel electrophoresis indicated the existence of isoinhibitors with pI values of 5.95, 6.25, 6.50, 6.90, and 7.15, respectively. The MALDI-TOF-TOF mass spectrum and N-terminal sequence 'DQHHSSKACC' suggested that the isolated RgPI is a member of the Bowman-Birk inhibitor family. RgPI exhibited noncompetitive type inhibitory activity against bovine pancreatic trypsin and chymotrypsin, with inhibition constants of 292 and 2265 nM, respectively. It was stable up to a temperature of 80 degrees C and was active over a wide pH range between 2 and 12. However, reduction with DTT or 2-mercaptoethanol resulted in loss of inhibitory activity against trypsin and chymotrypsin. It also decreased the activity of larval midgut trypsin-like proteinases in Manduca sexta . Its insecticidal property was further confirmed by reduction in the growth and development of these larvae, when supplemented in the diet.

Purification and characterization of a Bowman-Birk proteinase inhibitor from the seeds of black gram (Vigna mungo).

A proteinase inhibitor (BgPI) was purified from black gram, Vigna mungo (cv. TAU-1) seeds by using ammonium sulfate fractionation, followed by ion-exchange, affinity and gel-filtration chromatography. BgPI showed a single band in SDS-PAGE under non-reducing condition with an apparent molecular mass of approximately 8kDa correlating to the peak 8041.5Da in matrix assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrum. BgPI existed in different isoinhibitor forms with pI values ranging from 4.3 to 6.0. The internal sequence "SIPPQCHCADIR" of a peak 1453.7 m/z, obtained from MALDI-TOF-TOF showed 100% similarity with Bowman-Birk inhibitor (BBI) family. BgPI exhibited non-competitive-type inhibitory activity against both bovine pancreatic trypsin (K(i) of 309.8nM) and chymotrypsin (K(i) of 10.7muM), however, with a molar ratio of 1:2 with trypsin. BgPI was stable up to a temperature of 80 degrees C and active over a wide pH range between 2 and 12. The temperature-induced conformational changes in secondary structure are reversed when BgPI was cooled from 90 to 25 degrees C. Further, upon reduction with dithiothreitol, BgPI lost both its inhibitory activity as well as secondary structural conformation. Lysine residue(s) present in the reactive site of BgPI play an important role in inhibiting the bovine trypsin activity. The present study provides detailed biochemical characteristic features of a BBI type serine proteinase inhibitor isolated from V. mungo.

Inhibitors from pigeonpea active against lepidopteran gut proteinases.

The proteinase inhibitors (PIs) active against bovine pancreatic trypsin, chymotrypsin, and insect midgut trypsin-like proteinases were found in the seeds of 14 cultivars and eight wild types of pigeonpea, Cajanus cajan L.. The inhibitory activity of PIs against trypsin and chymotrypsin, as well as their activity profile on gelatin-polyacrylamide gel electrophoresis (PAGE) were identical among the various cultivars. In contrast to cultivars, the wild types showed differences in inhibitory activity of PIs and their activity profile on gelatin-PAGE. The PIs from all cultivars and few wild types showed 10- to 50-fold higher activity against midgut trypsin-like proteinases of Achaea janata (L.) (Lepidoptera: Noctuidae), compared with bovine pancreatic trypsin. However, the PIs from both cultivars and wild types showed three- to nine-fold less activity against Spodoptera litura (F.) (Lepidoptera: Noctuidae) midgut trypsin-like proteinases, compared with bovine pancreatic trypsin. This inhibitory potential of PIs from cultivars and wild types, toward midgut trypsin-like proteinases from A. janata was further evident by the strong activity profile visualized on gelatin-PAGE. These results further suggest that the inhibitory potential of PIs from pigeonpea cultivars and wild types could be exploited in management of nonhost insects.

Essentiality of mitochondrial oxidative metabolism for photosynthesis: optimization of carbon assimilation and protection against photoinhibition.

The review emphasizes the essentiality of mitochondrial oxidative metabolism for photosynthetic carbon assimilation. Photosynthetic activity in chloroplasts and oxidative metabolism in mitochondria interact with each other and stimulate their activities. During light, the partially modified TCA cycle supplies oxoglutarate to cytosol and chloroplasts. The marked stimulation of O2 uptake after few minutes of photosynthetic activity, termed as light enhanced dark respiration (LEDR), is now a well-known phenomenon. Both the cytochrome and alternative pathways of mitochondrial electron transport are important in such interactions. The function of chloroplast is optimized by the complementary nature of mitochondrial metabolism in multiple ways: facilitation of export of excess reduced equivalents from chloroplasts, shortening of photosynthetic induction, maintenance of photorespiratory activity, and supply of ATP for sucrose biosynthesis as well as other cytosolic needs. Further, the mitochondrial oxidative electron transport and phosphorylation also protects chloroplasts against photoinhibition. Besides mitochondrial respiration, reducing equivalents (and ATP) are used for other metabolic phenomena, such as sulfur or nitrogen metabolism and photorespiration. These reactions often involve peroxisomes and cytosol. The beneficial interaction between chloroplasts and mitochondria therefore extends invariably to also peroxisomes and cytosol. While the interorganelle exchange of metabolites is the known basis of such interaction, further experiments are warranted to identify other biochemical signals between them. The uses of techniques such as on-line mass spectrometric measurement, novel mutants/transgenics, and variability in metabolism by growth conditions hold a high promise to help the plant biologist to understand this

Consequence of restricted mitochondrial oxidative metabolism on photosynthetic carbon assimilation in mesophyll protoplasts: Decrease in light activation of four chloroplastic enzymes.

The patterns of light activation of 4 chloroplastic enzymes were examined in mesophyll protoplasts of pea (Pisum sativum) in the absence or presence of oligomycin (inhibitor of oxidative phosphorylation) or antimycin A (inhibitor of cytochrome pathway) or salicylhydroxamic acid (SHAM, inhibitor of alternative pathway). The results were compared with those of DCMU (inhibitor of photosynthetic electron transport). The light activation of NADP glyceraldehyde-3-phosphate dehydrogenase (NADP-GAPDH), fructose-1,6-bisphosphatase (FBPase), phosphoribulokinase (PRK) (enzymes of the Calvin cycle) and NADP malate dehydrogenase (NADP-MDH) (reflects chloroplast redox state) was more pronounced at limiting CO2 (0.1 mM NaHCO3) than that at optimal CO2 (1.0 mM NaHCO3). SHAM decreased markedly (up to 33%) the light activation of all 4 enzymes, while antimycin A or oligomycin exerted only a limited effect (<10% decrease). Antimycin A or oligomycin or SHAM had no significant effect on light activation of these 4 enzymes in isolated chloroplasts. However, DCMU caused a remarkable decrease in light activation of enzymes in both protoplasts (up to 78%) and chloroplasts (up to 69%). These results suggest that the restriction of alternative pathway of mitochondrial metabolism results in a marked decrease in the light activation of key chloroplastic enzymes in mesophyll protoplasts but not in isolated chloroplasts. Such a decrease in the light activation of enzymes could be also a secondary feedback effect because of the restriction on carbon assimilation.