Dissociation of enzymatic and pharmacological properties of piratoxins-I and -III, two myotoxic phospholipases A(2) from Bothrops pirajai snake venom

Piratoxins (PrTX) I and III are phospholipases A(2) (PLA(2)s) or PLA(2) homologue myotoxins isolated from Bothrops pirajai snake venom, which also induce myonecrosis, bactericidal activity against Escherichia coli, disruption of artificial membranes, and edema. PrTX-III is a catalytically active hemolytic and anticoagulant Asp49 PLA(2), while PrTX-I is a Lys49 PLA, homologue, which is catalytically inactive on artificial substrates, but promotes blockade of neuromuscular transmission. Chemical modifications of His, Lys, Tyr, and Trp residues of PrTX-I and PrTX-III were performed, together with cleavage of the N-terminal octapeptide by CNBr and inhibition by heparin and EDTA. The lethality, bactericidal activity, myotoxicity, neuromuscular effect, edema inducing effect, catalytic and anticoagulant activities, and the liposome-disruptive activity of the modified toxins were evaluated. A complex pattern of functional differences between the modified and native toxins was observed. However, in general, chemical modifications that significantly affected the diverse pharmacological effects of the toxins did not influence catalytic or membrane disrupting activities. Analysis of structural changes by circular dichroism spectroscopy demonstrated significant changes in the secondary structure only in the case of N-terminal octapeptide cleavage. These data indicate that PrTX-I and PrTX-III possess regions other than the catalytic site, which determine their toxic and pharmacological activities. (C) 2001 Academic Press.

Characterization of glycerol kinase from baker's yeast: Response surface modeling of the enzymatic reaction

The present study describes a methodology of dosage of glycerol kinase (GK) from baker's yeast. The standardization of the activity of the glycerol kinase from baker's yeast was accomplished using the diluted enzymatic preparation containing glycerol phosphate oxidase (GPO) and glycerol kinase. The mixture was incubated at 60 degrees C by 15 min and the reaction was stopped by the SDS solution addition. A first set of experiments was carried out in order to investigate the individual effect of temperature (7), pH and substrate concentration (S), on GK activity and stability. The pH and temperature stability tests showed that the enzyme presented a high stability to pH 6.0-8.0 and the thermal stability were completely maintained up to 50 degrees C during 1 h. The K(m) of the enzyme for glycerol was calculated to be 2 mM and V(max) to be 1.15 U/mL. In addition, modeling and optimization of reaction conditions was attempted by response surface methodology (RSM). Higher activity values will be attained at temperatures between 52 and 56 degrees C, pH around 10.2-10.5 and substrate concentrations from 150 to 170 mM.This low cost method for glycerol kinase dosage in a sequence of reactions is of great importance for many industries, like food, sugar and alcohol. RSM showed to be an adequate approach for modeling the reaction and optimization of reaction conditions to maximize glycerol kinase activity. (C) 2007 Elsevier B.V. All rights reserved.

Enzymatic variability among venoms from different subspecies of Apis mellifera (Hymenoptera: Apidae)

The enzymatic variability was analyzed in venom extracts from bees reared in different colonies of the Africanized, A. m. ligustica and A. m. carnica subspecies. The implications of this variation focused on the biochemistry differentiation and immunogenicity of these venoms. The results showed the existence of a huge variability among the subspecies as well as among the colonies for three out of the six tested components - hyaluronidase, acid phosphatase and proteases - suggesting the utilization of these features as possible biochemical markers. Furthermore, although not statistically significant, it was found that the Africanized bee venom presented slightly higher levels of protein content and esterase activity, when compared to the other subspecies. If the esterase plays a role in the pain intensity caused by the sting, as suggested elsewhere, this might suggest a reason for a bigger algogenicity of this venom in relation to that of European bees. On the other hand, A. m. ligustica bees presented the highest levels of proteolytic and acid phosphatase activities, whose functions are not enlightened in Hymenoptera venoms. The A. m. carnica workers presented the highest hyaluronidase and the lowest acid phosphatase activity levels. The extremely variable results among colonies of the same subspecies and among subspecies, for the tested venom components, justify the absence of correlation between allergic reactions and tests with pooled venom.

Antioxidant enzyme activity and hydrogen peroxide content during the drying of Arabica coffee beans

The development of the germination process and drought stress during the drying of coffee can generate reactive oxygen species, which can be neutralized by way of antioxidant mechanisms. No studies related to antioxidant enzymes during the drying of coffee were found in the literature, and considering their importance, the enzymatic activities of superoxide dismutase (SOD), guaiacol peroxidase (GPOX) and glutathione reductase (GR), and also the hydrogen peroxide content were evaluated during the drying of two types of coffee bean, one processed as natural coffee and the other as pulped natural coffee. The results showed a reduction in the SOD, GPOX and GR enzymatic activities of the natural coffee as compared to the pulped natural coffee during the drying period. Moreover, the hydrogen peroxide content of the natural coffee was greater than that of the pulped natural coffee. These results suggest the development of oxidative stress during the coffee drying process, controlled more efficiently in pulped natural coffee by the early action of GPOX during the drying process. Nevertheless, differential responses by SOD isoenzymes and possibly the role of other peroxidases also appear to be involved in the responses observed. © 2013 Springer-Verlag Berlin Heidelberg.

Xylo-oligosaccharides from lignocellulosic materials: Chemical structure, health benefits and production by chemical and enzymatic hydrolysis

Currently, there is worldwide interest in the technological use of agro-industrial residues as a renewable source of food and biofuels. Lignocellulosic materials (LCMs) are a rich source of cellulose and hemicellulose. Hemicellulose is rich in xylan, a polysaccharide used to develop technology for producing alcohol, xylose, xylitol and xylo-oligosaccharides (XOSs). The XOSs are unusual oligosaccharides whose main constituent is xylose linked by β 1-4 bonds. The XOS applications described in this paper highlight that they are considered soluble dietary fibers that have prebiotic activity, favoring the improvement of bowel functions and immune function and having antimicrobial and other health benefits. These effects open a new perspective on potential applications for animal production and human consumption. The raw materials that are rich in hemicellulose include sugar cane bagasse, corncobs, rice husks, olive pits, barley straw, tobacco stalk, cotton stalk, sunflower stalk and wheat straw. The XOS-yielding treatments that have been studied include acid hydrolysis, alkaline hydrolysis, auto-hydrolysis and enzymatic hydrolysis, but the breaking of bonds present in these compounds is relatively difficult and costly, thus limiting the production of XOS. To obviate this limitation, a thorough evaluation of the most convenient methods and the opportunities for innovation in this area is needed. Another challenge is the screening and taxonomy of microorganisms that produce the xylanolytic complex and enzymes and reaction mechanisms involved. Among the standing out microorganisms involved in lignocellulose degradation are Trichoderma harzianum, Cellulosimicrobium cellulans, Penicillium janczewskii, Penicillium echinulatu, Trichoderma reesei and Aspergillus awamori. The enzyme complex predominantly comprises endoxylanase and enzymes that remove hemicellulose side groups such as the acetyl group. The complex has low β-xylosidase activities because β-xylosidase stimulates the production of xylose instead of XOS; xylose, in turn, inhibits the enzymes that produce XOS. The enzymatic conversion of xylan in XOS is the preferred route for the food industries because of problems associated with chemical technologies (e.g., acid hydrolysis) due to the release of toxic and undesired products, such as furfural. The improvement of the bioprocess for XOS production and its benefits for several applications are discussed in this study. © 2012 Elsevier Ltd.

Storage of sugarcane stalks (Saccharum officinarum cv. SP 79-1011) in low oxygen atmospheres and the effects on enzymatic browning

Although sugarcane consumption is very popular worldwide there are few studies regarding the postharvest storage of stalks that focus on controlling enzymatic browning. The objective of this study was to evaluate the quality of sugarcane stalks stored at 10±1°C in controlled atmosphere with various oxygen (O2) levels (1kPa, 5kPa, 10kPa, 15kPa, and 21kPa) and to verify the effect of this storage on the activities of polyphenoloxidase (PPO) and peroxidase (POD). At 1kPa and 5kPa O2, the stalks' respiratory rate, POD activity, and reducing sugar content were significantly less (P<0.05) than at higher O2 concentrations. In contrast, low O2 levels did not affect PPO activity or influence cane coloration. These results can be used to guide the choice of plastic films, making it possible to store sugarcane stalks in a controlled atmosphere. © 2013 Elsevier B.V.

Activity of some isoenzymatic systems in stored coffee grains

Considering the worldwide consumption of coffee, it is natural that throughout the history many people have dedicated the research to markers that contribute somehow on gauging its quality. This research aimed to evaluate the biochemical performance of arabica coffee during storage. Coffee in beans (natural) and in parchment (pulped) dried in concrete terrace and in dryer with heated air were packed in jute bags and stored in not controlled environmental conditions. Enzymatic activities of superoxide dismutase, catalase, peroxidase, polyphenoloxidase, esterase and lipoxygenase in coffee grains were evaluated at zero, three, six, nine and twelve months by means of electrophoresis. Independently of the drying method, the activity of isoenzymatic complexes highlighted deteriorative processes in stored grains of coffee. The treatments 60/40º C and 60º C used to reduce the water content imposed a greater stress condition, accelerated metabolism of natural coffee in the storage with decreased activity of defense mechanisms due to latent damage in these grains. Natural coffees are more sensible to high drying temperatures and its quality reduces faster than pulped coffee in the storage.

Enteral nutrition feeding alters antioxidant activity in unstimulated whole saliva composition of patients with neurological disorders

Patients with neurological disorders have an increased risk of oral and systemic diseases due to compromised oral hygiene. If patients lose the ability to swallow and chew food as a result of their disorder, enteral nutrition is often utilized. However, this type of feeding may modify salivary antioxidant defenses, resulting in increased oxidative damage and the emergence of various diseases. The aim of this study was to evaluate the effects of enteral nutrition on biochemical parameters in the unstimulated whole saliva composition of patients with neurological disorders. For this, enzymatic (superoxide dismutase - SOD; glutathione peroxidase - GPx) and non-enzymatic (uric acid; ferric ion reducing antioxidant power - FRAP) antioxidant activity, as well as a marker for oxidative damage (thiobarbituric acid reactive substances - TBARS) were analyzed. Unstimulated whole saliva was collected from 12 patients with neurological disorders and tube-feeding (tube-fed group - TFG), 15 patients with neurological disorders and normal feeding via the mouth (non-tube-fed group - NTFG), and 12 volunteers without neurological disorders (control group - CG). The daily oral hygiene procedures of TFG and NTFG patients were similar and dental care was provided monthly by the same institution's dentist. All patients exhibited adequate oral health conditions. The salivary levels of FRAP, uric acid, SOD, GPx, TBARS, and total protein were compared between studied groups. FRAP was increased (p < 0.05) in the NTFG (4651 +/- 192.5 mmol/mL) and the TFG (4743 +/- 116.7 mmol/mL) when compared with the CG (1844 +/- 343.8 mmol/mL). GPx values were lower (p < 0.05) in the NTGF (8.24 +/- 1.09 mmol/min/mg) and the TFG (8.37 +/- 1.60 mmol/min/mg) than in the CG (15.30 +/- 2.61 mmol/min/mg). Uric acid in the TFG (1.57 +/- 0.23 mg/dL) was significantly lower than in the NTFG (2.34 +/- 0.20 mg/dL) and the CG (3.49 +/- 0.21 mg/dL). Protein was significantly lower in the TFG (5.35 +/- 0.27 g/dL) than in the NTFG (7.22 +/- 0.57 g/dL) and the CG (7.86 +/- 0.54 g/dL). There was no difference in the salivary flow rate and SOD between groups. Enteral nutrition in patients with neurological disorders was associated with lower oxidative damage, resulting in increased salivary. antioxidant capacity. These results emphasize the importance of oral care for this population to prevent oral and systemic diseases. (C) 2014 Elsevier Ltd. All rights reserved.

Enzymatic production of functional oligosaccharides - A review

Interest in oligosaccharide production and its general characteristics is growing. The physiological effects resulting from its ingestion make them more attractive than its sweetness, and the versatility of these carbohydrates allows their use for human and animal nutrition, pharmacology and the cosmetics industry, among others. Several microorganisms are involved in enzyme production to create oligomers with biological activity, including fructooligosaccharides, galactooligosaccharides and aminoglucanoligosaccharides. Some oligomers are currently on the market, but the search for new microorganisms producing enzymes, high-yield processes for obtaining oligosaccharides, different physiological effects, and a correlation between chemical structure and function continues.

Identification and enzymatic characterization of acid phosphatase from Burkholderia gladioli

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The long-term administration of calcineurin inhibitors decreases antioxidant enzyme activity in the rat parotid and submandibular salivary glands

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)