Effect of Cervitec (R) on mutans streptococci in plaque and on caries formation on occlusal fissures of erupting permanent molars

The aim of this study was to evaluate the effect of Cervitec(R) on the abundance of mutans streptococci (MS) in occlusal dental plaque and on 2-year caries increment of partly erupting first permanent molars. Sixteen healthy schoolchildren aged 6-8 years, with at least 2 sound contralateral partly erupted permanent molars, received diet counselling and daily parental supervised toothbrushing with a fluoride dentifrice. Stimulated saliva samples were collected at baseline and after 1 year to evaluate MS levels. In a split-mouth design, Cervitec varnish was applied to one of the teeth at baseline and after 3 and 6 months, while the other tooth in the same jaw was a control. At the 9-month follow-up the teeth were in occlusal contact. At this time, varnish was not applied. At 3 and 6 months after the first application of varnish a significant suppression of MS was observed in plaque. Caries investigations, performed at baseline and every 3 months during the 2 years after the start of the study, showed that all the teeth treated with the varnish were free of caries after 2 years, whereas 8/16 control teeth developed incipient caries. In conclusion, our results suggest that treatment with Cervitec reduces MS in plaque on erupting permanent molars and can lead to a significant decrease in caries incidence. Copyright (C) 2002 S. Karger AG, Basel.

Purification, crystallization and Patterson search of haemoglobin IV from the armoured catfish Liposarcus anisitsi

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

Partial purification and characterization of pectin methylesterase from acerola (Malpighia glabra L.)

The enzyme pectin methylesterase (PME) is present in acerola fruit and was partially purified by gel filtration on Sephadex G-100. The results of gel filtration showed different PME isoforms. The total PME (precipitated by 70% salt saturation) and one of these isoforms (fraction from Sephadex G-100 elution) that showed a molecular mass of 15.5 +/- 1.0 kDa were studied. The optimum pH values of both forms were 9.0. The total and the partially purified PME showed that PME specific activity increases with temperature, the total acerola PME retained 13.5% of its specific activity after 90 min of incubation at 98 degreesC. The partially purified acerola (PME isoform) showed 125.5% of its specific activity after 90 min of incubation at 98 degreesC. The K-m values of the total PME and the partially purified PME isoform were 0.081 and 0.12 mg/mL, respectively. The V-max values of the total PME and the partially purified PME were 2.92 and 6.21 mumol/min/mL/mg of protein, respectively.

Partial purification, heat stability and kinetic characterization of the pectinmethylesterase from Brazilian guava, Paluma cultivars

Pectinmethylesterase (PME) was extracted from guava fruit (Psidium guajava L.), cultivar Paluma, by 70% ammonium sulphate saturation and partially purified by gel filtration on Sephadex G100. Gel filtration showed PME isoenzymes with different values of molecular mass. Two samples were examined: concPME (70% saturation by ammonium sulphate) and Iso4 PME (one of the isoforms from gel filtration with the greatest specific activity). Optimum pH of the enzyme (for both samples) was 8.5 and optimum temperature ranged from 75 and 85 degrees C. The optimum sodium chloride concentration was 0.15 M. The K-M and V-max ranged from 0.32 to 0.23 mg m1(-1) and 244 to 53.2 mu mol/min, respectively, for concPME and Iso4PME. The activation energies (E-a) were 64.5 and 103 kJ/mol, respectively, for concPME and Iso4PME. Guava PME, cv Paluma, is a very thermostable enzyme, showing great heat stability at all temperatures studied. (c) 2005 Elsevier Ltd. All rights reserved.

Purification and characterization of polygalacturonase produced by thermophilic Thermoascus aurantiacus CBMAI-756 in submerged fermentation

An extracellular polygalacturonase was isolated from 5-day culture filtrates of Thermoascus aurantiacus CBMAI-756 and purified by gel filtration and ion-exchange chromatography. The enzyme was maximally active at pH 5.5 and 60-65 degrees C. The apparent K (m) with citrus pectin was 1.46 mg/ml and the V (max) was 2433.3 mu mol/min/mg. The apparent molecular weight of the enzyme was 30 kDa. The enzyme was 100% stable at 50 degrees C for 1 h and showed a half-life of 10 min at 60 degrees C. Polygalacturonase was stable at pH 5.0-5.5 and maintained 33% of initial activity at pH 9.0. Metal ions, such as Zn+2, Mn+2, and Hg+2, inhibited 50, 75 and 100% of enzyme activity. The purified polygalacturonase was shown to be an endo/exo-enzyme, releasing mono, di and tri-galacturonic acids within 10 min of hydrolysis.

Partial purification and some properties of a T2 ribonuclease from Aspergillus flavipes

A ribonuclease was partially purified from the culture medium of Aspergillus flavipes (IZ:1501), after 96 h of cultivation by chromatography on DEAE-cellulose and Sephadex G100 columns. The molecular weight of the RNase was estimated to be 40 kD by gel filtration using Sephadex G100, and the optimum pH and temperature were 4.0 and 50-55 degrees C, respectively. Catalytic activity was inhibited by Zn+2, Fe+3, Hg+2 and Ag+ ions. The enzyme did not show an exact base specificity and produced four kinds of 3'-nucleotides from yeast RNA.

Purification and biological activity of the thrombin-like substance isolated from Bothrops insularis venom

The venom of Bothrops insidaris snake, known in Brazil as jararaca ilhoa, contains a variety of proteolytic enzymes such as a thrombin-like substance that is responsible for various pharmacological effects. B. insularis venom chromatography profile showed an elution of seven main fractions. The thrombin-like activity was detected in fractions I and 111, the latter being subjected to two other chromatographic procedures, so to say DEAE and Hi Trap Benzamidine. The purity degree of this fraction was confirmed by analytical reverse phase HPLC, which displayed only one main fraction confirmed by SDS-PAGE constituting fraction III. About 5 mu g of fraction III protein potentiated the secretion of insulin induced by 2.8mM of glucose in rats isolated pancreatic beta-cells treated; the increase being around 3-fold higher than its respective control. B. insidaris lectin (BiLec; 10 mu g/mL) was also studied as to its effect on the renal function of isolated perfused rat kidneys with the use of six Wistar rats. BiLec increased perfusion pressure (PP), renal vascular resistence (RVR), urinary flow (UF) and glomerular filtration rate (GFR). Sodium (%TNa+) and chloride tubular reabsorption (%TCl-) decreased at 120 min, without alteration in potassium transport. In conclusion, the thrombin-like substance isolated from B. insularis venom induced an increase in insulin secretion, in vitro, and transiently altered vascular, glomerular and tubular parameters in the isolated rat kidney. (c) 2006 Elsevier Ltd. All rights reserved.

Purification and characterization of beta-fructosidase with inulinase activity from Aspergillus niger-245

Aspergillus niger - 245 a strain isolated from soil samples showed good beta -fructosidase activity when inoculated in medium formulated with dahlia extract tubers. The enzyme was purified by precipitation in ammonium sulphate and percolated in DEAE-Sephadex A-50 and CM-cellulose columns, witch showed a single peack in all the purification steps, maintaining the I/S ratio between 0.32 to, 0.39. Optimum pH for inulinase activity (I) was between 4.0 - 4.5 and for invertase activity (S) between 2.5 and 50. The optimum temperature was 60 degrees .C for both activities and no loss in activity was observed when it was maintained at this temperature for 30 min. The K-m value was 1.44 and 5.0 respectively, for I and S and V-m value 10.48 and 30.55 respectively. The I activity was strongly inhibited by Hg2+ and Ag+ and 2 x 10(-3) M of glucose, but not by fructose at the same concentration. The enzyme showed an exo-action mechanism acting on the inulin of different origins. In assay conditions total hydrolysis of all the frutans was obtained although it has shown larger activity on the chicory inulin than that one from artichoke Jerusalem and dahlia, in the first 30 min. The obtained results suggested that the enzyme presented good potential for industrial application in the preparing the fructose syrups.

Purification and characterization of jararassin-I, a thrombin-like enzyme from Bothrops jararaca snake venom

A thrombin-like serine protease, jararassin-I, was isolated from the venom of Bothrops jararaca. The protein was obtained in high yield and purity by a single chromatographic step using the affinity resin Benzamidine-Sepharose CL-6B. SDS-PAGE and dynamic light scattering analyses indicated that the molecular mass of the enzyme was about 30 kD. The enzyme possessed fibrinogenolytic and coagulant activities. The jararassin-I degraded the Bbeta chain of fibrinogen while the Aalpha chain and gammachain were unchanged. Proteases inhibitors, PMSF and benzamidine inhibited the coagulant activity. These results showed jararassin-I is a serine protease similar to coagulating thrombin-like snake venom proteases, but it specifically cleaves Bbeta chain of bovine fibrinogen. Single crystals of enzyme were obtained (0.2 mmx0.2 mmx0.2 mm) and used for X-ray diffraction experiments.