The L-amino acid oxidase of Malayan pit viper (Calloselasma rhodostoma) venom was purified to electrophoretic homogeneity. The molecular weight of the enzyme was 132,000 as determined by Sephadex G-200 gel filtration chromatography and 66,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It is a glycoprotein, has an isoelectric point of 4.4, and contains 2 mol of flavin mononucleotide per mole of enzyme. The N-terminal amino acid sequence of the enzyme was A-D-D-R-N-P-L-A-E-E-F-Q-E-N-N-Y-E-E-F-L. Kinetic studies suggest the presence of a alkyl side-chain binding site in the enzyme and that the binding site comprises at least four hydrophobic subsites. The characteristics of the binding site differ slightly from those of cobra venom L-amino acid oxidases.
Indirect ELISA shows that the antibodies to Calloselasma rhodostoma venom hemorrhagin (CR-HMG), thrombin-like enzyme (CR-TLE) and L-amino acid oxidase (CR-LAAO) exhibited strong to moderate cross-reactions with most crotalid and viperid venoms, but only anti-CR-LAAO cross-reacted with the elapid venoms. However, the indirect ELISA failed to detect some antigenic similarities demonstrable by cross-neutralization study. The double-sandwich ELISA for the three anti-C. rhodostoma venom components exhibited a much lower level of cross-reactions than the indirect ELISA.
The study was conducted to determine thiocyanate (SCN-) and hypothiocyanite (OSCN-) concentrations in resting (RWS) and stimulated whole saliva (SWS) and stimulated parotid saliva (SPS) of 20 healthy young adults aged 21-29 y. Samples of saliva were collected at 12:30, immediately before lunch. Resting saliva was collected by expectoration, and stimulated saliva was collected during the uniform chewing of paraffin wax. Parotid secretion was collected using a modified Carlsson-Crittenden cup (Carlsson et al., Am, J. Physiol., 26, 169-177, 1910). SCN- concentration was determined by the ferric nitrate method (Betts et al., J. Am. Chem. Soc., 75, 5721-5727, 1953) whilst OSCN- was assayed using 2-mercaptoethanol as a reducing agent (Pruitt et al., Caries Res., 16, 315-323, 1982). In RWS, SWS and SPS, the mean SCN- concentrations (in mM) were 1.48 +/- 0.59(S.D.), 0.90 +/- 0.56(S.D.) and 1.24 +/- 0.65(S.D.) whilst the mean OSCN- concentrations (in microM) were 31.21 +/- 13.54(S.D.), 24.90 +/- 12.61 and 30.19 +/- 23.35(S.D.) in the respective salivas. The presence of OSCN- in the secretion collected from the parotid gland supported previous findings by Tenovuo and Pruitt (Tenovuo et al., J. Oral Path, ol. 13, 573-584, 1984), who suggested an endogenous glandular (eukaryotic) source of hydrogen peroxide (H2O2), since parotid saliva from healthy glands is devoid of bacteria and leukocytes.
By a combination of PCR and direct-cycle sequencing using consensus primers, we analyzed approximately 400-bp fragments within the NS3 genes of twenty-one dengue virus type 3 strains isolated from five neighboring Southeast Asian countries at different time intervals from 1956 to 1992. The majority of base disparities were silent mutations, with few predicted amino acid substitutions, thus emphasizing the strict conservation of the NS3 gene. Phylogenetic trees constructed on the basis of these nucleotide differences revealed distinct but related clusters of strains from the Philippines, Indonesia, and strains from Singapore and Malaysia of the 1970s and early 1980s, while the Thai cluster was relatively more distant. This genetic relationship was compatible with that proposed by other workers who have studied other dengue 3 virus genes such as E, M and prM. However, we observed that the more recent, epidemic-associated dengue 3 strains from Singapore and Malaysia of the late 1980s and early 1990s were more closely related to the Thai cluster, implying their evolution from the latter, and emphasizing the importance of viral spread via increasing travel within the Southeast Asian area and elsewhere. Nucleotide sequence analysis of the NS3 genes of dengue viruses can serve to advance the understanding of the epidemiology and evolution of these viruses.
Fish has been known as a source of nonoccupational mercury exposure to fish-consuming population groups. In this study, hair samples collected from fishermen and their families residing in an industrialized area in Penang and a nonindustrialized area in Terengganu were analyzed for mercury by neutron activation. The range, arithmetic mean, geometric mean, and median of the mercury concentrations for the groups in Penang and in Terengganu were 0.45-16.68, 3.61, 3.49, and 2.96 and 6.79-18.31, 12.08, 11.69, and 12.05 mg/kg, respectively. Somewhat lower values than from the Penang group were found in a group from Selangor consisting mainly of office workers. The group in Penang took about 40-100 g of fish/d, whereas the group in Terengganu consumed twice as much. This shows that hair mercury levels depend on a fish consumption pattern, and not on the location of the population. The levels of mercury found in this study were similar to those reported by other workers for fish-consuming population groups worldwide.
Trace elements, such as As, Co, Cr, Hg, Sb, and Zn, were determined by neutron activation analysis (NAA), whereas Cd, Cu, and Pb were determined by graphite furnace atomic absorption spectroscopy (GFAAS) in clam, crab, prawn, swamp cerith, and mussel samples after digestion by microwave heating under controlled conditions before eluting the solutions through a column of a chelating resin, Chelex-100. The standard used in the determination of percentage volatile elements retained by microwave digestion and also in the activation process was Lobster Hepatopancreas TORT-1, whereas known mixed standards were prepared from nitrate salts to determine the efficiency of the separation procedure at a controlled pH. Mercury and lead detected in crabs exceeded the maximum permissible level. Some species also showed a high affinity toward certain elements, and their levels of accumulation in the tissues of these species corresponded with the concentration of these elements in sediments, especially at sites in the vicinity of an industrial zone.
In this study, the depth of cure of composite resins cured within simulated root canals by means of light-transmitting plastic posts was compared to that achieved by the conventional light-curing method. Six sizes of posts with diameters of 1.05 mm, 1.20 mm, 1.35 mm, 1.50 mm, 1.65 mm, and 1.80 mm were investigated. In general, the larger the post diameter, the greater was the depth of cure. There were significant differences in the depth of cure between the control and all sizes of posts investigated. There were also significant differences between the various post diameters except for the 1.35 mm and 1.50 mm diameter posts. It was possible to achieve a depth of cure exceeding 11 mm using these light-transmitting posts.
Heart mitochondria freshly isolated from ginseng treated rats respired higher at ADP-induced, state 3 respiratory rates and with greater respiratory indices. These mitochondria were less susceptible to experimentally-induced functional impairment. Control heart mitochondria incubated with ginseng extract also showed that ginseng prevented mitochondria from incubation induced deterioration with NAD-linked substrates. Comparison of force of contraction of isolated, perfused and electrically paced hearts showed that deterioration of the force of heart contraction was consistently smaller throughout the experiment in hearts from ginseng treated rats. These results indicated that Panax ginseng was able to delay experimentally induced heart mitochondrial impairment and muscle contraction deterioration.
Aspergillus flavus produced approximately 50 U/mL of amylolytic activity when grown in liquid medium with raw low-grade tapioca starch as substrate. Electrophoretic analysis of the culture filtrate showed the presence of only one amylolytic enzyme, identified as an alpha-amylase as evidenced by (i) rapid loss of color in iodine-stained starch and (ii) production of a mixture of glucose, maltose, maltotriose and maltotetraose as starch digestion products. The enzyme was purified by ammonium sulfate precipitation and ion-exchange chromatography and was found to be homogeneous on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified enzyme had a molar mass of 52.5 +/- 2.5 kDa with an isoelectric point at pH 3.5. The enzyme was found to have maximum activity at pH 6.0 and was stable in a pH range from 5.0 to 8.5. The optimum temperature for the enzyme was 55 degrees C and it was stable for 1 h up to 50 degrees C. The Km and V for gelatinized tapioca starch were 0.5 g/L and 108.67 mumol reducing sugars per mg protein per min, respectively.
The hydroxide ion-catalyzed hydrolysis of securinine involves the ring opening of the lactone moiety. The rate of hydrolysis is insensitive to the ionic strength. The observed pseudo-first-order rate constants reveal a decrease of approximately 4-fold due to the increase in the MeCN content from 4 to 50% (v/v) in mixed aqueous solvent. The temperature dependence of the rate of hydrolysis follows the Eyring equation, which yields delta H* and delta S* as 11.0 kcal mol-1 and -34.5 cal deg-1 mol-1, respectively. The hydroxyl carboxylate product of the alkaline hydrolysis of securinine is shown to undergo cyclization in acidic medium to yield securinine. The observed pseudo-first-order rate constants for cyclization increase linearly with an increase in [H+]. The change in the content of MeCN from 3.8 to 47.2% (v/v) in mixed aqueous solvents does not show an effect on the rate of the cyclization reaction. The most plausible mechanisms for alkaline hydrolysis and acid cyclization reactions are also discussed.
Matched MeSH terms: Alkaloids/chemistry*; Central Nervous System Stimulants/chemistry*
The effects of ovariectomy and hormone replacement in control and carcinogen treated female rats were investigated by measuring whole blood and liver glutathione (WGSH, HGSH), glutathione S-transferase (GST), glutathione peroxidase (GPx), and glutathione reductase (GRx) and histological evaluation. Hepatocarcinogenesis was induced by diethylnitrosamine and 2-acetylaminofluorene. In control rats not receiving carcinogen, ovariectomy significantly increased the GST and GRx activities. Replacement with either estrogen or progesterone reduced the GST activities to below intact female values whereas replacement of both hormones together brought the GST activities to that of intact females. GRx activities were brought to intact female values by replacement with estrogen or progesterone, either singly or in combination. Neither ovariectomy nor sex hormone/s replacement influenced the levels of WGSH, HGSH and GPx activities. Carcinogen administration to intact rats increased all the parameters measured. Ovariectomized rats treated with carcinogen showed lower GPx and GRx activities at 2 mths. However, replacement with either progesterone or combined estrogen and progesterone increased GPx and GRx activities to original values. On the other hand GST and GPx activities in ovariectomized rats which had carcinogen treatment were lower than intact rats after 5 mths. Replacement with hormones either singly or both brought GST and GPx activities up to intact rat levels receiving carcinogen. The levels of WGSH, HGSH and GRx activities (5 mths) in carcinogen treated rats were not influenced by ovariectomy and/or hormone/s replacement. The results from this study suggested that ovariectomy reduced the severity of hepatocarcinogenesis which was restored by sex hormone/s replacement.
Ethanolic extract of Cassia alata leaves was investigated for its antimicrobial activities on several microorganisms including bacteria, yeast, dermatophytic fungi and non-dermatophytic fungi. In vitro, the extract exhibited high activity against various species of dermatophytic fungi but low activity against non-dermatophytic fungi. However, bacterial and yeast species showed resistance against in vitro treatment with the extract. The minimum inhibitory concentration (MIC) values of the extract revealed that Trichophyton mentagorphytes var. interdigitale, Trichophyton mentagrophytes var. mentagorophytes, Trichophyton rubrum and Microsporum gypseum had the MIC of 125 mg/ml, whereas Microsporum canis had the MIC of 62.5 mg/ml. The inhibition can be observed on the macroconidia of Microsporum gypseum which resulted in structural degeneration beyond repair. The mechanism of inhibition can be related to the cell leakage as observed by irregular, wrinkle shape and loss in rigidity of the macroconidia.
A lamellar liquid crystalline region was identified in a typical skin lotion formulation system composed of a mixture of isostearic acid and triethanolamine (TEA) at 65:35 (w/w), decane, and water (the temperature was controlled at 30 degrees C). The interlayer spacings were determined by a small-angle X-ray diffraction technique. Incorporation of a natural dye, curcumin, resulted in lower interlayer spacings and higher penetration of water into the layered structure. However, the higher penetration of water was not apparent at all compositions of isostearic acid:TEA, decane, and water.
Many proteins derived from the latex of Hevea brasiliensis that remain soluble in trichloroacetic acid (TCA) can be precipitated by phosphotungstic acid (PTA). A combination of 5% TCA and 0.2% PTA precipitates a wide range of proteins effectively even when they are present in low concentrations (below 1 microgram ml-1). In addition to its protein purification function, acid precipitation also increases the sensitivity of the subsequent protein assay by allowing the test sample to be concentrated. Another advantage of protein precipitation by TCA and PTA is that very small amounts of protein (of the order of 10 micrograms) can be repeatably recovered without the use of precipitate-bulking agents such as sodium deoxycholate. This general procedure of protein purification and concentration is simple and rapid, but the use of PTA may not be fully compatible with the Bradford protein assay. A modified Lowry microassay is described which enables about 3 micrograms ml-1 to be quantitated at the photometric absorbance of 0.05. When used in conjunction with protein concentration by precipitating with TCA/PTA, approximately 0.4 microgram ml-1 protein present in 6 ml of solution can be assayed.
A novel derivative of sucrose, beta-(3,6-di-O-feruloyl)-fructofuranosyl-alpha-(2,3,4,6-tetra-O-ac etyl)- glucopyranoside, was isolated from the wood of Bhesa paniculata. Its structure was determined by a combination of 2D 1H-1H and 1H-13C correlation NMR spectroscopy. The known compounds, glycerol 1-9',12'-octadecadienoate, beta-sitosterol, (+/-)-pinoresinol, methyl 3,4-dihydroxybenzoate, 4-hydroxy-3-methoxybenzoic acid, anofinic acid and 2-(1'-methylethenyl)-benzofuran-5-carboxylic acid were also isolated.