Reducing sugars can induce the oxidative inactivation of rhodanese.

The enzyme rhodanese (thiosulfate sulfurtransferase, EC 2.8.1.1) is inactivated on incubation with reducing sugars such as glucose, mannose, or fructose, but is stable with non-reducing sugars or related polyhydroxy compounds. The enzyme is inactivated with (ES) or without (E) the transferable sulfur atom, although E is considerably more sensitive, and inactivation is accentuated by cyanide. Inactivation of E is accompanied by increased proteolytic susceptibility, a decreased sulfhydryl titer, a red-shift and quenching of the protein fluorescence, and the appearance of hydrophobic surfaces. Superoxide dismutase and/or catalase protect rhodanese. Inactive enzyme can be partially reactivated during assay and almost completely reactivated by incubation with thiosulfate, lauryl maltoside, and 2-mercaptoethanol. These results are similar to those observed when rhodanese is inactivated by hydrogen peroxide. These observations, as well as the cyanide-dependent, oxidative inactivation by phenylglyoxal, are explained by invoking the formation of reactive oxygen species such as superoxide or hydrogen peroxide from autooxidation of alpha-hydroxy carbonyl compounds, which can be facilitated by cyanide.     

Recovery of parasitic nematodes from fish by digestion or elution.

Two methods, digestion and elution, were used to recover parasitic nematodes from 470 flatfish belonging to species in the family Pleuronectidae. Samples of similar fish were collected from market lots; half of each sample was subjected to digestion, and half was subjected to elution (sedimentation). The edible (flesh) and the inedible (viscera) portions of each fish were analyzed separately. The total number of nematodes recovered by digestion was 1,110, which was not significantly greater than the 922 nematodes recovered by elution. However, digestion recovered 1,062 nematodes of the anisakine genera Anisakis and Phocanema, which are potentially pathogenic for human consumers of raw of semiraw fish. This number is significantly greater than the 608 pathogenic nematodes recovered by elution. Digestion also recovered 242 more nematodes from the edible flesh than did elution. Conversely, more nonpathogenic nematodes were recovered by elution. Approximately half the fish (240) had been collected in Boston markets, and the other half (230) had been collected in San Francisco markets. Fish from San Francisco each contained an average of eight nematodes, and those from Boston contained an average of less than one nematode per fish.     

Factors in the rat submaxillary gland that stimulate growth of cultured glioma cells: identification and partial characterization

The effect of rat submaxillary extract on the growth of rat C6 glioma cells in serum-free culture has been examined. Extracts (10-15 microgram/ml) of submaxillary glands from both male and female rats markedly enhanced the growth of serum-deprived C6 cells and, in combination with insulin, transferrin, and NIH-LH (a source of fibroblast growth factor), were able to stimulate C6 cell growth to an extent comparable to that achieved with an optimal amount of fetal calf serum. The mitogenic activity of rat submaxillary extracts was found to be heat-labile, acid-stable, and partially inactivated by protease and 2-mercaptoethanol. Under our assay conditions, biologically active preparations of purified mouse submaxillary gland epidermal growth factor (EGF) or nerve growth factor (NGF) were not mitogenic for C6 cells, nor was the mitogenic activity of rat submaxillary extracts inhibited by antiserum to these mouse submaxillary gland growth factors. These results suggest that the active component(s) of rat submaxillary extracts is unrelated to either EGF or NGF. The growth-enhancing effect also appears unrelated to esteropeptidase activity present in these extracts since the mitogenic activity was unaffected by several protease inhibitors. Moreover, two purified mouse submaxillary gland arginylesteropeptidases, EGF-binding protein and gamma-subunit of 7 S NGF, were unable to elicit a comparable growth response even when added to cell culture medium at unreasonably high concentrations. The C6 cell mitogenic activity of crude submaxillary extracts could be separated into two biologically similar components by either gel filtration on Sephadex G-100, preparative isoelectric focusing in a pH gradient of 3-10, or adsorption to DEAE-cellulose followed by elution with a sodium chloride gradient. One of the active components was acidic in nature and had an apparent molecular weight of 40,000, while the other was near neutral in charge and possessed a molecular weight of approximately 20,000. The relationship between these two C6 cell mitogenic components and the rat submaxillary gland component responsible for stimulating Balb/c-3T3 cell growth in serum-free, factor supplemented medium (McClure et al., 1979, J. Cell Biol. 83:96a) is also discussed.     

Usefulness of the stomacher in a microbiological regulatory laboratory.

The relative efficiency of the Waring blender, the Stomacher 400, and the Stomacher 3500 for preparing food samples for microbiological analysis was studied. Comparative aerobic plate count (APC) values were determined on 671 samples, representing 30 categories of foods. Of the 26 categories of nonfatty foods, the blender gave significantly higher geometric mean APC values than those given by the Stomacher 400 and the Stomacher 3500 in 65 and 69 percent of the categories, respectively. In a comparison of the two Stomacher models, the Stomacher 400 gave significantly higher geometric mean APC values than these given by the Stomacher 3500 in 73 percent of the food categories. Addition of Tween 80 to four categories of fatty foods at concentrations of 0.5, 1.0, and 2.0 percent did not raise the APC values given by either model of stomacher to the levels given by the Waring blender. Overall, the efficiency of both models of Stomacher, relative to the blender and to each other, was specific and depended upon the particular food being analyzed.     

p-Chlorphenylalanine effect on phenylalanine hydroxylase in hepatoma cells in culture.

We have investigated the p-chlorophenylalanine-dependent loss of phenylalanine hydroxylase activity in cultured hepatoma cells. The similarity of the effect of p-chlorophenylalanine on phenylalanine hydroxylase in the hepatoma cells and that reported from studies in vivo indicates that the loss of phenylalanine hydroxylase activity is due to a direct interaction of the amino acid analogue with the liver. We can find no evidence that the loss of phenylalanine hydroxylase activity is due to: a direct inactivation of the hydroxylase by p-chlorophenylalanine or an inhibitor produced by p-chlorophenylalanine treatment; an effect similar to that of p-fluorophenylalanine; or leakage of enzyme from the cells during p-chlorophenylalanine treatment. The data presented indicate: (a) the p-chlorophenylalanine effect is rather specific for phenylalanine hydroxylase; (b) following p-chlorophenylalanine removal, new protein synthesis is necessary for restoration of the hydroxylase activity; (c) the rate of loss of phenylalanine hydroxylase activity after the addition of p-chlorophenylalanine is much faster than the rate of restoration of the hydroxylase activity after removal of p-chlorophenylalanine; (d) even in the presence of p-chlorophenylalanine, hydrocortisone greatly stimulates the hydroxylase activity; (e) the cell density-dependent increase of phenylalanine hydroxylase activity is blocked by p-chlorophenylalanine. A discussion of the possible mechanisms of p-chlorophenylalanine-dependent loss of phenylalanine hydroxylase is presented. To measure very low leanine-dependent loss of phenylalanine hydroxylase is presented. To measure very low levels of phenylalanine hydroxylase activity, a new procedure, based on isotope dilution, was developed for isolating the tyrosine formed during the enzymatic reaction.     

CHLOROGENIC ACID ACCUMULATION AND WOUND HEALING IN SWEET POTATO ROOTS

Mc Clure , T. T. (Plant Pest Control Division, ARS, USDA, Washington, D. C.) Chlorogenic acid accumulation and wound healing in sweet potato roots . Amer. Jour. Bot. 47(4) : 277—280. Illus. 1960.–Chlorogenic acid accumulation in cells adjacent to a wound occurs before suberization and wound-periderm formation. Suberization during wound healing was highly correlated with chlorogenic acid accumulation and with wound-periderm formation. The possible role of chlorogenic acid as a source of chemical units for suberization is suggested. Histochemical tests indicate that suberization during wound healing may be a form of lignification. Over 5 times as much lignin was found by chemical analysis in the tissues of healed surfaces as in controls.     

Computerized Interpretation of the Electron Ionization Mass Spectra of Nucleoside TMS Derivatives

Abstract

A computer program has been developed for the automated interpretation of mass spectra of TMS derivatives of nucleosides found in human urine. The m/z values in the unknown spectrum are compared to m/z values of 3 different ion series commonly observed in the mass spectra of nucleoside TMS derivatives.¹ If a correlation exists, the unknown spectra are marked with color according to the scheme: 1) blue—molecular ion series, 2) red—base ion series and 3) yellow—sugar ion series. The program suggests a structural assignment for each of the marked ions and calculates a series related ion current. The calculated ion current is used to assign the of sugar contained in the unknown nucleoside.