James R. Heitz

Separation and characterization of venom components in Loxosceles reclusa—III. Hydrolytic enzyme activity☆

Abstract Fractions 3, 4, 5, 6, 8 and 9 of the prep-disc electrophoretic separation of Loxosceles reclusa spider venom showed enzymic activity toward the hydrolysis of indoxyl-acetate and were therefore considered as hydrolytic enzyme components in the spider venom. Fraction 5 was designated as a lipase due to its higher affinity for the longer chain synthetic substrates. Fraction 6 was designated as a nonspecific esterase due to its action on alkyl and phosphate esters. Fraction 9 was designated as the primary alkaline phosphatase activity in the venom. Electrophoretic assays revealed two venom protein bands exhibiting alkaline phosphatase and three venom proteins exhibiting esterase activity. Two of the bands were arylesterases and one was an aliesterase; acetylcholinesterase was not detected.

Characteristics of an alkaline phosphatase activity in brown recluse venom.

Abstract The venom of the brown recluse spider (Loxosceles reclusa) has been shown by fluorometric assay to contain an alkaline phosphatase activity. Similar Km values were determined for four substrates, differing widely as to size, suggesting a broad specificity. Kinetic inactivation by fluorescein mercuric acetate and p-hydroxymercuribenzoate suggests that this is a sulfhydryl enzyme. The importance of a disulfide bond to the tertiary structure of the enzyme is indicated by the complete loss of activity upon addition of mercaptoethanol.

Physiological effects of crude oil exposure in the striped mullet, Mugil cephalus

Abstract Juvenile mullet ( Mugil cephalus ) were exposed to a surface slick of Empire Mix crude oil for a three week period in a simulated estuarine ecosystem. Liver weight to body weight ratios were increased in the mullet from the oil-treated ponds when compared to those from the control ponds. Activities of alkaline phosphatase, which were elevated in gill and muscle of oil-treated mullet, and β-glucuronidase, which was elevated in the muscle of oil-treated mullet, may be related to the degree of stress the animals were experiencing. Malic dehydrogenase, which was depressed in the livers and elevated in the muscle of oil-treated organisms, indicate changes in aerobic metabolism in response to the stress of crude oil exposure. Muscle acetylcholinesterase was not affected by oil exposure.

Pesticidal applications of halogenated xanthene dyes

James Robert Heitz, born 1941. Prof. of Biochemistry and Molecular Biology, Mississippi State University, Mississippi State, MS. President, PhotoDye International, Inc. A.B. in Chemistry (1963), Bellarmine College; Ph.D. in Biochemistry (1967), U. of Tennessee (Knoxville). 1968-70, Postdoctoral at the McCollum-Pratt Institute, The Johns Hopkins University, 1970-74, Asst. Prof.; 1974-79, Assoc. Prof.; 1979-present, Prof., all at MSU. 1974-present, joint appointment, Dept. of Chemistry; 1975-present, joint appointment, Dept. of Entomology, both at MSU; 1987-89, Manager, Center for Agricultural Product Research and Development; 1988-94, Head, Analytical Support and Food Safety Laboratory, both at MSU. Main field of interest: The effects of dyes in agricultural systems. Fellow, Division of Agrochemicals, American Chemical Society.

Enzyme activities following chronic exposure to crude oil in a simulated ecosystem: I. American oysters and brown shrimp

Abstract Enzyme activities were investigated in whole-body homogenates from oysters and hepatopancreas homogenates from shrimp which had been exposed to crude oil for 8 months in a simulated estuarine ecosystem. Enzymes assayed included acetylcholinesterase, alkaline phosphatase, β-glucuronidase, glutamic-pyruvic transaminase, lactic dehydrogenase, and malic dehydrogenase. Few seasonal trends in enzyme activity were observed in either species. Several alterations in enzyme activity were noted in oil-treated shrimp and oysters 6–8 months following the oil spill when the animals were stressed and may reflect physiological changes in animals which are severely stressed. However, few chronic alterations in enzyme activity were observed which could be attributed to the oil spill.

Variation in enzyme activities of the American oyster (Crassostrea virginica) relative to size and season

Abstract 1. 1. The specific activities and subcellular distribution of eleven enzymes were determined in whole body homogenates of the American oyster, Crassostrea virginica . 2. 2. The activities of lactic dehydrogenase, cytochrome oxidase, and acid phosphatase were inversely proportional to organism weight. 3. 3. Seasonal fluctuations were observed for malic dehydrogenase, glutamic oxaloacetic transaminase, glutamic pyruvic transaminase, acetylcholinesterase, alkaline phosphatase and acid phosphatase.

Enzyme activities following chronic exposure to crude oil in a simulated ecosystem. II. Striped mullet.

Abstract Enzyme activities were investigated in brain, gill, liver, and muscle homogenates from striped mullet which had been exposed to crude oil for 10 months in a simulated estuarine ecosystem. Enzymes assayed included acetylcholinesterase, alkaline phosphatase, β-glucuronidase, glutamic-pyruvic transaminase, lactic dehydrogenase, and malic dehydrogenase. Few seasonal trends in enzyme activities were observed. Alterations in some enzyme activities, particularly acetylcholinesterase, β-glucuronidase, and malic dehydrogenase, may have reflected physiological changes in the mullet resulting from stress. In general, there were few chronic alterations in mullet enzyme activities resulting from the oil spill.

Dye-sensitized photoinactivation of the lactic dehydrogenase and acetylcholinesterase from the boll weevil, Anthonomous grandis

Abstract The lactic dehydrogenase and acetylcholinesterase enzymes of the boll weevil, Anthonomous grandis , have been shown to be inactivated by dye-sensitized photooxidation mediated by substituted xanthenes. The efficiency of the photooxidation reaction was correlated with the degree of halogenation of the dye molecule, the efficiency of the dye in singlet oxygen formation, and the strength of binding to lactic dehydrogenase. Changes in the in vivo levels of these enzymes due to ingestion of rose bengal by adult weevils are not further modified in the presence of light.

High-performance liquid chromatographic determination of ecdysteroid titers in the house fly

Abstract An efficient, rapid and sensitive method has been developed for extraction, clean-up, and analysis of ecdysteroid titers in the larval stage of the house fly. A methanolic extract was filtered, defatted with hexane, and then passed through a C18 reversed-phase Sep-Pak cartridge, which retained the ecdysteroids. Polar interfering materials were removed by eluting the cartridge with water, followed by 5% acetonitrile and 20% methanol. The methanol eluate was used for quantitative analysis of ecdysteroids by high-performance liquid chromatography on a C18 μBondapak reversed-phase column with 16% aqueous acetonitrile. The detection limits for 20-hydroxyecdysone and ecdysone were 10 and 20 ng, respectively.

The Purification of Xanthene Dyes by Reverse Phase High Performance Liquid Chromatography

Abstract A reliable method for the separation of fluorescein dyes from their impurities was developed using high performance liquid chromatography and involved a μBondapak C18 reverse phase column and mixtures of methanol and ammonium acetate buffer. This technique was used to verify the purity of commercial products as well as to aid in the development of an empirical theory related to retention of halogenated fluorescein dyes by reverse phase columns.