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L S Gan

Publications and source records attributed to L S Gan.

25 records · Page 2Linked to original sources
PubMed1989

A rapid method for isolation of human hemoglobin benzo[a]pyrene diol epoxide derived adducts using high performance liquid chromatography.

A method is described to isolate rapidly human hemoglobin-benzo[a]pyrene diol epoxide adducts. A combination of 300 A pore size C4 reversed phase HPLC to effect separation of adducted protein from native protein, and mu-bore C18 reversed phase HPLC to isolate and partially characterize proteolytic peptide adducts (by UV), was used.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed1989

Quantitation of methylated hemoglobin via hydrolysis of methyl esters to yield methanol.

Methyl esters formed by methylation of carboxylic acid groups on hemoglobin can be quantitated via GC/MS negative-ion chemical ionization analysis of the pentafluorobenzoate derivative of the methanol released from the hemoglobin upon base hydrolysis. Although the method requires extensive cleanup of extracting solvents because of the ubiquitous presence of trace levels of methanol, it is possible to quantitate adducted hemoglobin at levels as low as 4 ng per gram of hemoglobin using [13C, 2H3]methyl alcohol as the internal standard. In principle this method can be used in humans as a dosimeter for exposure to environmental methylating or alkylating agents.

Animals↗
PubMed1988

Serum albumin adducts in the molecular epidemiology of aflatoxin carcinogenesis: correlation with aflatoxin B1 intake and urinary excretion of aflatoxin M1.

Aflatoxin-serum albumin adducts in the blood of 42 residents of Guangxi Province, People's Republic of China, were determined and compared with intake of aflatoxin B1 (AFB1) and excretion of aflatoxin M1 (AFM1) in urine. Blood specimens were obtained during the same period that urine was collected and that diet was sampled. Serum albumin was isolated from blood by affinity chromatography on Reactive Blue 2-Sepharose and subjected to enzymatic proteolysis using Pronase. Immunoreactive products were purified by immunoaffinity chromatography and quantified by competitive radioimmunoassay. A highly significant correlation (r = 0.60, P less than 0.00003) of adduct level with AFM1 excretion was observed. An equally highly significant correlation of adduct level with intake (r = 0.69, P less than 0.000001) was also observed. From the slope of the regression line for adduct level as a function of intake, it was calculated that 1.4-2.3% of ingested AFB1 becomes covalently bound to serum albumin, a value very similar to that observed when rats are administered AFB1.

Aflatoxin B1↗
PubMed1985

Effects of acetylenic and olefinic pyrenes upon cytochrome P-450 dependent benzo[a]pyrene hydroxylase activity in liver microsomes.

1-Ethynylpyrene, trans-, & cis-1-(2-bromovinyl)pyrene, methyl 1-pyrenyl acetylene, and phenyl 1-pyrenyl acetylene are substrates for cytochrome P-450 dependent monooxygenases and also inhibitors of cytochrome P-450 dependent benzo[a]pyrene hydroxylase activities in liver microsomes from 5,6-benzoflavone or phenobarbital pretreated rats. 1-Ethynylpyrene, trans-1-(2-bromovinyl)pyrene, and methyl 1-pyrenyl acetylene cause a mechanism based inhibition (suicide inhibition) of the benzo[a]pyrene hydroxylase activities in microsomes from 5,6-benzoflavone or phenobarbital pretreated rats, while cis-1-(2-bromovinyl)pyrene only causes suicide inhibition of the hydroxylse activities in the 5,6-benzoflavone induced microsomes and phenyl 1-pyrenyl acetylene does not cause a detectable suicide inhibition of these activities in either type of microsome. Incubation with NADPH and 1-ethynylpyrene, trans-, or cis-1-(2-bromovinyl)pyrene causes a loss of the P-450 content in the microsomes from 5,6-benzoflavone or phenobarbital pretreated rats, but incubations with methyl 1-pyrenyl acetylene or phenyl 1-pyrenyl acetylene did not cause a loss of the P-450 content of either microsomal preparation.

Animals↗
PubMed1984

1-Ethynylpyrene, a suicide inhibitor of cytochrome P-450 dependent benzo[a]pyrene hydroxylase activity in liver microsomes.

The preparation of 1-ethynylpyrene (EP) is described. Incubation of EP with liver microsomes in the presence of NADPH yields fluorescent products, but the production of these products ceases after 15 min. Addition of fresh microsomes restores the original rate of EP metabolism. The metabolism of EP is initially more rapid in microsomes from 5,6-benzoflavone- (BF) pretreated rats than in microsomes from phenobarbital- (PB) pretreated rats or from untreated, control animals. EP strongly inhibits the hydroxylation of benzo[a]pyrene (BP) by liver microsomes; after 20 min in the presence of EP, BP metabolism nearly ceases. Addition of fresh microsomes restores the original rate of BP hydroxylation. EP more effectively inhibits the oxidation of BP in liver microsomes from rats pretreated with BF than from rats pretreated with PB or from untreated, control animals. The inhibition of BP hydroxylation activity due to EP is dependent upon NADPH and is apparently irreversible. Kinetic analyses demonstrate that the observed inhibition of BP hydroxylation is due to loss of the enzymatic activity by a process that is first order in EP and that reaches a limiting value at infinite EP concentrations. One such first-order process, with a t 1/2 of 3.5 min and a Ks for EP of 40 microM, is observed in microsomes from BF-pretreated rats. Two such first-order processes, one with t 1/2 of 6.9 min and Ks of 46 microM and one with t 1/2 of 12.7 min and Ks of 33 microM, are observed in microsomes from PB-pretreated rats. It is proposed that a self-catalyzed inhibition (suicide inhibition) of the cytochrome P-450 dependent BP hydroxylation occurs in the presence of EP. Incubation with EP under conditions that result in loss of about 90% of the BP hydroxylase activity in microsomes from BF-pretreated rats and about 66% of the activity in microsomes from PB-pretreated rats causes the loss of only 6 and 12% of the cytochrome P-450, respectively. It is concluded that loss of P-450 content is an insensitive measure of the effect of this inhibitor upon this cytochrome P-450 dependent enzyme activity. The selectivity of the loss of P-450 due to the incubation of the different microsomal preparations with EP is also observed to be different than the selectivity for loss of BP hydroxylase activity. It is proposed that the suicide inhibition of cytochrome P-450 dependent enzymes by alkynes need not involve heme alkylation and a resulting loss of P-450 content.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMedDate not supplied

Benzo[a]pyrene diol epoxide adduct formation in mouse and human hemoglobin: physicochemical basis for dosimetry.

The interactions of human hemoglobin (hHb) with the anti-diol epoxide of benzo[a]pyrene (aBaPDE) and with the corresponding tetrols formed by hydrolysis of the epoxide were investigated with the aim of characterizing the covalent adducts formed by reaction of the epoxide with the protein. The major product (80% of the total) was determined to be an ester resulting from oxirane ring opening by one or several (unidentified) carboxylate group(s). Minor products were characterized as adducts formed by reaction with amino or heterocyclic nitrogen by comparison of their UV spectra with those of model compounds. There was no evidence for reaction with cysteine. Formation of ester adducts by aBaPDE with mouse hemoglobin (mHb) following administration of BaP to mice was also investigated. It was found that esters constituted the majority of the adducts formed by aBaPDE and a substantial fraction of the total adducts formed. The esters formed by mHb were significantly less stable than those formed by hHb, both in vivo and in vitro. The instability of mHb ester adducts is believed by the responsible for differences among previous descriptions of the in vivo binding of BaP to mHb.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗