• Applied Biological Chemistry
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• v.26 no.2
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• pp.125-131
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• 1983

Formal net charges, bond populations, atomic orbital coefficients, energy components and conformation of dimethyl-2,2-dichlorovinylphosphate have been studied theoretically by using the CNDO/2 molecular orbital calculation method in attempt to describe the reactivity and the stability of the molecule. From the analysis of rate equation, molecular orbital calculations and identification of the hydrolysis products, 2,2-dichloroacetaldehyde and dimethylphosphoric acid, a mechanism of the hydrolysis of dimethyl-2,2-dichlorovinylphosphate(DDVP) has been proposed. The hydrolysis of DDVP proceeds through the mechanism of nucleophilic addition, typical Micheal reaction in basic media. Therefore, it appears probable that the attack by strong nucleophile, hydroxide ion occurs at the increased positive charge $C_2({\alpha})$ atom of a staggered conformation due to the inductive effect (-)I>(+)R of 2,2-dichlorovinyl, electron-attracting group. And then, the hydrolytic scission involves the $C_2({\alpha})-O_3$, ${\pi}-anti-bonding\;orbital({\pi}^*)$ in the subsequent reaction in aqueous solution.

• Development and Reproduction
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• v.4 no.1
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• pp.115-123
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• 2000

A few enzymeimmunoassay (EfA) for testosterone (T) have been reported but was not suitable for all biological samples. The present study was designed to develop a rapid, ultrasensitive EIA and to apply this technique for study the physiological changes of T in biological samples. Saliva samples were collected at 06:00～09:00 hour during one menstrual cycle from 18 normally menstruating women and on 09:00～10:00 hour from 20 normal men. The present study shows an established EIA for testosterone, using horseradish peroxidase (HRP), which was covalently bonded to testosterone-3-carboxymethyloxime (T-3-CMO). One batch of T-antisera was also covalently linked to microcrystalline cellulose particles by a mixed anhydride method in order to facilitate separation of bound and free steroids. The established EIA was validated in terms of sensitivity, accuracy, specificity, precisions etc., comparing with conventional radioimmunoassay. The sensitivity of the established EIA was less than 25 pg/tube. The correlation coefficients between the expected T-values and observed T-values measured by EIA or RIA were r=0.985 and r=0.941 respectively. The cross reactivity of antiserum in EIA was a little higher than that of RIA, especially by 5 ${\alpha}$-DHT. The intra- and inter-assay precisions of the present EIA were similar to those of RIA. The present study also demonstrates that the normal T-values in saliva of Korean male ＆ female samples are 265.65${\pm}$15.80 pmol／l and 109.74${\pm}$ 12.01 pmol／l, respectively. The present EIA seems to be established and suitable for use in the endocrinological studies. The advantages of this EIA system also might make the present T-EIA an ideal procedure for use in a routine assay of ordinary laboratory with a conventional spectrophotometer.