Oxidative stress causes plasma protein modification.

We investigated the effect of oxidative systems on plasma proteins using Chloramine-T, a source of free radicals. Plasma specimens from 10 healthy volunteers were treated with 40 mmol/L Chloramine-T (1:1 v/v). Total protein and plasma carbonyl levels were evaluated spectrophotometrically. Identification of plasma proteins modifications was performed by SDS-PAGE, protein and lipid electrophoresis. Protein fragmentation was evaluated by HPLC. Total protein levels of oxidised plasmas were significantly lower (4.08 ± 0.12 g/dL) than control (7.86 ± 0.03 g/dL) (P <0.01). Plasma carbonyl levels were higher (1.94 ± 0.38 nmol/mg protein) in oxidised plasma than that of control (0.03 ± 0.01 nmol/mg protein) (P<0.01). Plasma oxidation had no significant effect on the levels of proteins and lipids. Protein fragmentations were detected in oxidised groups compared to those of the control. We conclude that protein modifications have direct effect on the protein functions, which are related to stress agent, its treatment period(s), and the methodology used for evaluating such experimental results.

Use of chloramine-T and two dyes in the sensitive determination of stavudine in pharmaceuticals

Three new methods are described for the assay of stavudine (STV) in bulk drug and in dosage forms using chloramine-T (CAT) and two dyes, methyl orange and indigocarmine, as reagents. Titrimetry involves treating STV with a measured excess of CAT in hydrochloric acid medium, and after the oxidation of STV is judged to be complete, the unreacted oxidant is determined iodometrically. Spectrophotometric methods entail the addition of a known excess of CAT to STV in hydrochloric acid medium followed by determination of residual oxidant by reacting with a fixed amount of either methyl orange and measuring the absorbance at 520 nm (Method A) or indigo carmine and measuring the absorbance at 610 nm (Method B). In all the methods, the amount of CAT reacted corresponds to the amount of STV. In titrimetric method, the reaction follows 1:1 stoichiometry (STV: CAT), and is applicable over the range 1.5-10 mg of STV. In spectrophotometric methods, the absorbance is found to increase linearly with concentration of STV. The systems obey Beer's law for 0.2-2.0 and 1.0-10.0 mg/mL for method A and method B, respectively. The apparent molar absorptivities are calculated to be 5.7x10(4) and 1.5x10(4) L/mol/cm for method A and method B, respectively, and the corresponding Sandell sensitivity values are 0.004 and 0.015 µg/cm². The limits of detection and quantification are reported for both methods. Intra-day and inter-day precision and accuracy of the developed methods were evaluated as per the current ICH guidelines. The methods were successfully applied to the assay of STV in tablet and capsule formulations and the results were compared with those of a reference method by applying Student's t-test and F-test. No interference was observed from common tablet adjuvants. The accuracy and reliability of the methods were further ascertained by performing recovery experiments via standard-addition method.

Descrevem-se três novos métodos para o ensaio de estavudina (STV) na matéria-prima e nas formulações utilizando-se clroamina-T (CAT) e dois corantes, alaranjado de metila e índigo carmim como reagentes. A titulação envolve o tratamento de STV com excesso medido de CAT em meio de ácido clorídrico, e, quando a oxidação se completar, o oxidante que não reagiu é determinado iodometricamente. Os métodos espectrofotométricos compreendem a adição de excesso conhecido de CAT ao STV em ácido clorídrico, seguida da determinação do oxidante residual por meio da reação com quantidade fixada de alaranjado de metila, medindo-se a absorvância a 520 nm (Método A) ou índigo carmim, medindo-se a absorvância a 610 nm (Método B). Em todos os métodos, a quantidade de CAT que reagiu corresponde à quantidade de STV. No método titulométrico, a reação segue a estequiometria 1:1 (STV:CAT) e é aplicável na faixa de 1,5 a 10 mg de STV. Nos métodos espectrofotométricos, a absorvância aumenta linearmente com a concentração de STV. Os sistemas obedecem a lei de Beer nos intervalos de 0,2 a 2,0 mg/mL e 1,0 a 10,00 mg/mL para os métodos A e B, respectivamente, e os valores de sensibilidade de Sandell correspondentes são 0,004 e 0,015 µg/cm². Os limites de detecção e de quantificação são apresentados para ambos os métodos. A precisão e a exatidão intra-dia e inter-dia dos métodos desenvolvidos são avaliadas de acordo com as normas ICH. Os métodos foram aplicados com êxito aos ensaios de STV em comprimidos e em cápsulas e os resultados foram comparáveis com aqueles obtidos com o método de referência, utilizando-se o teste t de Student e o teste F. Não se observou interferência dos adjuvantes comuns em comprimidos. A exatidão e a confiabilidade dos métodos foram ajustadas por meio de experimentos de recuperação via método de adição de padrão.

The Effect of Iguratimod on the Collagen Secretion of 3T6 Cells / 医学研究杂志

Objective To study the effect of Iguratimod on the collagen secretion of 3T6 cells. Method The median toxic concen- tration of Iguratimod was determination by MTT Assay. Iguratimod' s median toxic concentration to 3T6 cell was about 1142.6mg/L. Taking into the consideration of the cells performance after administration under spectacles, we chosen the administration concentration as 334.8 mg/L(DMS0:1?l/256?l) ,167.4mg/L,83.7mg/L and 41.85 mg/L. After administration of 12 hours, 24 hours, 36 hours and 48 hours, the supernatant was collected for assayed respectively. The collagens content was detected by the method of chloramine T. Results The collagen concentration of the drug concentration 334.375mg/L group increased (P

An Improved Method for Determination of Cyanide in Drinking Water / 环境与健康杂志

Objective To resolve the problem that qualified chloramine T was difficult to purchase for some basic laboratories Methods Using the solution of available chlorine prepared by bleaching power instead of chloramine T, the method for determination of cyanide in drinking water was improved Results The data obtained by this assay accorded with Beers Law in the range of 0～2 0 ?g/25 ml cyanide The detection limit,relative standard deviation,and recovery rate were 0 04?g,2 69%～3 09% and 97 50%～99 17% respectively Conclusion This method for determination of cyanide in drinking water was simple and easy to operate and presented good precision and accuracy