ABSTRACT
【Objective】 To develop and verify high performance liquid chromatography (HPLC) detection for tromethamine (Tris) residues in human coagulation factor Ⅷ. 【Methods】 Alanine was used as internal standard, and AQC for pre-column derivation. Inertsil® ODS-SP was adopted, and acetic acid-sodium acetate buffer and acetonitrile were used for gradient elution at the flow rate of 1 mL/min and column temperature of 37℃. The 25 μL sample was loaded and determined by UV-detector with detection wavelength at 248 nm. This method was then verified. 【Results】 Glycine, sodium citrate and calcium chloride showed no interference with the detection of tromethamine (Tris) residues. The recovery rate of spike samples was within 90.0%~100.8%. The RSD in repeatability test were 2.5%, 0.7% and 1.2%, respectively, and in intermediate precision test 0.8%. The tromethamine (Tris) at concentrates of (0.5~4.0)μg/mL showed good linear relationship to the peak area to internal standard, of which the regression equation was Y=0.012 9X-0.000 782, R=0.999 97.The quantitative detection limit was 0.04μg/mL. 【Conclusion】 The HPLC for determination of tromethamine (Tris) residues in human coagulation factor Ⅷ was successfully developed, which showed good linearity as well as high specificity, precision and accuracy.
ABSTRACT
The effect of CdS quantum dots (QDs) on the electrochemiluminescence (ECL) signal of Ru(bpy)32+ was studied. It was found that CdS QDs could enhance the anodic ECL of Ru(bpy)32+ by 4 times. The sensitization mechanism was discussed and the influence factors including concentrations of Ru (bpy)32+ and CdS QDs, pH of solution and scan rate on ECL intensity were investigated. On the basis of quenching effect of catechol on the ECL signal of CdS QDs-Ru(bpy)32+,a system for sensitive determination of catechol was established with a detection limit of 5.5 nmol/L (S/N=3). This method was applied to the detection of catechol in tea sample with satisfactory results.
ABSTRACT
O presente estudo teve como objetivo avaliar a capacidade crioprotetora das lipoproteínas de baixa densidade (LDL) presentes no plasma de gema de ovo, adicionado ao trihidroxiaminometano (TRIS) para congelar sêmen ovino. Trinta e seis ejaculados foram coletados para formar 12 "pool". Cada alíquota de sêmen foi diluída em TRIS-gema de ovo (TRISG) ou TRIS- plasma de gema de ovo (TRISP) antes de congelar o sêmen. Para a obtenção do plasma da gema de ovo, foi utilizado o método de ultracentrifugação. Após o descongelamento, não houve diferença entre os dois extensores em relação aos parâmetros seminais (motilidade, viabilidade, membrana acrossômica e plasma). No entanto, no Teste de Termo Resistência Lenta (TTRL - 4h/38°C), o sêmen congelado com TRISP resultou no aumento do número de espermatozoides com acrossoma intacto (P<0,032). Conclui-se que o diluente TRISP é uma alternativa para criopreservação do sêmen de carneiros, pois permite uma melhora da característica seminal em relação ao diluente TRIS gema, mas sem a necessidade do processo de purificação das LDL.
The present study aimed to evaluate the cryoprotectant low-density lipoprotein (LDL) present in the plasma of egg yolk added to the extender trihidroxiaminometano (TRIS) for freezing ram semen. Thirty-six ejaculates were collected to form 12 pool. Each aliquot of semen was diluted in TRIS-egg yolk (TRISG) or TRIS-egg yolk plasma (TRISP) before freezing the semen. The plasma of egg yolk was obtained by ultracentrifugation. After thawing, no difference was detected between the two extenders in relation to seminal parameters (motility, viability, plasma membrane and acrosome). However, in the thermal resistance slow test (4h in a water bath at 38°C), the semen frozen with TRISP resulted in higher number of sperm with intact acrosome than those with TRISG (P<0.032). It was concluded that the TRISP extender is an alternative for cryopreservation of ram semen, once it improves the semen characteristics in comparison to TRISG, avoiding the purification process of LDL.
ABSTRACT
The hydrolysis of p-nitrophenyl phosphate (pNPP) by calf intestinal alkaline phosphatase (CIAP) was investigated with respect to kinetic parameters such as Vmax, Km and Kcat under varying pH, buffers, substrate concentration, temperature and period of incubation. Highest activity was obtained with Tris-HCl at pH 11, while in the case of glycine-NaOH buffer the peak activity was recorded at pH 9.5. The enzyme showed the following kinetic characteristics with pNPP in 50 mM Tris-HCl at pH 11 and 100 mM glycine-NaOH at pH 9.5 at an incubation temperature of 37°C: Vmax, 3.12 and 1.6 µmoles min-1 unit-1; Km, 7.6 × 10-4 M and 4 × 10-4 M; and Kcat, 82.98 s-1 and 42.55 s-1, respectively. CIAP displayed a high temperature optimum of 45°C at pH 11. The kinetic behaviour of the enzyme under different parameters suggested that the enzyme might undergo subtle conformational changes in response to the buffers displaying unique characteristics. Bioprecipitation of Cu2+ from 50 ppm of CuCl2 solution was studied where 64.3% of precipitation was obtained. Pi generated from CIAP-mediated hydrolysis of pNPP was found to bind with copper and precipitated as copper-phosphate. Thus, CIAP could be used as a test candidate in bioremediation of heavy metals from industrial wastes through generation of metal-phosphate complexes.
Subject(s)
Alkaline Phosphatase/chemistry , Alkaline Phosphatase/metabolism , Animals , Cattle/metabolism , Enzyme Activation , Enzyme Stability , Hydrolysis , Kinetics , Nitrophenols/chemistry , Organophosphorus Compounds/chemistryABSTRACT
The enantioseparations of 38 racemates on Chiralcel OD, Chiralpak AD, Chiralpak IA and(S, S)-Whelk-01 were presented by HPLC. Those enantiomers come from the amines, alcohols, ethers, ketones, aromatic derivatives, heterocyclic compounds, amide acids and medicines etc. With the mobile phase of n-hexane)/isopropanol(90∶ 10, V/V), n-hexane/isopropanol/trifluoracetic acid(90∶ 10∶ 0.2, V/V) or n-hexane/isopropanol/triethylamine(90∶ 10∶ 0.2, V/V), over 70% enantioseparations were obtained for OD, AD and IA columns). The order of enantioseparation selectivity for four columns was OD>AD>IA>(S,S)-Whelk-01, and among those columns there was a big chiral discriminating complementarity. This investigation was useful for choosing chiral columns to separate chiral compounds.
ABSTRACT
Among various ECL systems,such as 9,10-diphenylanthraeene,lucigenin,tris(2,2'-bipyridyl) ruthenium,peroxyoxalate,luminol,graphene,and nanocrystals,Ru(bpy)_3~(2+) ECL is one of the most widely studied ECL systems in recent years due to its broad applications in immunoassays,DNA probe assays,core-actants analysis,and aptasensors. In this review,the progress in Ru(bpy)_3~(2+) ECL has been summarized on the whole,and the future research trends have been proposed.
ABSTRACT
An electrogenerated chemiluminescence ( ECL) sensor was fabricated by employing ruthenium complex polymer as ECL signal and ionic liquid as binding reagent and a sensitive ECL method for the determination of heroin was developed. The ECL sensor was prepared by thoroughly mixing ruthenium complex polymer,graphite powder and ionic liquid,which shows good electrochemical and ECL behaviors. The detection limit for tripropylamine (TPA) was one order of magnitude lower than that observed at the paraffin modified carbon paste electrode. ECL intensity of the fabricated sensor was greatly enhanced in the presence of heroin and a sensitive ECL method was proposed for the determination of heroin based on the enhancement ECL. The ECL intensity was linear with the concentration of heroin in the range from 2. 0 ×10~(-9) mol/L to 2. 0×10~(-5) mol/L and the detection limit was 8×10~(-10) mol/L. The ECL sensor exhibited a long-term stability,fine reproducibility with relative standard deviation less than 5% for 5. 0×10~(-9) mol/L heroin in 60 continuous determinations. The developed method allows the detection of heroin in a serum sample with recovery in 94% -101%.
ABSTRACT
BACKGROUND: Plasmid-mediated AmpC beta-lactamases (PABLs) have been detected in the strains of Escherichia coli, Klebsiella spp., Proteus mirabilis, and Salmonella spp. PABLs may be difficult to detect and might interfere in the therapeutic and infection-control processes. Although several PABL-detection methods based on phenotypes have been reported, the Clinical and Laboratory Standards Institute currently does not recommend a routine detection method for PABLs. The aim of this study is to compare the performances of 3 phenotypic PABL detection methods. METHODS: Total 276 non-duplicated clinical isolates of E. coli (N=97), K. pneumoniae (N=136), and P. mirabilis (N=43) were collected from 14 hospitals in Korea between April and June 2007 in a non-consecutive and non-random manner. Multiplex PCR was performed to detect the PABL genes. Further, 3 phenotypic detection methods-cephamycin-Hodge test, Tris-EDTA (TE) disk test, and combination-disk test with 3-aminophenylboronic acid (BA)-were performed using cefoxitin and cefotetan disks. RESULTS: PABL genes were detected by multiplex PCR in 122/276 isolates, including 14/97 E. coli, 105/136 K. pneumoniae, and 3/43 P. mirabilis isolates. The combination-disk test with BA showed higher sensitivity (98.4%), specificity (92.2%), and efficiency (96.3%) than the cephamycin-Hodge (76.2%, 96.1%, and 88.6%, respectively) and the TE-disk (80.3%, 91.6%, and 87.9%, respectively) tests. CONCLUSIONS: The combination-disk test with BA is a simple, efficient, and interpretable test that can be applicable in clinical laboratories involved in the detection of PABLs in clinical isolates of E. coli, K. pneumoniae, and P. mirabilis.
Subject(s)
Humans , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/analysis , Cefotetan/pharmacology , Cefoxitin/pharmacology , Disk Diffusion Antimicrobial Tests/methods , Escherichia coli/genetics , Klebsiella pneumoniae/genetics , Phenotype , Plasmids , Proteus mirabilis/genetics , Sensitivity and Specificity , beta-Lactamases/analysisABSTRACT
BACKGROUND: Epoxy resin compounds are one of the common causes of occupational allergic contact dermatitis. In Korea, most cases of allergic contact dermatitis from epoxy resin compounds have been caused by the epoxy resin itself. We report a rare case of allergic contact dermatitis which was caused by epichlorohydrin, an ingredient of epoxy resin and 2,4,6-tris-(dimethylaminomethyl)phenol (tris-DMP), a kind of hardeners. CASE REPORT: A 43-year-old man, who had worked at the epoxy resin glue manufacturing factory since 1999, presented with mild and intermittent erythematous papules and rashes on his face, neck, trunk, and both arms. He was dealing with epoxy resin, epichlorohydrin, bisphenol A and hardeners. After a new hardener was added in August 2008, his skin lesions worsened from what he had experienced in the past. A skin patch test was performed to identify the causative chemicals of the skin lesion. Epichlorohydrin and tris-DMP elicited positive reactions after 48 hours and increased after 96 hours. CONCLUSION: This case confirmed occupational allergic contact dermatitis caused by epichlorohydrin and tris-DMP, an ingredient of epoxy resin and a hardener, respectively.
Subject(s)
Adult , Humans , Adhesives , Arm , Benzhydryl Compounds , Dermatitis, Allergic Contact , Epichlorohydrin , Exanthema , Korea , Neck , Patch Tests , Phenols , SkinABSTRACT
Objective To investigate the electrochemiluminescence (ECL) behavior of cloperustine hydrochloride.Methods ECL intensity of tris (2,2'-bipyridyl) rutheniumo(Ⅱ) was enhanced, the method for the determination of dupernstine hydrochloride was established using capillary electrophoresis (CE) coupled with electrochemilumolinescence (ECL) detection. Results Under the optimum conditions, ECL intensity varied linearly with cloperastine hydrochloride concentration from 7.0×10-6 g/mL to 1.0×10-4 g/mL. The detection limit (S/N=3) was 8.05×10-7g/mL. he relative standard deviation of the ECL intensity and the migration time for 11 consecutive injections of 1.0 ×10-5g/mL cloperastine hydrochloride was 2.9% and 1.5%, respectively. This method was successfully applied to eloperastine hydrochloride tablet determination. Conclusion The method has been established, validated and applied for determination of cloperastine hydrochloride.
ABSTRACT
Objective: To investigate the electrochemiluminescence (ECL) behavior of cloperastine hydrochloride. Methods: ECL intensity of tris (2,2′-bipyridyl) rutheniumo(II) was enhanced, the method for the determination of cloperastine hydrochloride was established using capillary electrophoresis (CE) coupled with electrochemilumolinescence (ECL) detection. Results: Under the optimum conditions, ECL intensity varied linearly with cloperastine hydrochloride concentration from 7.0 × 10-6g/mL to 1.0 × 10-4g/mL. The detection limit (S/N=3) was 8.05 × 10-7 g/mL. The relative standard deviation of the ECL intensity and the migration time for 11 consecutive iajections of 1.0 × 10-5 g/mL cloperastine hydrochloride was 2.9% and 1.5%, respectively. This method was successfully applied to cloperastine hydrochloride tablet determination. Conclusion: The method has been established, validated and applied for determination of cloperastine hydrochloride.