Measurement of elastin hydrolysis by reverse-phase HPLC with on-line post-colum derivatization

Summary Excessive breakdown of elastin, a structural protein, may be related to aortic disease and emphysema. Since L-valyl-L-proline occurs in high concentrations in elastin, a rapid and sensitive method using HPLC with post-column on-line derivatization was used to measure the dipeptide from swine aortic tissue, and the amount of elastin present was determined. Elastin was extracted by alkaline hydrolysis. After neutralization and filtration, the sample was injected onto a ODS-2 gel column, and the dipeptide was eluted by a linear gradient of 0 to 10% of 1-propanol in 50 mM heptafluorobutyrate, pH 3, at a flow rate of 1 ml/min. The eluent was reacted with fluorescamine at pH 8.6, and fluorescence was detected at an excitation wavelength of 395 nm and a 455 nm cutoff emission filter.Presented at the 17th International Symposium on Chromatography, September 25–30, 1988, Vienna, Austria.     

A method for the determination of histamine in wine by HPLC with precolumn derivatization with phenylisothiocyanate

Summary A method for determining histamine in wine by precolumn derivatization with PITC (phenylisothiocyanate) with reversed-phase HPLC and UV detection is reported. Histamine can be determined together with the 24 amino acids within 40 min, or separately in a shorter time (less than 4 min) if a prior solid phase extraction clean-up is used.     

新型紫外衍生试剂用于D-氨基酸的毛细管电泳分离

采用一种新型紫外衍生试剂对乙酰氨基苯磺酰氟(PAABS-F)作为柱前衍生试剂,成功地对19种标准D-氨基酸和甘氨酸进行了衍生和毛细管电泳分离。详细研究了各种分离条件对毛细管电泳分离的影响。实验结果表明:采用20 mmol/L硼砂(pH 9.3),126 mmol/L十二烷基硫酸钠,8 mmol/Lβ-环糊精和20mmol/L NaC l时分离结果最佳,16 m in内实现了20种氨基酸的基线分离。     

Luminarin 9: A new fluorescent reagent for the precolumn derivatizatioin of main volatile and non-volatile N-nitrosamines in RP-HPLC

Summary A sensitive precolumn fluorescence derivatization method for low level detection of the, volatile (N-nitrosodimethylamine and N-nitrosopyrrolidine) and non-volatile N-nitrosamines (N-nitrosoproline and N-nitrosodiethanolamine) an high-performance liquid chromatography was developed. This method is based on the denitrosation of the compounds of interest by a mixture of hydrobromic acid and acetic acid to produce the corresponding secondary amines. These are, then, able to react with, a quinolizinocoumarin derivative (luminarin 9^®) to form highly fluorescent labelled N-nitrosamine derivatives. The structural elucidation of the luminarin 9^® derivatives of N-nitrosoproline and N-nitrosodimethylamine by way of example, were established by liquid chromatography-mass spectrometry (LC-MS) and by direct chemical ionization-mass spectrometry (CI-MS). The separation, derivatization and detection conditioins were optimized for all the studied compounds. The detection limits (signal to noise ratio=3) were between 0.4 and 1.0 pmol injected depending on the compound. The calibration graphs were linear for derivatized amounts in the range of 0.5–40 nmol for N-nitrosodimethylamine and N-nitrosopyrrolidine, 0.4–2- nmol for N-nitrosoproline and 1.0–40 nmol for N-nitrosodiethanolamine. The repeatability (RSD less than 3.5%, n=6) and reproducibility (RSD less than 4.8%, n-6) were satisfactory.     

Enhancing chemical analysis with signal derivatization using simple available software packages

Derivative techniques for analytical signal processing are useful for solving some noise and signal resolution problems in various fields of study such as titrimetry, spectrophotometry, chromatography and electrochemistry. The broad use of these techniques, however, is often limited by costly inflexible built-in software packages in commercial analytical instruments. We propose here the application of commercial simple software packages such as Microsoft® Excel and Microcal Origin for signal smoothing and fitting, and for obtaining derivative analytical signals in batch and flow-based analyses, including potentiometric titration, spectrophotometry, chromatography, voltammetry and sequential injection analysis (SIA). The worldwide (especially Excel) software packages are easy-to-use for less experienced users and have also capabilities for advanced users, and therefore employing such packages can result in expansion of useful derivative techniques. We demonstrate application of the available package-aided derivative capabilities for enhancing some chemical analyses, including potentiometric acid–base titration, Bradford assay of protein, chromatographic separation of ajmaline and reserpine and anodic stripping voltammetry of copper. The derivative signals from smoothed and fitted curves offer better accuracy and precision, even for non-resolving peaks and tailing peaks. In some cases, the optimization of experimental conditions is not further required, which can lead to fast method development.     

免疫亲和柱净化/柱前衍生化-高效液相色谱荧光检测法测定粮谷中的T-2毒素

建立了免疫亲和柱净化/柱前衍生化-高效液相色谱荧光检测器测定粮谷中T-2毒素含量的方法。样品经甲醇-水(体积比为80∶20)混合溶剂提取,通过免疫亲和柱(IAC)净化,以氰酸蒽(1-AN)为衍生化试剂、4-二甲基氨基吡啶(DMAP)为催化剂进行衍生,以ZORBAX Eclipse XDB-C18 柱为分离柱,乙腈-水(体积比为80∶20)为流动相进行高效液相色谱分离及荧光检测，荧光检测的激发波长为381 nm，发射波长为470 nm。T-2毒素的质量浓度为0.01~1.5 mg/L时与峰高呈良好的线性,相关系数为0.9985。在0.01~1.5 μg/g添加水平下，回收率为79.7%~94.5%,相对标准偏差小于7%；检出限（S/N＝3）为0.01 μg/g。该方法净化效果好,灵敏度高,操作简便快速。