Fingerprinting quality control of Qianghuo by high-performance liquid chromatography-photodiode array detection

A novel, simple and accurate fingerprint method was developed using high-performance liquid chromatography-photodiode array detection (HPLC-DAD) for the quality control of Qianghuo, a Tibetan folk and Chinese herbal medicine used as a diaphoretic, an antifebrile and an anodyne. For the first time, the feasibility and advantages of employing chromatographic fingerprint were investigated and demonstrated for the evaluation of Qianghuo by systematically comparing chromatograms of aqueous extracts with the professional analytical software recommended by State Food and Drug Administration (SFDA). Our results revealed that the chromatographic fingerprint combing similarity evaluation could efficiently identify and distinguish raw herbs of Qianghno from different sources and different species. The effects on Notopterygium forbesii Boiss (Apiaceae) chromatographic fingerprints resulted from collecting locations, harvesting time were also examined. (c) 2006 Elsevier lrelanc Ltd. All rights reserved.

Enhancement of atmospheric pressure chemical ionization for the determination of free and glycine-conjugated bile acids in human serum

A highly sensitive and accurate method based on the precolumn derivatization of bile acids (BA) with a high ionization efficiency labeling reagent 1,2-benzo-3,4-dihydrocarbazole-9-ethyl-benzenesulfonate (BDEBS) coupled with LC/MS has been developed. After derivatization, BA molecules introduced a weak basic nitrogen atom into the molecular core structure that was readily ionized in commonly used acidic HPLC mobile phases. Derivatives were sufficiently stable to be efficiently analyzed by atmospheric pressure chemical ionization (APCI)-MS/MS in positive-ion mode. The MS/MS spectra of BA derivatives showed an intense protonated molecular ion at m/z [M + H](+). The collision-induced dissociation of the molecular ion produced fragment ions at [MH - H2O](+), [MH - 2H(2)O](+), [MH - 3H(2)O](+). The characteristic fragment ions were at m/z 320.8, 262.8, and 243.7 corresponding to a cleavage of N - CO, O - CO, and C - OCC, respectively, and bonds of derivatized molecules. The selected reaction monitoring, based on the m/z [M + H]+ -> [MH - H2O](+), [MH - H2O](+), [MH - 2H(2)O](+), [MH-3H(2)O](+), 320.8, 262.8, and 243.7 transitions, was highly specific for the BA derivatives. The LODs for APCI in a positive-ion mode, at an S/N of 5, were 44.36-153.6 fmol. The validation results showed high accuracy in the range of 93-107% and the mean interday precision for all standards was < 15% at broad linear dynamic ranges (0.0244-25nmol/mL). Good linear responses were observed with coefficients of > 0.9935 in APCI/MS detection. Therefore, the facile BDEBS derivatization coupled with mass spectrometric analysis allowed the development of a highly sensitive and specific method for the quantitation of trace levels of the free and glycine-conjugated BA from human serum samples.

2-(2-Phenyl-1H-phenanthro-[9,10-d]imidazole-1-yl)-acetic acid (PPIA) and its application for determination of amines by high performance liquid chromatography with fluorescence detection and identification with mass spectroscopy/atmospheric pressure chemical ionization

A simple, sensitive, and mild method for the determination of amino compounds based on a condensation reaction with 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC-HCI) as the dehydrant with fluorescence detection has been developed. Amines were derivatized to their acidamides with labeling reagent 2-(2-phenyl-1H-phenanthro-[9,10-d]imidazole-1-yl)-acetic acid (PPIA). Studies on derivatization conditions indicated that the coupling reaction proceeded rapidly and smoothly in the presence of a base catalyst in acetonitrile to give the corresponding sensitively fluorescent derivatives with an excitation maximum at lambda(ex) 260nm and an emission maximum at lambda(em) 380nm. The labeled derivatives exhibited high stability and were enough to be efficiently analyzed by high-performance liquid chromatography. Identification of derivatives was carried out by online post-column mass spectrometry (LC/APCI-MS/MS) and showed an intense protonated molecular ion corresponding m/z [MH](+) under APCI in positive-ion mode. At the same time, the fluorescence properties of derivatives in various solvents or at different temperature were investigated. The method, in conjunction with a gradient elution, offered a baseline resolution of the common amine derivatives on a reversed-phase Eclipse XDB-C-8 column. LC separation for the derivatized amines showed good reproducibility with acetonitrile-water as mobile phase. Detection limits calculated from 0.78 pmol injection, at a signal-to-noise ratio of 3, were 3.1-18.2 fmol. The mean intra- and inter-assay precision for all amine levels were < 3.85% and 2.11%, respectively. Excellent linear responses were observed with coefficients of > 0.9996. The established method for the determination of aliphatic amines from real wastewater and biological samples was satisfactory. (c) 2006 Elsevier B.V. All rights reserved.

Development of a sensitive fluorescent derivatization reagent 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC) and its application for determination of amino acids from seeds and bryophyte plants using high-performance liquid chromatography with fluorescence detection and identification with electrospray ionization mass spectrometry

A pre-column derivatization method for the sensitive determination of amino acids and peptides using the tagging reagent 1,2-benzo-3,4dihydrocarbazole-9-ethyl chloroformate (BCEOC) followed by high-performance liquid chromatography with fluorescence detection has been developed. Identification of derivatives was carried out by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS/MS). The chromophore of 2-(9-carbazole)-ethyl chloroformate (CEOC) reagent was replaced by 1,2-benzo-3,4-dihydrocarbazole functional group, which resulted in a sensitive fluorescence derivatizing reagent BCEOC. BCEOC can easily and quickly label peptides and amino acids. Derivatives are stable enough to be efficiently analyzed by high-performance liquid chromatography. The derivatives showed an intense protonated molecular ion corresponding m/z (M + H)(+) under electrospray ionization (ESI) positive-ion mode with an exception being Tyr detected at negative mode. The collision-induced dissociation of protonated molecular ion formed a product at m/z 246.2 corresponding to the cleavage of C-O bond of BCEOC molecule. Studies on derivatization demonstrate excellent derivative yields over the pH 9.0-10.0. Maximal yields close to 100% are observed with a 3-4-fold molar reagent excess. Derivatives exhibit strong fluorescence and extracted detzvatization solution with n-hexane/ethyl acetate (10:1, v/v) allows for the direct injection with no significant interference from the major fluorescent reagent degradation by-products, such as 1,2-benzo-3,4-dihydrocarbazole-9-ethanol (BDC-OH) (a major by-product), mono- 1,2-benzo-3,4-dihydrocarbazole-9-ethyl carbonate (BCEOC-OH) and bis-(1,2-benzo-3,4-dihydrocarbazole-9-ethyl) carbonate (BCEOC)(2). In addition, the detection responses for BCEOC derivatives are compared to those obtained with previously synthesized 2-(9-carbazole)-ethyl chloroformate (CEOC) in our laboratory. The ratios AC(BCEOC)/AC(CEOC) = 2.05-6.51 for fluorescence responses are observed (here, AC is relative fluorescence response). Separation of the derivatized peptides and amino acids had been optimized on Hypersil BDS C-18 column. Detection limits were calculated from 1.0 pmol injection at a signal-to-noise ratio of 3, and were 6.3 (Lys)-177.6 (His) fmol. The mean interday accuracy ranged from 92 to 106% for fluorescence detection with mean %CV < 7.5. The mean interday precision for all standards was < 10% of the expected concentration. Excellent linear responses were observed with coefficients of > 0.9999. Good compositional data could be obtained from the analysis of derivatized protein hydrolysates containing as little as 50.5 ng of sample. Therefore, the facile BCEOC derivatization coupled with mass spectrometry allowed the development of a highly sensitive and specific method for the quantitative analysis of trace levels of amino acids and peptides from biological and natural environmental samples. (c) 2005 Elsevier B.V. All rights reserved.

高效液相色谱法分离测定Fe(Ⅱ)和络合物Fe(Ⅱ)-PDT的两种几何异构体

铁在自然界中广泛分布，其环境和生态效应主要取决于它们的理化性质、形态、溶解性以及与溶解有机质的结合程度。氧化还原性敏感元素铁在自然界主要以Fe(Ⅱ) 和Fe(Ⅲ)两种价态存在。铁在水环境中的地球化学循环作用主要通过其自身的各种价态之间转化以及与周围其他物质(微量元素,营养盐,有机质)之间的相互作用来实现的。不同形态的铁具有不同的理化性质，其环境效应和生物有效性都是不同，不同形态的铁还可对其他重金属在水体中的循环迁移、赋存形态和生物有效性等均产生不同程度的一定的影响。因此，为铁的环境地球化学行为提供更为准确的信息，就必须进行铁的形态分析，由于Fe(Ⅱ)的氧化还原敏感性，故对铁进行形态分析的前提就是准确测定Fe(Ⅱ)的含量，用普通分光光度法测定Fe(Ⅱ)特别是低含量的Fe(Ⅱ)时，结果的不准确性以及高检出限等缺点都严重制约了准确测定环境中Fe(Ⅱ)的含量。为克服用普通分光光度法测定Fe(Ⅱ)时易受样品溶液其他组分的基体干扰以及灵敏度低等缺点，我们采用了用3-(2-吡啶基)-5,6-二苯基-1,2,4-三嗪（PDT）作为柱前衍生试剂，反相离子对高效液相色谱法来测定Fe(Ⅱ)。由于络合物Fe(II)-PDT存在两种几何异构体，故在建立反相离子对高效液相色谱法来测定Fe(Ⅱ)的同时，我们进一步研究了PDT及Fe(II)-PDT的两种几何异构体色谱分离与测定的实验研究，取得了如下进展： 1．研究了有机萃取分光光度法测定Fe(Ⅱ)，实验中发现加入离子对试剂四溴酚酞乙酯钾(TPBE-K)，由于TPBE和Fe(Ⅱ)-PDT络合物生成三元离子缔合物，导致最大特征吸收波长红移，由555nm增加至610nm，三元离子缔合物[Fe(PDT)3] [TBPE]2的摩尔吸收系数由2.4×104 l•mol-1•cm-1增加至1.9×105 l•mol-1•cm-1，显著改善了方法的灵敏度，降低了检出限，在Fe(Ⅱ)含量为0~0.3μg/ml范围内，方法的线性关系良好，线性方程为y＝0.3582x＋0.01，R2＝0.9974，检出限为0.96ng/ml(S/N＝3)，达到ng/ml水平，成功应用于测定泉水及表层湖水的Fe(Ⅱ)及总铁含量。 2．将3-(2-吡啶基)-5,6-二苯基-1,2,4-三嗪(PDT)作为衍生显色络合试剂应用于分光光度法来测定Fe(Ⅱ)已得到较为充分的报道。在上述研究基础上，首次研究了将PDT作为柱前衍生络合试剂，应用于反相离子对高效液相色谱法来测定Fe(Ⅱ)，不仅克服了常规络合衍生分光光度法过量络合剂干扰试验结果的缺点，而且进一步优化选择了检测波长，由于HPLC的应用，突破了在分光光度法中只能在可见波长范围内选择检测波长为555nm，将检测波长选择在Fe(Ⅱ)-PDT吸收更强烈的紫外区295nm处，使得灵敏度得到了显著提高。在Fe(Ⅱ)浓度为0～3.5×10-6mol/l时，峰面积A和峰高H均和标准溶液的浓度C(×10-7mol/l)的线性关系良好,线性回归方程分别为：A=7.4951x-0.0785(R2=0.9999) 和H=0.5126x- 0.0915(R2=0.9997)，方法的检测限为0.35ng/ml (S/N＝3)。本方法测定Fe(Ⅱ)时，不需复杂的前处理步骤，如固相萃取、有机溶液萃取等，直接柱前衍生HPLC方法测定，方便、快速且灵敏度高，低于众多分光光度报道的检出限水准7～170ng/ml范围。 3．由于络合物Fe(II)-PDT存在面式和经式两种几何异构体，故在建立起反相离子对高效液相色谱法来测定Fe(Ⅱ)新方法的同时，进一步研究了PDT及Fe(II)-PDT的两种几何异构体色谱分离及测定实验研究，关于这方面的试验研究，目前来说还没有相关的研究报道，所以将反相离子对高效液相色谱法应用到分离及测定PDT及Fe(II)-PDT的两种几何异构体中，这也是本论文的第二个创新之处。在本实验中，所用的色谱固定相为普通的反相键合C8柱，但由于优化选择了最佳流动相条件，使得PDT及Fe(II)-PDT的两种几何异构体直接在C8柱上得到完全的基线分离。讨论了流动相中有机改性剂乙腈不同比例、不同种类的离子对试剂(高氯酸钠和十二烷基硫酸钠)及不同浓度对PDT和Fe(II)-PDT两种几何异构体的色谱行为的影响。根据不同试验条件下，对所获得的色谱参数(容量因子、分离度、选择性因子、保留时间等)进行了较为深入的分析，据此探讨了色谱分离异构体过程的机理。 4．在上述实验的基础之上，引入更能灵活的调节洗脱强度和被分离物质分离度的三元流动相(乙腈/甲醇/水=20/50/30)，优化选择了三元流动相中有机改性剂的比例以及离子对试剂的种类及浓度，使得异构体的分离度和选择性因子均得到满意结果。对一系列不同浓度的Fe(II)-PDT异构体分离，面式和经式异构体的面积和浓度之间的线性关系在4~25×10-7mol/l范围内良好，线性关系分别为：y = 3.6333x - 2.3378(R2 = 0.9957，经式异构体)，y = 1.8219x - 1.0783(R2 = 0.9964, 面式异构体)。面式和经式异构体的检测限分别为4.28ng/ml和3.44ng/ml (S/N＝3)。 5．进一步研究发现Fe(II)-PDT的两种几何异构体之间存在相互转化的动态平衡，故根据异构体含量随时间的变化就可以探讨在转变过程中的动力学和热力学参数。用高效液相色谱法研究了异构体转变过程中色谱峰高和面积随时间的变化，并依此首次得出了两种几何异构体相互转化可逆过程中的热力学和动力学参数，如熵变、焓变、自由能变化、反应速率常数等，对两种几何异构体之间的动力学转变平衡机理进行合理的解释提供了有益的参考。各不同温度下(30℃，35℃，40℃，45℃)，反应时间在3倍半衰期以内，两种几何异构体之间的相互转变平衡均符合动力学一级反应。各不同温度下Xeln[(Xe-X0)/Xe-X)]随时间变化的线性关系良好，分别为：y=0.0821x+0.7288(R2=0.9911,T=45℃)；y = 0.0486x + 0.598(R2 = 0.9987, T=40℃); y = 0.0216x + 0.5861(R2 = 0.9987, T=35℃)；y = 0.0124x + 0.591(R2 = 0.9988, T=30℃)。 首次将PDT作为柱前衍生络合试剂，应用于反相离子对高效液相色谱法来测定环境中的Fe(Ⅱ)，且无需复杂的样品前处理过程(固相萃取，有机溶液萃取等浓缩富积过程)，克服了普通分光光度法检测限高、易受样品基体效应的干扰等缺点；在建立起的高效液相色谱法测定Fe(Ⅱ)的基础上，进一步对络合物Fe(II)-PDT的两种几何异构体色谱分离及动力学转变实验研究，则是对建立起的反相离子对高效液相色谱法测定Fe(Ⅱ)试验研究基础上的进一步延伸和扩展，是对测定Fe(Ⅱ)的高效液相色谱法的机理的进一步认识和探讨。PDT作为柱前衍生络合试剂首次应用于高效液相色谱测定Fe(Ⅱ)为铁的环境地球化学工作者提供了一种灵敏度高的测定Fe(Ⅱ) 的新途径，为环境样品中Fe(Ⅱ)含量的准确测定提供保证，对铁的环境地球化学研究，如铁形态分析、铁的水-沉积物界面行为等提供更翔实可靠的信息。

贵阳城市污水、河流及养鱼水域抗生素的地球化学行为

抗生素是广泛应用于人类和动物疾病治疗的药物，近年来在养殖业中抗生素更是常常作为动物催长剂。这些抗生素会通过人畜排泄物、药品的丢弃、污水排放、农业废物堆积等途径进入到环境中。环境中抗生素的残留会导致细菌耐药性的产生、微生态平衡的破坏，以及对生态环境和人类难以预知的影响。抗生素污染是近十年来才在国际上引起注意的新型环境问题，目前研究非常薄弱，要深入认识抗生素的环境影响特征就迫切需要对抗生素在环境中的迁移、归宿等地球化学行为进行针对性的研究。本文研究了贵阳城市污水、南明河及乌江渡水库中氯霉素（CAP）、土霉素（OTC）、四环素（TC）和金霉素（CTC）的环境地球化学特征，包括抗生素的分布、迁移特征及影响因素，并分析了贵阳市动物源性食品中抗生素的残留特征，探讨了环境因素对抗生素稳定性及吸附/解吸的影响，以期了解抗生素在环境中的降解、迁移等环境行为的特征，并据此评价抗生素的环境影响特点，为认识抗生素的污染特点、发展相应的控制、治理措施提供科学依据。主要内容包括： 1. 对环境样品中CAP和TCs的前处理过程和检测方法进行了优化。对水样的前处理采用固相萃取的方法，对土壤和沉积物的前处理采用EDTA-McIlvaine振荡萃取、SAX-HLB净化富集的方法；选择HPLC-UV作为分析的手段，对样品中的4种抗生素进行了检测，各化合物得到了很好的分离，该方法简便、快捷、重现性好。 2. 证实了抗生素在城市污水中的普遍存在。贵阳城市污水中以CAP的污染最为突出，其含量明显高于广州地区的城市污水；污水中四环素类药物（TCs）的含量和大多数欧美国家及地区城市污水中TCs的含量相当。这一结果证明，污水中抗生素的污染非常严重，对地表水环境形成严重的威胁。污水中抗生素的来源包括：人服用抗生素药物以及食用含有抗生素残留的食品之后的排泄物；食品中残留的抗生素随剩余食品的丢弃而进入污水。污水中抗生素的含量都具有明显的季节变化规律，冬春季含量明显高于夏秋季，我们认为这与各季节用水量和疾病特点有关。污水中抗生素的大量存在说明：1）抗生素存在滥用的现象，特别是CAP可能存在严重的违法使用，这一现象应当重视；2）污水中的抗生素会威胁水环境质量，应加强处理和控制技术。 3. 证明了城市河流受到多种来源的抗生素影响，其中城市污水是最重要的源，而且抗生素在河流中呈现一定的持久性。南明河上游河段中抗生素的来源包括郊区城镇污水、养鱼场、制药厂污水排放及地表垃圾。城市污水是抗生素的一大排放源，受其影响，排污口下游的河段受到的抗生素污染尤为严重，此外还有来自下游郊区城镇和农业污水的影响。对南明河中抗生素的来源及行为特点的剖析表明：1）抗生素在南明河的水和沉积物中具有不同的分布特征：河水中以CAP的污染最为严重，各水样中CAP的含量基本都高于TCs；而大部分沉积物中TCs的含量都高于CAP。CAP和TCs在水－沉积物之间的分布特征表明，它们在水环境中的地球化学行为明显受到吸附/解吸特点的影响，CAP的亲水性强，而TCs的亲粒性强。2）虽然在养殖业中已禁止使用CAP，但河水和污水中CAP的大量存在，这一结果揭露了CAP仍然存在大量违法使用的事实。3）河水中抗生素含量的季节差异明显，冬春季抗生素的含量高于夏秋季，其原因主要是夏秋季河水流量大，对河水中抗生素的稀释作用强。沉积物中抗生素的含量没有明显的季节变化规律，各季节抗生素的平均含量差异较小。4）受污水影响，各季节排污口下游的南明河水受到的抗生素污染尤为严重，污水口上下游之间的含量差异明显。沉积物在冬季受城市污水的影响大，在排污口下游的抗生素的含量明显高于上游，而春、夏、秋季受污水影响不明显。这可能是因为在春、夏和秋季其它污染源的影响较大，因此污水的影响强度减弱。夏季上游沉积物中CAP的含量甚至高于下游，原因一方面是因为夏季城市污水中抗生素含量低，对南明河的影响减小，另一方面可能因为夏季雨水多、水量大，将上游的地表垃圾以及积累在小河沟中的垃圾冲刷到南明河中，而这些工业及生活垃圾通常是抗生素的一大来源。 抗生素在河流中呈现了一定的持久性，在20 km的范围内水和沉积物显示了明显的污染水平，可能会对水生生态以及流域的环境造成一定的影响。此外，贵阳市南明河两岸的农田通常是用河水进行灌溉的，从而可能使抗生素的污染危及到食物链，对人体的健康有潜在的威胁。 4. 由于食品（如鱼肉、猪肉等）中残留的抗生素是污水中抗生素的重要来源，因此我们研究了养殖业（水产养殖区）中抗生素的环境行为特点，还研究了常见动物源性食品中抗生素的残留特点。对养鱼区的研究表明，养鱼饲料中含有大量的抗生素，是养鱼区水环境中抗生素的来源。由于饲料的颗粒大，且河水流速快，因此饲料中的抗生素不会对水体造成很大的影响，但是在网箱区的沉积物中形成了高含量的残留，除此以外，在乌江渡主航道的河心沉积物中抗生素的含量也相当可观。这说明该区水产养殖的抗生素主要影响沉积物，且相应的污染不是局部的，而会影响到整个养鱼区。从沉积物中抗生素平均含量的季节差异来看，各季TCs的用药量可能不同，秋季的用量可能最大，而其它季节用量相当；CAP在每个季节的用量可能相当。尽管分析的饲料中CAP的含量高于TCs，但在各季节乌江渡水库主航道沉积物中TCs的含量几乎都远高于CAP，这是由于TCs比CAP更易存在于沉积物中。乌江渡水库上下游主航道沉积物中抗生素的含量差异日益减小，说明了乌江渡水库的污染已日益严重。这些抗生素虽然暂时被固定在了沉积物中，活性相对较小，但是在外界条件的影响下，它们的吸附/解吸作用是可以改变的。因此，乌江渡沉积物中高含量的抗生素残留对水生生态环境具有潜在的威胁。 对贵阳市动物源性食品（鱼肉、猪肉和牛奶）中抗生素的残留特征的分析表明，这些食品中普遍残留有高含量的抗生素，被人食用后，其中的抗生素也会被人体摄入，一部分抗生素会随着人的排泄物而进入到污水中，并最终进入环境中；另一部分抗生素会积蓄在人体中，到一定浓度后就会对人体造成伤害。鲩鱼、鲫鱼、鲤鱼、鳙鱼和鲢鱼中CAP、OTC、TC和CTC的平均含量范围分别在57.1～365.5 µg/kg、37.0～60.7µg/kg、34.3～72.4 µg/kg和22.3～68.0 µg/kg之间。秋季的长吻鲍、团头鲂、鲈鱼、长江洄鱼、对虾、黄鳝、泥鳅和猪肉中CAP、OTC、TC和CTC的平均含量分别为88.1 µg/kg、76.8 µg/kg、343.7 µg/kg和34.2 µg/kg；牛奶中的含量分别为4.2 µg/L、1.6 µg/L和2.5 µg/L和22.0 µg/L。 5. 确定了抗生素的稳定性、吸附/解吸作用与各因素的关系，据此可以探讨环境中抗生素的行为特点及机理，并为环境中抗生素的污染防治提供依据。在短时的加热中，TCs发生了部分降解，而CAP却几乎没有发生分解，说明烹饪过程中的加热不能去除食品中的抗生素，这些抗生素则会进入到人体和污水中。当pH在2～8之间时，TCs的性质较稳定；CAP在pH＝1～10的范围内性质都比较稳定。地表和地下水体的pH一般为7左右，说明了水环境中TCs受pH的影响小，而CAP几乎不受影响。在6个月的时间内，TCs在3种介质中都发生了不同程度的降解，但仍有大部分残留；CAP几乎没有发生明显的降解，这说明CAP和TCs的污染具有一定的持久性。TCs的吸附/解吸受颗粒物粒径的影响大，粒径越小，吸附越强；CAP不受影响。这证明了在环境中CAP更易以水溶态的形式存在，而TCs更易吸附在沉积物中。颗粒物中CaCO3含量越高，TC和CTC的吸附越强，证明CaCO3可以用来增强TC、CTC的吸附，这为环境中抗生素的治理提供了一条思路。

生物标志化合物数据处理及在重建古环境中的应用

本文主要地讨论了生物标志化合物数据的处理方法，并将此应用到古沉积环境的判别方面。全文共分三大部分：第一部分主要讨论是多元统计方法对那些研究的较成熟的烃类生物标志经合物参数在重建古沉积环境方面的应用。对分别采自不同沉积环境的共46个样品中生物标志化合参数的R型因子分析和对应分析的结果表明：生物标志化合物参数可以有效地将所研究的样品分划分为：淡水湖泊相沉积环境、半咸水/淡水环境、膏盐环境和泻湖沉积具火山灰沉积的环境，与此对应，生物标志化合物参数也可以划分为相应的四大类，即：（1）反映淡水湖泊沉积环境的特征为：含较丰富的高等植物输入的生物标志化合物，如：具较高的H/L，C31/C17，C27/C17比值等；（2）半咸水/淡水沉积环境以较高的Pr，Pr/Ph和高的藿/甾比为特征；（3）生物标志化合物参数组合{Ph/nC18，植烷，升藿烷指数，伽玛蜡烷指数， iC25+iC30和Tm/Ts}是反映膏盐环境的一类生物标志化合物参数；（4）可能可以用含较高的C28甾烷和β-胡萝卜烷来区分泻湖具火山灰沉的环境。正构烷烃的分布对于划分沉积环境远不如上述的综合生物标志化合物那样有效，但是，中等链长的正烷烃（C22，C24）在主因子分析的第二主因子轴上有较大的载荷，反晨了含较高中等链长的正烷烃是膏盐环境的一个特征，对其归一化数据的主因子分析的结果还表明，它是一类较好的物源（高等植物或低等生物输入）指示生物标志化合物。第二部分讨论的是膏盐环境中含硫化合物GC/MS数据的处理方法，通过初步确定碳数、碳数的校正、计算TPI（噻吩指数，Thiophene index）和CTPI、确定噻吩代码、建立TPmaster、绘制噻吩化合物分布的“指纹图”等步骤，在大多数噻吩化合物结构不明的情况下，快速地、清楚地、有条理地将这类化合物的所有异构体表达出来，同时还实现了这类化合物在不同样品间的自动对比和直观对比；大大的方便了这类化合物在地质体中应用的研究。在理论上，可以通过TPmater寻找新的异构体和新的噻吩化合物系列。第三部分在计算同位素分子峰发布的基础上，快速地、有效地处理了卟啉的质谱数据，即：卟啉Probe-MS数据的同位素校正。这一计算程序不仅可以计算混合分子同位素峰束中单个化合物的含量（即质谱数据的同位素校正），还可以计算任意化合物和混合物的分子同位素峰的分布类型，通过对比质谱数据，有助于地质体中新的卟啉化合物的发现和鉴定。在用该计算程序处理了采自不同沉积环境的七个样品的Probe-MS数据后，再加上自由基卟啉的HPLC特征，结果表明：可以用卟啉的分布将淡水/半咸水环境的沁阳凹陷、泻湖相具火山灰沉积的准葛尔盆地和四川三叠系海相区分开来。

Prediction of soil adsorption coefficients from retention parameters on three reversed-phase liquid chromatographic columns

Reversed-phase high-performance liquid chromatographic (RP-HPLC) retention parameters, which are determined by the intermolecular interactions in retention process, can be considered as the chemical molecular descriptors in linear free energy relationships (LFERs). On the basis of the characterization and comparison of octadecyl-bonded silica gel (ODS), cyano-bonded silica gel (CN), and phenyl-bonded silica gel (Ph) columns with linear solvation energy relationships (LSERs), a new multiple linear regression model using RP-HPLC retention parameters on ODS and CN columns as variables for estimation of soil adsorption coefficients was developed. It was tested on a set of reference substances from various chemical classes. The results showed that the multicolumn method was more promising than a single-column method was for the estimation of soil adsorption coefficients. The accuracy of the suggested model is identical with that of LSERs.

Drug-protein interaction studied by technique of microdialysis combined with high performance liquid chromatography

Drug-protein binding is an important process in determining the activity and fate of a pharmaceutical agent once it has entered the body. This review examines the method of microdialysis combined with high-performance liquid chromatography (HPLC) that has been developed;by ours to study such interactions, in which the microdialysis was applied to sample the free drug in the mixed solution of drug with protein, and HPLC to quantify the concentration of free drug in the microdialysate. This technique has successfully been used for determining various types of binding interactions between the low affinity drugs, high affinity drugs and enantiomers to HSA. For the case of competitive binding of two drugs to a protein in solution, a displacement equation has been derived and examined with four nonsteroidal anti-inflammatory drugs and HSA as model drugs and protein, respectively. Microdialysis with HPLC was adopted to determine simultaneously the free solute and displacing agent in drug-protein solutions. The method is able to locate the binding site and determine affinity constants even up to 10(7) L/mol accurately.

Theoretical study of the separation mechanism of ionizable compounds in capillary electrochromatography

The migration mechanism of ionizable compounds in capillary electrochromatography (CEC) is more complicated than in high performance liquid chromatography (HPLC) due to the involvement of electrophoresis and the second chemical equilibrium. The separation mechanism of ionizable compounds in CEC has been studied theoretically. The electrochromatographic capacity factors of ions (k *) in CEC and in the pressurized CEC are derived by phenomenological approach. The influence of pH, voltage, pressure on k* is discussed. in addition, the k * of weak acid and weak base are derived based on acid-base equilibrium and the influence of pH on k * is studied theoretically.

New method for analysis of Chinese herbal complex prescription and its application

Traditional Chinese medicine (TCM) is a great treasure of China, the analysis of which is an arduous task. The viewpoint that all chemical constituents of Chinese herbal complex prescription should be analyzed as a black box is elucidated for the first time. Intelligent multi-mode multi-column chromatographic system (IMMCC) with its hybrids is the basic method and HPLC Unified Method is the breakthrough for the black box analysis. Dang-Gui-Bu-Xue-Tang was selected as a typical TCM and a systematic separation method from non-aqueous mobile phase to pure water mobile phase was put forward in order to convert unknown sample to known sample. The a, c values and UV spectra of 66 components of Astragalus, 78 components of Angelica and 71 components of Dang-Gui-Bu-Xue-Tang were obtained. Intelligent optimization and peak identification method and software for complex samples were developed and the optimum multi-step multi-binary gradient curve of mobile phase for Astragalus was ascertained. The maximum error and minimum error of predicted retention time for all components of Astragalus are 8.62% and 0.05% respectively. All components of Astragalus were compared with those of Angelica and it is found that many components of Astragalus are the same as those of Angelica, while the contents of these components are different. Many components of Dang-Gui-Bu-Xue-Tang are also the same as those of Astragalus and Angelica with different contents.

Reversed-phase high-performance liquid chromatographic investigation of urinary normal and modified nucleosides of cancer patients

Post-transcriptional modifications in RNA give rise to free modified ribonucleosides circulating in the blood stream and excreted in urine. Due to their abnormal levels in conjunction with several tumor diseases, they have been suggested as possible tumor markers. The developed RP-HPLC method has been applied to analyze the urinary nucleosides in 34 urinary samples from 15 kinds of cancer patients. The statistical analyses showed the urinary nucleoside excretion, especially modified nucleoside levels, in cancer patients were significantly higher than those in normal healthy volunteers. Factor analysis was used to classify the patients with cancer and normal healthy humans. It was found that using 15 urinary nucleoside levels or only five modified nucleoside levels as data vectors the factor analysis plot displayed two almost separate clusters representing each group. (C) 1999 Elsevier Science B.V. All rights reserved.

Chiral separation of enantiomers of amino acid derivatives by high-performance liquid chromatography on a norvancomycin-bonded chiral stationary phase

A novel norvancomycin-bonded chiral stationary phase (NVC-CSP) was synthesized by using the chiral selector of norvancomycin. The chiral separation of enantiomers of several dansyl-amino acids by high-performance liquid chromatography (HPLC) in the reversed-phase mode is described. The effects of some parameters, such as organic modifier concentration, column temperature, pH and flow rate of the mobile phase, on the retention and enantioselectivity were investigated. The study showed that ionic, as well as hydrophobic interactions were engaged between the analyte and macrocycle in this chromatographic system. Increasing pH of buffers usually improved the chiral resolution for dansyl-alpha-amino-n-butyric acid (Dns-But), dansyl-methionine (Dns-Met) and dansyl-threonine (Dns-Thr), but not for dansyl-glutamic acid (Dns-Glu) which contains two carboxylic groups in its molecular structure. The natural logarithms of selectivity factors (In alpha) of all the investigated compounds depended linearly on the reciprocal of temperature (1/T), most processes of enantioseparation were controlled enthalpically. Interestingly, the process of enantioseparation for dansyl-threonine was enthalpy-controlled at pH of 3.5, while at pH of 7.0, it was entropy-controlled according to thermodynamic parameters Delta(R,S)DeltaHdegrees and Delta(R,S)DeltaSdegrees afforded by Van't Hoff plots. In order to get baseline separation for all the solutes researched, norvancomycin was also used as a chiral mobile phase additive. In combination with the NVC-CSP remarkable increases in enanselectivity were observed for all the compounds, as the result of a "synergistic" effect. (C) 2003 Elsevier B.V. All rights reserved.

Discriminating the xanthones in an extract of Swertia franchetiana by retention parameters

The quantitative structure-retention relationship is one of the most actively studied topics in the field of chromatography. In this paper, retention parameters of components were used to discriminate the xanthones in a methanol extract of Swertia franchetiana. The extract was analysed by HPLC under two different multistage linear gradient conditions and the retention parameters calculated from these retention data. It was found that the retention parameters of xanthones are in a specific region in the plot of log k(w) vs. S and the xanthones in the extract could be distinguished from other components by this feature. Furthermore, xanthone aglycones and xanthone glucosides could also be discriminated by retention parameters. Copyright (C) 2005 John Wiley Sons, Ltd.

Characteristics of elution profile in radial chromatography under linear conditions

Based on the mass balance equations of solute transfer in the radial chromatographic column, the theoretical expression to describe the column efficiency and shape of elution profile is obtained under linear isotherm case. Moreover, the tendency for the variation of column efficiency and symmetry of peak profile is systematically discussed. The results showed that in radial chromatography the relationship between the column efficiency and volumetric flow rate is similar with that relationship in axial chromatography; relatively high column efficiency still can be obtained under high flow rate in radial chromatography. Accompanying the increase of retention factor of solutes and injection time, the column efficiency decreases monotonously. The effect of column diameter and column length on the column efficiency interfere with each other. It is more advantageous to increase the column efficiency by applying columns with larger column diameter and shorter column length. According to the discussion of the effect of diffusion on the column efficiency, radial chromatography is proved to be suitable for the separation of samples with relatively high diffusion coefficient, which predicts its obvious advantage in the preparative separation of samples such as proteins and DNA.