超临界流体萃取技术与中药现代化

本文介绍了超临界流体萃取技术的基本原理和工艺流程、该技术在中草药有效成分提取分离中的应用以及超临界流体色谱技术。     

Validation of direct assay of an aqueous formulation of a drug compound AZY by chiral supercritical fluid chromatography (SFC)

Supercritical fluid chromatography (SFC) is increasingly being recognized as a powerful technique for analysis of pharmaceutical compounds in various dosage forms. Assay of aqueous formulations of research compounds by SFC is, however, a relatively unexplored area primarily due to the potential problems associated with it. This work describes the development of a direct assay of a chiral drug compound AZY in a 100% aqueous formulation by SFC, and its qualification following ICH and FDA validation guidelines on chromatographic methods. The results indicated that SFC has the potential for assaying aqueous formulations of research compounds with high degree of selectivity, accuracy, precision, robustness, sensitivity, and linearity over a wide range of concentrations. This work also confirmed a previous hypothesis that direct formulation assay by SFC approach is applicable to both acidic and basic pharmaceutical compounds with equal degree of success.     

Chiral interconversion monitoring of a drug candidate by supercritical fluid chromatography (SFC)

Stereoisomer interconversion of chiral drug substances is of significant importance if it occurs within pharmacological and pharmaceutical time scales and under physiological and shelf life conditions. Several analytical techniques exist for the determination of first order rate constants and enantiomerization energy barriers by dynamic and stopped flow chromatography, mathematical models and functions, and computer programs. The focus of this work is to utilize a simple supercritical fluid chromatography (SFC) chiral assay to determine the possibility of interconversion of the desired R and less active S isomers of a drug candidate. The rate constants of racemization and enantiomerization, the half life of racemization, and enantiomerization energy barriers were determined for the R → S (or, forward) and S → R (or, reverse) conversions. The method was selective and sensitive enough to detect less than 1% interconversion occuring under the conditions studied. The method also demonstrated that R ? S racemization was possible only under extreme conditions of prolonged heating (80 °C) and highly basic pH (9.5).     

超临界二氧化碳流体从荜茇中萃取胡椒碱

目的：建立超临界流体萃取法（ＳＦＥ）提取中药荜茇中的胡椒碱的方法。方法：经系统观察法考察了ＳＦＥ的最佳条件,并用反向高效液相色谱法进行分离测定。结果：ＳＦＥ的最佳萃取条件为：萃取压力３８．５ＭＰａ,萃取温度７０℃,改性剂量０．４ｍｌ,静脉萃取时间５ｍｉｎ及动态萃取体积５ｍｌ。结论：ＳＦＥ法简便快捷,效率较高,可以作为荜茇及其制剂分析时的预处理手段。     

超临界CO2萃取铜藻中岩藻黄质的工艺研究

摘 要: 目的 采用超临界CO2流体从铜藻中萃取岩藻黄质。方法 以岩藻黄质提取率为指标,采用单因素试验研究萃取时间、粉碎粒度、萃取压力、萃取温度对超临界CO2流体萃取岩藻黄质的影响。结果 通过正交试验得到最佳提取工艺为:样品粉碎粒度80目,在萃取温度40 ℃、萃取压力25 Mpa下萃取30 min,在最佳萃取条件下,岩藻黄质提取率为 1.12 mg?g-1。结论 优选出超临界二氧化碳萃取铜藻中岩藻黄质的最佳工艺,为进一步开发应用提供参考依据。     

超临界流体色谱拆分齐留通对映体

目的研究超临界流体色谱对齐留通对映体的分离效果。方法考察确定5种手性柱中分离效果最佳的柱子,在该柱上考察改性剂的种类、浓度、温度及背压对分离度的影响。结果采用Chiralpak AD-H柱、改性剂为异丙醇,异丙醇在流动相中比例为18%、背压为140 bar、温度为303 K时,齐留通对映体达到最佳分离,分离度为11.5,出峰时间分别为4.59、7.35min。结论所用超临界流体色谱的方法简单、快捷、经济有效,适用于齐留通对映体的分离。     

超临界流体色谱在药物分析中的研究进展

超临界流体色谱是以超临界流体为流动相,依靠流动相的溶剂化能力来进行分离分析的一种色谱,具有分离速度快、效能高、绿色环保等优点,是气相色谱和液相色谱的有力补充,具有广阔的应用前景。本文利用中国知网（CNKI）、ScienceDirect、PubMed等数据库检索了近几年来超临界流体色谱在药物分析中的研究文献,归纳了其在手性药物、天然产物、药物代谢和杂质检测中的应用进展,并对今后的进一步研究提出思考。     

Supercritical fluid chromatography-tandem mass spectrometry for fast bioanalysis of R/S-warfarin in human plasma

Chiral separation for the analysis of enantiomers in biological fluids by HPLC often takes relatively long chromatography time compared to achiral analysis. The advantage of fast mass transfer in packed-column supercritical fluid chromatography (pSFC) and the high-flow compatibility of APCI-MS/MS were applied to develop a fast bioanalytical method for R/S-warfarin in human plasma. Presented here are the main challenges encountered during method development of a semi-automated liquid extraction SFC-MS/MS method. The selection of internal standard, robustness of the SFC equipment, and carryover issues are discussed. The method has high-throughput: the chromatography time is at least two-fold faster than the our fastest previous method; and the liquid/liquid extraction time of 96 samples is less than 20 min using a Tecan Genesis^® RSP 100 pipetting station and a Tomtec Quadra-96^® workstation. The standard curve range was 13.6–2500 ng/ml. Precision of QC concentrations from four validation runs was 7.0% for R-warfarin and 6.0% C.V. for S-warfarin; and the bias was 3.7 and 3.2% R.E., respectively. The method is sensitive, accurate, selective and robust, and was applied to a drug-interaction clinical study with rapid turnaround of sample analysis.     

Exploration of liquid and supercritical fluid chromatographic chiral separation and purification of Nutlin-3--a small molecule antagonist of MDM2

Inhibition of the MDM2–p53 interaction can stabilize the p53 protein and offer a novel strategy for cancer therapy. The imidazoline compound (Nutlin-3) is a promising small molecule antagonist of the MDM2–p53 interaction. This compound was synthesized as a racemic mixture, and one enantiomer is 100–200-fold more active than the other enantiomer. In this study, various enantiomeric separation approaches were explored to resolve the Nutlin-3 enantiomers using chiral supercritical fluid chromatography (SFC) as well as chiral liquid chromatography (LC) under normal phase mode, reversed phase mode and polar organic phase mode. The chiral SFC method based on Chiralcel OD column showed superior separation in terms of selectivity and efficiency. Optimization of the chiral separation method enabled high throughput preparative scale purification. Ultimately, 5 g of racemic mixture were purified on Prep-SFC in 75 min with the recovery rate above 92%.     

Application of packed column supercritical fluid chromatography to the analysis of barbiturates

The analysis of barbiturates using packed column supercritical fluid chromatography (SFC) has been investigated. The separations were carried out on either an ODS bonded silica or polystyrene—divinylbenzene polymer column with carbon dioxide as the mobile phase and flame ionisation detection. All the barbiturates were strongly adsorbed on the ODS-silica column, but reasonable separations were obtained by using the polymer column.     

Chiral separation of potent corticotropin-releasing factor-1 receptor antagonists by supercritical fluid chromatography

Pyrazinones bearing an N-1-alkyl chain with a chiral center have been reported as potent antagonists of the corticotropin-releasing factor-1 receptor (CRF1R). Separation of individual enantiomers for preclinical testing was an important aspect of lead optimization. To evaluate the applicability and efficiency of supercritical fluid chromatography (SFC) for enantiomeric resolution of this class of compounds, enantiomeric pairs of eight pyrazinones with different structural characteristics were tested under an array of SFC conditions. The results showed that pyrazinones with a 1-cyclopropyl-2-methoxyethyl substituent were readily separated with a Chiralpak AD-H or Chiralcel OD-H column with ethanol as the modifier. On the other hand, analogs with a less polar alkyl substituent were not amenable to the general method and required further optimization of the chromatographic conditions. In addition, structural variations on the pyrazinone core and aromatic moiety had an impact on the chiral resolution of this class of compounds. This investigation led to the development of efficient chiral SFC methods for separating all eight pyrazinone enantiomeric pairs encompassing an array of structural variations.     

Simultaneous determination of roxithromycin and ambroxol hydrochloride in a new tablet formulation by liquid chromatography

A rapid and accurate liquid chromatographic method is described for the simultaneous determination of roxithromycin and ambroxol hydrochloride in a new tablet formulation. Chromatographic separation of the two drugs was achieved on a Diamonsil™ C₁₈ column (200 mm×4.6 mm, 5 μm). The mobile phase consisting of a mixture of acetonitrile, methanol and 0.5% ammonium acetate (39:11:50 (v/v), pH 5.5) was delivered at a flow rate of 1.0 ml/min. Detection was performed at 220 nm. Linearity, accuracy and precision were found to be acceptable over the concentration range of 201.2–2012.0 μg/ml for roxithromycin and 42.7–427.0 μg/ml for ambroxol hydrochloride, respectively. Separation was complete in less than 10 min. The proposed method can be used for the quality control of formulation products.     

A comparison between solid phase extraction and supercritical fluid extraction for the determination of fluconazole from animal feed

The application of supercritical fluid extraction with carbon dioxide and modified carbon dioxide for the determination of fluconazole from an animal feed was studied. A fractional factorial design approach was used to examine the significant experimental variables for quantitative extraction of fluconazole. Gas chromatography with either flame ionisation or mass selective detection was used for quantitation of the extracts. The results indicated that modifier (methanol) had the greatest effect on the recovery of fluconazole from the animal feed.     

超临界流体萃取法毛细管气相色谱法分析牡丹皮及制剂中丹皮酚的含量

采用超临界流体萃取法(supercritical fluid extraction,SFE)提取中药牡丹皮及其成方制剂中丹皮酚,以氯仿作改性剂,在温度90℃,压力28MPa下,二氧化碳动态萃取体积3ml;静态萃取时间5min。此法简便快速,萃取完全。用大孔径毛细管柱气相色谱法作含量监测,结果:相关性好(γ=0.9999),中药与制剂的回收率分别为97.8%,RSD=2.35%(n=3);100.3%,RSD=1.89%(n=3)。为中药有效成分的提取和质量控制提供了一种有效可靠的方法。     

均匀设计方法在香附超临界流体萃取中的应用

目的：研究超临界CO2流体萃取（SFE）香附有效成份的最佳条件。方法：采用均匀设计方法,以萃取压力、萃取温度;解析压力、解析温度4个因素,每个因素7个水平进行实验。结果：在SFE中最佳萃取条件：萃取压力15Mpa,萃取温度40℃：解析压力8Mpa以上,解析温度36℃。结论：本法是一种简便、高选择性和高效率的提取方法。     

Intelligent polymeric micelles: development and application as drug delivery for docetaxel

Recent years, docetaxel (DTX)-loaded intelligent polymeric micelles have been regarded as a promising vehicle for DTX for the reason that compared with conventional DTX-loaded micelles, DTX-loaded intelligent micelles not only preserve the basic functions of micelles such as DTX solubilization, enhanced accumulation in tumor tissue, and improved bioavailability and biocompatibility of DTX, but also possess other new properties, for instance, tumor-specific DTX delivery and series of responses to endogenous or exogenous stimulations. In this paper, basic theories and action mechanism of intelligent polymeric micelles are discussed in detail, especially the related theories of DTX-loaded stimuli-responsive micelles. The relevant examples of stimuli-responsive DTX-loaded micelles are also provided in this paper to sufficiently illustrate the advantages of relevant technology for the clinical application of anticancer drug, especially for the medical application of DTX.     

Development of a liquid chromatography method for the analysis of josamycin

Out of three methods for the analysis of josamycin, the best one was selected and used as starting point for further development. A central composite design was applied to find the most influencing parameters and to optimize the chromatographic conditions and a full factorial design was used to perform a robustness study. The final method uses a Hypersil ODS column 5mum, 250mmx4.6mm i.d. maintained at 45 degrees C. The mobile phase is composed of acetonitrile-phosphate buffer (pH 3, 0.2moll(-1))-tetrabutylammonium hydrogen sulphate 0.2moll(-1)-water (21:5:3:71, v/v/v/v). Strongly retained impurities after the main peak require gradient elution, which is obtained by increasing linearly the acetonitrile concentration (from 21% to 50%, v/v) and decreasing the TBA concentration (from 3% to 0%, v/v) in the mobile phase. The total run time was 65min. UV detection is performed at 232nm and the flow rate is 1ml/min. The method shows good selectivity, precision, linearity and sensitivity. Five commercial bulk samples were analyzed.     

黄荆子超临界CO_2萃取物化学成分的研究

目的分析黄荆子超临界CO2萃取物的挥发性成分。方法采用超临界CO2萃取法对黄荆子药材进行提取,运用毛细管气相色谱-质谱法对萃取物的挥发性成分进行分析,用气相色谱面积归一化法测定各成分的相对百分含量。结果经毛细管色谱分离出91个峰,并鉴定出了其中的60个化合物,所鉴定的化学成分约占色谱峰总面积的96%。其主要化学成分为正癸醇(71.22%)、2,5,5,8a—四甲基—八氢—2H—苯并吡喃(4.96%)、β—石竹烯(2.29%)、环己烯(1.86%)、蛇床子素(1.77%)、4—羟基—4—甲基—2—戊酮(1.61%)、9—(3—丁烯基)蒽(1.11%)等。结论本研究可为进一步开发利用黄荆子提供科学依据。     

键合型多糖手性固定相超临界流体色谱用于取代苯基哌嗪衍生物的拆分(英文)

合成了六个新型的1-取代苯基-4-[3-(吲哚-4-氧基)-2-羟丙基]-哌嗪消旋体化合物1–6,并利用键合型多糖手性固定相超临界流体色谱完成了对其的手性拆分研究。实验表明,基于固定相Chiralpak ⅠA的拆分优于其它两种键合型固定相(Chiralpak ⅠB、Chiralpak ⅠC),改性剂异丙醇的效果优于乙醇、甲醇、四氢呋喃、乙腈、二氯甲烷。本文还研究了有机改性剂、背压与柱温对六个化合物对映体拆分的影响。结果表明,改性剂组成最大程度地影响了分离时间,柱温的改变对分离时间的影响较小但最大程度地影响了对映体选择性的变化。最优的拆分条件为:固定相为Chiralpak ⅠA柱,柱温为35℃,背压为120bar,流动相为35%异丙醇(含0.1%二乙胺),流速为3.0mL/min,紫外检测波长为283nm。该六个消旋体化合物在10分钟内得到了良好的拆分。超临界流体色谱法是拆分该类化合物对映体的有效方法。