手性药物的HPLC检测和圆二色谱检测

智能文字提取功能测试中

目前，在生产过程中很少能控制化合物的手性形式。这种缺乏控制通常导致两种手性形式产生等量。在分子水平上，这两个手性伴侣具有相同的物理性质，因此分离起来非常困难和昂贵。分离的一个更优越的替代方案是首先形成单一纯手性化合物的方法。本申请说明将描述使用HPLC对几种非处方药物的手性检测。本应用说明结论：高效液相色谱CD检测法是一种分离和检测包括药物在内的手性化合物的优良方法。荧光检测器和CD检测器都对浓度非常敏感；然而荧光检测器不能给出关于哪种对映异构体是哪种的任何指示。光学旋转检测器能够区分对映体，但在所有研究的材料中发现灵敏度要低得多。一般来说，与OR一起使用的最佳材料是糖。Jasco公司目前提供世界上唯一的高效液相色谱CD检测器。HPLC APPLICATION NOTE 01-04Detection of Chiral Drugs UsingHPLC with CD Detection Recent American Food and Drug Administration guidelines have effectively determined that mixtures of chiral compounds can no longer be brought to the pharmaceuticals market place. For new pharmaceuticals， each chiral form must be pure and separately assessed for registration. The need for chiral detection in the development of pharmaceuticals is increasing due to regulatory changes and the demand for single isomers. The key pharmaceutical industry objective is now racemic switching' and single-isomer drugs. These guidelines require a means for chiral detection and compound separation. Introduction Many molecules made up of the same number and type of atoms come in two forms commonly referred to as left-handed and right-handed isomers. This is not just some interesting quirk of nature： left-handed and right-handed forms of the same molecules can have profoundly different properties because of their chiralform. In the food industry for example， limonene (left-handed) smells like lemons； limonene (right-handed)smells like oranges. One form of asparagine is used as a sweetener i n foods and beverages， whilst its chiral partner has a bitter taste. Different chiral forms of molecules can have profoundly different biological effects. For example， one chiral form of the drug naproxen has 28 times the anti-inflammatory activity of the opposite chiral relative.Another example is the synthetic form of dopamine used to treat Parkinson’s disease： one isomer acts on the nerve cells to control the patient’s tremors whilst the other form is actually toxic to nerve cells. In crop protection it has been found that one chiral form of the pesticide Permethrine is much more toxic to insects than the other. Currently there is little control over which chiral form of a chemical compound is formed during a production process. This lack of control generally results in production of equal amounts of both chiral forms. At the molecular level the two chiral partners have identical physical properties and therefore are very difficult and expensive to separate.A far superior alternative to separation would be processes in which single pure chiral compounds are formed in the first place.This application note will describe chiral detection of several over the counter medications using HPLC Experimental A prescription Naproxen tablet (500mg) and an over the counter naproxen sodium tablet (220mg)， were separately ground. The tablets were dissolved in ethanol and sonicated for 15 minutes. These solutions were then diluted to a final concentration of 5ug/uL and filtered through a 0.45um PVDF syringe filter. A Jasco HPLC system with a CD， OR and fluorescence detector was used.Injections size was 1uL. Separations were accomplished using a ChiralPak AD column (4.6 x 250mm) with a 100% EtOH with (0.1% TFA) mobile phase and a flow rate of 1.0 mL/min. CD/UV were monitored at 260nm for the naproxen and 310nm for the Coumadin. Prescription Nexium and Prilosec were also examined by grinding the tablets and dissolving them i n ethanol，a Or n 0d 1 g sonicating for 20 minutes. These solutions were brought to a final concentration Of 1pg/uL with an injection size of 2uL. Separations were accomplished using a ChiralPak AD column (4.6 x 250mm) with a 100% EtOH mobile phase and a flow rate of 0.9mL/min. Results and Discussion Figure 1. Naproxen chromatograms. Top-CD detector，middle- UV detector， and bottom- OR detector. The chromatograms of Naproxen and the naproxen sodium are shown i n Figures 1-2. Naproxen is used to relieve the pain， and inflammation caused by arthritis，and other inflammatory conditions. One chiral form of the drug naproxen (S-enantiomer) has 28 times the anti-inf l ammatory activity of the opposite chiral relative.The (R) isomer i s reported to be a liver toxin. Both the Naproxen and the sodium salt showed only one peak in the CD chromatogram as expected.. This indicates that both the prescription and the over the counter versions have are meeting the requirements to be enantiomerically pure. Figure 2. Naproxen-sodium chromatograms. Top-CD detector，middle - UV detector， and bottom- OR detector. Figure 3. Prilosec chromatograms.Top-CD detector，middle- UV detector， and bottom-OR detector. The chromatograms for the Nexium and the Prilosec are shown in Figures 33-4. Prilosec is S a proton pump inhibitor used to treat ulcers， heartburn， acid reflux， or Zollinger-Ellison syndrome Nexium， (esomeprazole)， is the S enantiomer of omeprazole， the active ingredient in Prilosec. Nexium was one of the first pharmaceuticals to be repatented and marketed as a pure enantiomer of a formerly mixed enantiomer drug. The Nexium aS a superior clinical efficacy due to i ts higher and more consistent bioavailability. As expected， the CD of the prilosec shows the inclusion of both enantiomers while in the Nexium only the esomeprazole enantiomer is detected. Figure 4. Nexium chromatograms. Top- CD detector，middle- UV detector， and bottom-OR detector. Conclusions HPLC with CD detection is an excellent method for the separation and detection of chiral compounds including pharmaceuticals. Both the fluorescence and the CD detectors were very sensitive with respect to concentration； however the fluorescence detector can not give any indication as to which enantiomer is which.The optical rotation detector is capable of discriminating the enantiomers， but in al l the materials studied was found to be much less sensitive. In general the best materials for use with the OR are sugars. Jasco Inc.currently offers the world’s only CD detector for HPLC.