Determination Of Three Kinds Of Primary Amine Bisphosphonates Drugs By HPLC And Determination Of Four Kinds Of Anthraquinone In Rhubarb By GC-MS

PART I DETERMINATION OF THREE KINDS OF PRIMARY AMINE BISPHOSPHONATES DRUGS BY HPLCBisphosphonates are a class of highly effective inhibitors of bone resorption, clinical courses for a variety of bone related diseases (such as osteoporosis, paget disease and malignant hypercalcemia etc.). Most bisphosphonates have low bioavailability after oral administration. However, they are difficult to be separated and enriched because of their strong polarity. Furthermore, they are hardly maitained in reversed phase chromatographic column. And most bisphosphonates can not be detected directly by HPLC-UV or fluorescence because they have no chromophore group.Alendronate, pamidronate and neridronic acid belong to primary amines of bisphosphonates.They have been widely used in the treatment of osteoporosis, bone metabolism disorders in the elderly, and the various osseous lesion associated with tumor in clinic.Therefore the establishment of rapid detection method for these three medicines has important theoretical and practical significance. An HPLC method has been developed for three bisphosphonates after pre-column derivatization with FMOC-C1 in this study.Objective:To establish an HPLC method for the determination of alendronate sodium tablets, disodium pamidronate injection and neridronic acid raw material medicine by the FMOC-C1 as the pre-column derivatization reagent. Methods:The analytical column was LinkSi C18 (250×4.6mm,5.0um).The mobile phase consisted of methanol-acetonitrile-0.4%EDTA with pH 7.16 (5:25:70).The flow rate is at 1mL· min-1.The detection wavelength was 264 nm. Results:Three drugs showed good linearity(r>0.9995) over the concentration range of 1.0-40 ugmL-1.The regression and LOD of the three drugs are as following:alendronate, y=0.941x-0.697,LOD=0.2ugmL-1;pamidronate,y=0.447x-0.240,LOD=0.5u gmL-1;neridronic acid,y=0.730x-0.316,LOD=0.1ugmL-1. Conclusion: Determination of three kinds of bisphosphonate by the HPLC-UV method is accurate, simple, rapid, qualitative.It can be used to determine the content of the sample and preparation.PART II DETERMINATION OF FOUR KINDS OF ANTHRAQUINONE IN RHUBARB BY GC-MSRhubarb as a Chinese traditional medicine,has many pharmacological effects. In particular,it has a good effect on constipation, stagnation of food, abdominal pain, acute appendicitis, acute infectious hepatitis, acute conjunctivitis and other diseases. Modern pharmacological experiments showed that anthraquinone compounds contained in rhubarb have a broad spectrum of biological activity, and they are also as a major component of rhubarb effect.There is existing some literature on the chemical constituents of rhubarb. Its main activity constituents are these free anthraquinone as aloe emodin, rhein, emodin, chrysophanol and physcion. Now,the determination of the anthraquinone compounds in rhubarb mainly implemented by HPLC. In this paper, we use GC-MS to detect the content of these anthraquinones by the selected ion mode which has good specificity and high sensitivity.It provides a basis for the analysis of this kind of compounds in biological samples. At the same time, analyse the main pyrolysis approach of four analytes by EI-MS.It provided a basis for further study on the structure modification of the four kinds of compounds and for the development of the drugs.Objective:To develop a method of GC-MS for the determination of chrysophanol, physcion, aloe-emodin and emodin in rhubarb. Analyse the fragmentation pathways of the four kinds of anthraquinones in Rhubarb by EI-MS. Method:The analytes was analyzed quantitatively on the TG-5MS quartz capillary chromatographic column by selective ion monitoring.The sample was extracted by ultrasound in acetic ether using osthole as an internal standard. Results:There was a good linear (r>0.9998) relationship in the range of 3.2-30μgmL-1 in four compounds.The average recovery were 98.82%,97.90%,102.94%and 96.89%and the detection limit were 0.1ngmL-1,0.5 ng·mL-1,10ng·mL-1 and 20 ng·mL-1. Conclusion:The method is convenient and rapid, it can be applied for determination of chrysophanol, physcion, aloe-emodin and emodin in rhubarb.