The establishment and validation of efficient assays for anti-IIa and anti-Xa activities of heparin sodium and heparin calcium.

Heparin is used as an anticoagulant drug. The anticoagulation process is mainly caused by the interaction of heparin with antithrombin followed by inhibition of anticoagulant factor IIa and factor Xa. The anti-factor IIa and anti-factor Xa activities of heparin are critical for its anticoagulant effect; however, physicochemical methods that can reflect these activities have not been established. Thus, the measurements of anti-IIa and anti-Xa activities by biological assay are critical for the quality control of heparin products. Currently in the Japanese Pharmacopoeia (JP), the activities of heparin sodium and heparin calcium are measured by an anti-Xa activity assay (anti-Xa assay), but anti-IIa activity is not measured. Here, we established an anti-IIa activity assay (anti-IIa assay) and an anti-Xa assay having good accuracy and precision. When samples having a relative activity of 0.8, 1.0 and 1.2 were measured by the established anti-IIa and anti-Xa assays in nine laboratories, good accuracy (100.0-102.8% and 101.6-102.8%, respectively), good intermediate precision (1.9-2.1% and 2.4-4.2%, respectively) and good reproducibility (4.0-4.8% and 3.6-6.4%, respectively) were obtained. The established anti-IIa and anti-Xa assays have similar protocols, and could be performed by a single person without a special machine. The established assays would be useful for quality control of heparin.

Coronary abnormal response has increased in Japanese patients: analysis of 17 years' spasm provocation tests in 2093 cases.

BACKGROUND: Abnormal coronary response on acetylcholine test is observed in patients with early coronary atherosclerosis. OBJECTIVES: We analyzed retrospectively the abnormal response rate during 17 years of spasm provocation tests in 2093 consecutive patients. METHODS: We performed 2093 spasm provocation tests, consisting of 1198 acetylcholine tests and 895 ergonovine tests, between January 1991 and December 2007. Spasm provocation test was mainly performed in patients with ischemic heart disease. Abnormal response was defined as transient >90% luminal narrowing during spasm provocation tests. We classified these 17 years into two periods: former period from January 1991 to December 2000, and the latter period from January 2001 to December 2007. In the former period, 1300 spasm provocation tests were performed and 793 spasm provocation tests were done in the latter period. RESULTS: The incidences of hypertension, dyslipidemia, and diabetes mellitus were significantly increased in the latter period. The values of total cholesterol, triglycerides, and fasting blood sugar were also significantly increased in the latter period. The frequency of abnormal response in the latter period was significantly higher than that in the former period (46.0% vs. 33.2%, p<0.05). The frequency of abnormal coronary response to acetylcholine in the latter period was significantly higher than that in the former period (60.0% vs. 34.0%, p<0.01), whereas there was no difference concerning abnormal response of ergonovine between the two periods (31.9% vs. 30.7%, ns). CONCLUSIONS: In Japanese patients, abnormal coronary response to acetylcholine has increased and coronary endothelial dysfunction is suggested to have progressed.