1.Effects of vitamin D on learning and memory and expression of Mcoln-1 in brain tissue of rats with traumatic brain injury
Guangkui HAN ; Yueshu ZHAO ; Cuilian SUN ; Changmeng CUI
Chinese Journal of Behavioral Medicine and Brain Science 2020;29(7):589-593
Objective:To explore the effects of vitamin D (VD) on lysosome activity and calcium channel protein Mcoln-1 in hippocampus and cortex of rats after traumatic brain injury (TBI).Methods:Forty-five SD rats were randomly divided into control group (Sham group), TBI model group (TBI group) and calcitriol treatment group (Calcitriol group). TBI models were established by electronic cortical impactor in TBI group and Calcitriol group. The rats in Calcitriol group were given of calitriol (1 μg/kg) by intraperitoneal injection at 30 min, 24 h and 48 h after TBI.Western blot was used to detect the expression of Mcoln-1, LAMP-1 and cathepsin-B in hippocampus and cortex region of rats. Immunofluorescence was used to detect the co-localization of Mcoln-1 and neurons three days later. Morris water maze test was performed on the 8th, 9th and 10th day after surgery in all groups.Results:Morris water maze results showed that compared with sham group((19.54±3.54)s, (18.64±4.63)s, (17.64±5.88)s), the latency of seeking platform escape ((58.75±6.65)s, (50.64±5.56)s, (42.64±5.87)s) were significantly increased in TBI group ( t=18.042, 14.325, 10.117; all P<0.05). On the 8th, 9th and 10th day after modeling, the escape latency of Calcitriol group((44.54±3.75)s, (30.74±4.74)s, (24.43±4.75)s)were significantly lower than those of TBI group ( t=6.539, 8.909, 7.369, all P<0.05). Western blot results showed that the expression of Mcoln-1 in cortex and hippocampus of TBI group were not significantly different from those of Sham group (both P>0.05). Compared with TBI group, the expression of Mcoln-1 protein in cortex and hippocampus of rats in Calcitriol group were significantly increased ( t=18.862, 17.336, both P<0.05). Immunofluorescence showed that the expression of Mcoln-1 and NeuN (neuron marker) co-located in the cortex and hippocampus of rats in the Calcitriol group. Conclusion:VD can improve learning and memory dysfunction after TBI in rats, and its mechanism may be related to the activation of Mcoln-1 calcium channel.
2.Identification and Content Determination of Main Unknown Impurity in Oxiracetam Capsule
Jing ZHANG ; Ting SUN ; Changmeng ZHAO ; Jianguo JIANG
China Pharmacy 2020;31(6):682-686
OBJECTIVE:To identify the main unknow impurity of Oxiracetam capsule and determine its content ,so as to improve the standard of quality control. METHODS :Two-dimensional UPLC -IT-TOF-MS was adopted to qualitatively analyze the unknown impurity. One-dimensional liquid chromatogram analysis was performed on ST PAK C 18 ES column with mobile phase consisted of 0.02 mol/L sodium dihydrogen phosphate solution at the flow rate of 0.5 mL/min. The column temperature was set at 30 ℃,sample size was 20 μL. The detection wavelength was set at 210 nm. Two-dimensional liquid chromatogram analysis was performed on Techmate C 18-STⅡ column with mobile phase consisted of 0.02 mol/L ammonium acetate solution at the flow rate of 0.5 mL/min. The column temperature was 30 ℃. Mass spectrometry was adopted (electropray ionization source ,MS+ and MS - mode data acquisition ). After the target impurity was located by one-dimensional liquid chromatography ,it was transferred to two-dimensional liquid chromatography-mass spectrometry system for qualitative analysis. The unknown impurity structure was inferred by means of molecular formula prediction module “Accurate Mass Calculator ”in LCMS Solution ,and the refined impurity products by preparation and purification were standardized and confirmed . The impurity content was determined by HPLC (with the same condition of one-dimensional liquid chromatography for qualitative analysis ). RESULTS :The main unknown impurity in Oxiracetam capsules is oxiracetam acid. The content of the refined product was 99.5% after preparation and purification. The contents of oxiracetam acid in 9 batches of Oxiracetam capsules were 0.05% -0.14% . CONCLUSIONS :The established two-dementional UPLC-IT-TOF-MS method can accurately locate the peak position of the impurity oxiracetam acid ,and analyze its structure,while the corresponding content determination method can better separate the impurity from the main drug and other components,with good sensitivity ,precision,repeatability,stability and accuracy. The quality of the finished product of Oxiracetam capsules can be well controlled by using above method .