Quantitative determination of spinosin in rat plasma by liquid chromatography-tandem mass spectrometry method

A sensitive method for the quantitative determination of spinosin in rat plasma was developed and validated using high-performance liquid chromatographic separation with tandem mass spectrometric detection. The analytes of interest were extracted from rat plasma samples by methyl tert-butyl ether (MTBE) after acidification with 1.0% acetic acid aqueous solution. Chromatographic separation was achieved on an Agilent Zorbax SB-C(18) (50 mm x 4.6 mm, 5 microm) using a isocratic mobile phase consisting of acetonitrile-water (30:70, v/v) with 1% isopropyl alcohol and 0.01% heptafluorobutyric acid. The flow rate was 0.2 ml/min. The column temperature was maintained at 25 degrees C. Detection was performed on a triple quadrupole tandem mass spectrometer by selected reaction monitoring (SRM) mode via electrospray ionization (ESI). The calibration curve was linear over the range of 1.00-400 ng/ml in rat plasma, with 1.00 ng/ml of the lower limit of quantification (LLOQ). The inter- and intra-day precisions and accuracy for all samples were satisfactory. The validated method was successfully applied for the pharmacokinetic study of spinosin in rat. After oral administration of 20mg/kg spinosin to rats, the main pharmacokinetic parameters of T(max), C(max), T(0.5) and AUC(0-T) were 5.33+/-0.58 h, 132.2+/-10.6 ng/ml, 4.89+/-0.37 h, 1.02+/-0.09 microg h/l, respectively.     

Spinosin,a C-Glucosylflavone,from Zizyphus jujuba var. spinosa Ameliorates Aβ1–42 Oligomer-Induced Memory Impairment in Mice

Alzheimer’s disease (AD) is a neurodegenerative disorder associated with progressive memory loss and neuronal cell death. Although numerous previous studies have been focused on disease progression or reverse pathological symptoms, therapeutic strategies for AD are limited. Alternatively, the identification of traditional herbal medicines or their active compounds has received much attention. The aims of the present study were to characterize the ameliorating effects of spinosin, a C-glucosylflavone isolated from Zizyphus jujuba var. spinosa, on memory impairment or the pathological changes induced through amyloid-β_1–42 oligomer (AβO) in mice. Memory impairment was induced by intracerebroventricular injection of AβO (50 μM) and spinosin (5, 10, and 20 mg/kg) was administered for 7 days. In the behavioral tasks, the subchronic administration of spinosin (20 mg/kg, p.o.) significantly ameliorated AβO-induced cognitive impairment in the passive avoidance task or the Y-maze task. To identify the effects of spinosin on the pathological changes induced through AβO, immunohistochemistry and Western blot analyses were performed. Spinosin treatment also reduced the number of activated microglia and astrocytes observed after AβO injection. In addition, spinosin rescued the AβO-induced decrease in choline acetyltransferase expression levels. These results suggest that spinosin ameliorated memory impairment induced through AβO, and these effects were regulated, in part, through neuroprotective activity via the anti-inflammatory effects of spinosin. Therefore, spinosin might be a useful agent against the amyloid b protein-induced cognitive dysfunction observed in AD patients.     

LC-MS/MS法测定酸枣仁中酸枣仁皂苷A、B和斯皮诺素的含量

目的建立简单、快速、灵敏同时测定酸枣仁中酸枣仁皂苷A、B和斯皮诺素的LC-MS/MS法。方法采用Phenomenex Gemini C1(850 mm×2.0 mm,5μm)色谱柱;流动相:甲醇-(5 mmol.L-1甲酸铵-0.1%甲酸缓冲液)梯度洗脱;流速为0.25 mL.min-1;柱温为40℃;质谱条件:采用负离子的模式,以MRM配对离子方式进行定量,酸枣仁皂苷A、B和斯皮诺素的选择检测离子质荷比分别为m/z 1251.9→1251.9,1089.9→1089.9和607.3→427.1。结果酸枣仁皂苷A、B和斯皮诺素分别在50～1000 ng.mL-1、25～500 ng.mL-1和12.5～500 ng.mL-1的浓度范围内,线性关系良好;酸枣仁皂苷A、B和斯皮诺素的最低检测限(LOQ)分别为5,2.5和0.5 ng.mL-1;平均回收率为99.35%～101.10%,RSD5.01%(n=9)。三批酸枣仁中酸枣仁皂苷A、B和斯皮诺素的含量分别为1.28～1.38 mg.g-1、1.02～1.11 mg.g-1和1.05～1.24 mg.g-1。结论该法专属性强、快速灵敏,可用于酸枣仁中酸枣仁皂苷A、B和斯皮诺素含量的测定。     

酸枣仁化学成分研究Ⅱ

自酸枣仁(Zizyphus spinosus Hu)种子的甲醇提取部位(F₃)中分离出六个化合物,三个为中性皂甙,酸枣仁皂甙B(Ⅵ),酸枣仁皂甙A(Ⅶ),皂甙Ⅺ(结构待定);一个酚性化合物,阿魏酸(Ⅷ),二个黄酮,spinosin(Ⅸ)和zivulgarin(Ⅹ),其中阿魏酸为首次从酸枣仁中得到,zivulgarin为一新的黄酮类化合物。     

多指标正交试验法优选酸枣仁药材水提取工艺

目的：以浸膏收率、总皂苷含量、斯皮诺素含量为指标,优选酸枣仁药材水提取工艺。方法：采用正交试验,考察煎煮时间、提取次数和加水倍量3个因素,对酸枣仁药材水提取工艺条件进行优化。结果：酸枣仁药材最佳水提取工艺条件为加10倍量水浸泡过夜,加热煎煮3次,每次1h。结论：该提取工艺稳定可行,重现性好,适于酸枣仁药材水提取。     

Spinosin, a C-glycoside flavonoid from semen Zizhiphi Spinozae, potentiated pentobarbital-induced sleep via the serotonergic system

Semen Zizhiphi Spinozae has been used extensively for the treatment of insomnia. This study investigated the effect and possible mechanism of action of spinosin (also known as 2'-beta-o-glucopyranosyl swertisin), a major constituent of semen Zizhiphi Spinozae, on sleep in mice. The present results showed that spinosin significantly and dose-dependently augmented pentobarbital (45 mg/kg, i.p.)-induced sleep, reflected by increased sleep time and reduced sleep latency assessed with the loss-of-righting reflex, and these effects were potentiated by the 5-hydroxytryptamine (serotonin) precursor 5-hydroxytryptophan (5-HTP, 2.5 mg/kg,i.p.). With a subhypnotic dose of pentobarbital (28 mg/kg, i.p.), spinosin significantly increased the rate of sleep onset and exhibited a synergistic effect with 5-HTP (2.5 mg/kg, i.p.). Pretreatment with p-chlorophenylalanine (PCPA, 300 mg/kg, s.c.), an inhibitor of tryptophan hydroxylase, significantly decreased pentobarbital-induced sleep time, and spinosin significantly reversed this effect. The dopamine precursor L-3-(3, 4-dihydroxyphenylalanine (L-DOPA) reduced pentobarbital-induced sleep, an effect not significantly affected by spinosin. These results suggest that spinosin potentiated pentobarbital-induced sleep via a serotonergic mechanism.     

HPLC测定安神健脑液中棘苷的含量

目的:采用HPLC法测定安神健脑液中棘苷的含量。方法:采用Hypersil C₁₈（250 mm×4.6 mm,5μm）色谱柱,以乙腈-0.005 mol·L^-1磷酸二氢钾（12:88）作为流动相,流速为0.8 ml·min^-1,柱温:30℃,检测波长:334 nm。结果:棘苷在4.2～84.0μg·ml^-1范围内有良好的线性关系,平均加样回收率为98.8%,RSD=0.83%。结论:本方法快速简便,重复性好,结果准确,可用于该制剂中棘苷的定量分析。     

HPLC法测定四味安神颗粒中棘苷的含量

目的:采用HPLC法测定四味安神颗粒中尿囊素的含量。方法:采用ZORBAX SB-C18(250mm×4.6mm,5μm)色谱柱,以乙腈-0.005 mol·L-1磷酸二氢钾(13∶87,v/v)作为流动相,流速为0.8mL·min-1,柱温:30℃,检测波长:334nm。结果:棘苷在1.22~24.40μg·mL-1范围内有良好的线性关系,平均加样回收率为98.2%,RSD=1.06%。结论:本方法快速简便,重复性好,结果准确,可用于该制剂中棘苷的定量分析。     

蜜炙酸枣仁中斯皮诺素含量测定

目的:以高效液相色谱法测定蜜炙酸枣仁中斯皮诺素的含量。方法:采用反相高效液相色谱法,以Wonda Cract ODS-2(250 mm×4.6 mm,5μm)为色谱柱,乙腈-水为流动相,检测波长335 nm,流速1 mL/min,柱温30℃。结果:斯皮诺素的线性范围0.005~0.3 mg/mL,r=0.9965,平均回收率为99.74%,RSD=1.46%。结论:该方法准确、重现性好,可为蜜炙酸枣仁的炮制工艺研究及质量控制提供检测标准。     

斯皮诺素在体胃肠吸收动力学研究

目的:研究斯皮诺素在大鼠胃、肠及各肠段的吸收动力学,为其剂型设计提供生物药剂学依据.方法:采用大鼠在体胃肠吸收模型,用高效液相色谱法测定胃肠灌注液中药物的浓度.结果:不同质量浓度(10,20,40 mg·L-1)的斯皮诺素在大鼠胃部2h的吸收百分率分别为20.35％,21.88％,20.23％;药物在十二指肠、空肠、回肠、结肠的吸收速率常数为(0.026 4±0.001 4),(0.023 4±0.0009),(0.022 6±0.0009),(0.026 5±0.0006) h-1;药物质量浓度为10,20,40 mg·L-1时,肠的吸收速率常数分别为(0.023 1±0.001 5),(0.024 1±0.001 6),(0.023 3±0.002 9)h-1;当pH为6.5,7.2,7.8时,肠的吸收速率常数分别为(0.023 5±0.001 5),(0.024 1±0.001 6),(0.022 1 ±0.002 6)h-1.结论:斯皮诺素在大鼠胃肠道各部分均有吸收,且吸收呈一级动力学过程,吸收机制为被动扩散;药物在大鼠肠内吸收不受药物浓度和pH的影响;药物的吸收按十二指肠、结肠、空肠、回肠的顺序依次下降,药物在胃中的吸收较好.