目的 建立以多糖分子量(molecular weight,Mw)及其分布为质量控制指标的黄精和酒黄精配方颗粒质量标准.方法 通过酶解法考察去除辅料的适用性及其最佳反应条件,选取合适的辅料制备黄精和酒黄精配方颗粒,以多糖MW 及其分布情况为指标,采用高效凝胶渗透色谱法(high performance gel permeation chromatography,HPGPC)建立配方颗粒特征图谱.结果高温α-淀粉酶或糖化酶单独使用,均不能将可溶性淀粉和糊精酶解完全,而两种酶协同处理的酶解效果更好.高温α-淀粉酶和糖化酶最佳加酶量分别为辅料:酶用量 1∶1.5 和 1∶2(g/mL).黄精和酒黄精多糖的差异主要在MW>10 kDa部分,经酶法协同处理后,可溶性淀粉在MW>10 kDa部分的糖链已完全水解.各批次黄精配方颗粒以及酒黄精配方颗粒间特征图谱的相似度均大于0.9;且黄精与酒黄精配方颗粒之间的相似度均小于0.5.结论 该方法稳定、可靠,重复性好,能很好地区分黄精和酒黄精配方颗粒,可用于黄精和酒黄精配方颗粒的质量控制.

目的:探讨中药乳香醋炙前后对链脲佐菌素(streptozotocin,STZ)联合高脂饲料诱导2型糖尿病大鼠的改善作用,为扩展中医药的应用和阐明炮制增效机理奠定基础.方法:除正常组外将24只2型糖尿病模型大鼠随机分为模型组、乳香组、醋乳香组及阳性药组,以腹腔注射STZ的方法辅以高脂饲料诱导糖尿病模型.正常组及模型组给予0.5％羧甲基纤维素钠(carboxymethylcellulose sodium,CMC-Na),阳性药组给予盐酸二甲双胍的0.5％CMC-Na混悬液,乳香组和醋乳香组分别给予临床等效剂量乳香及醋乳香的0.5％CMC-Na混悬液进行灌胃给药,共给药2周,给药期间每周进行体质量及空腹血糖的监测.末次给药后,采用ELISA法测定血中胰岛素、糖化血红蛋白(glycosylated hemoglobin,GHb)、一氧化氮(nitric oxide,NO)和内皮素-1(endothelin-1,ET-1)水平并比较胰腺病理切片.结果:与模型组比较,乳香和醋乳香在给药2周后,体质量和胰岛素水平显著提升(P<0.05或P<0.01),而空腹血糖值、GHb、NO及ET-1水平显著降低(P<0.05或P<0.01).胰腺病理切片显示,醋乳香组糖尿病大鼠胰岛萎缩和胰泡细胞变性程度较乳香组降低,且醋乳香的改善作用优于乳香.结论:乳香醋炙后对糖尿病大鼠的改善作用增强.

目的:比较乳香醋炙前后对血瘀模型大鼠的活血作用.方法:采用全血血小板聚集试验进行体外评价,以二磷酸腺苷(adenosine diphosphate,ADP)为诱导剂,2％氢氧化钾为对照组,阿司匹林为阳性药组,电阻法测定乳香和醋乳香供试品溶液对大鼠体外全血血小板聚合值的影响.体内评价采用30只清洁级SD大鼠,除正常组外以注射肾上腺素和冰水浴刺激的方法建立寒凝血瘀动物模型,将24只血瘀模型大鼠随机分为模型组、乳香组、醋乳香组及阳性药组每组各6只.正常组和模型组均给予0.5％羧甲基纤维素钠(carboxymethylcellulose sodium,CMC-Na),阳性药组给予阿司匹林0.5％CMC-Na混悬液,乳香组和醋乳香组分别给予临床等效剂量乳香及醋乳香的0.5％CMC-Na混悬液进行干预,末次给药后检测各组全血黏度、血浆黏度、凝血酶原时间(prothrombin time,PT)、纤维蛋白原(fibrinogen,FIB)、活化部分凝血活酶时间(activated partial thromboplastin time,APTT)、凝血酶时间(thrombin time,TT)、血沉及红细胞压积水平.结果:体外实验结果表明,与对照组比较,醋乳香组和乳香组的全血血小板聚合值和斜率均显著降低(P<0.05,P<0.01),且醋乳香的抑制全血血小板聚集作用优于乳香.体内实验结果表明,与模型组比较,醋乳香组低切变率的全血黏度、血浆黏度、红细胞压积及FIB显著下降,而TT和PT值显著提升(P<0.05,P<0.01);乳香组仅有红细胞压积明显下降(P<0.05),TT值显著上升(P<0.01).结论:乳香和醋乳香均具有抑制全血血小板聚集、改善寒凝血瘀大鼠模型的血液流变学和抗凝血作用,且醋炙后的改善作用更好.

目的 从对初级胆汁酸合成的影响,探讨醋炙增强乳香改善溃疡性结肠炎(ulcerative colitis,UC)的作用机制.方法 基于2,4,6-三硝基苯磺酸(2,4,6-trinitrobenzenesulfonic acid,TNBS)法建立UC大鼠模型,采用超高效液相色谱-质谱联用(UPLC-TQ-MS)法测定大鼠血浆中6种初级胆汁酸含量;进一步借助L-02肝细胞模型,围绕初级胆汁酸合成场所肝脏,探究乳香醋炙前后及醋炙前后差异成分3-乙酰-9,11-去氢-β-乳香酸(3-acetyl-9,11-dehydro-β-boswellic acid,ADHBA)的干预作用,以ELISA法考察胆汁酸前体胆固醇变化,采用qRT-PCR法测定游离型初级胆汁酸合成酶胆固醇7α-羟化酶(cholesterol7α-hydroxylase,CYP7A1)、甾醇12α-羟化酶(sterol12α-hydroxylase,CYP8B1)、甾醇-27-羟化酶(27-hydroxycholesterol,CYP27A1)、氧甾醇-7α-羟化酶(oxysterol-7α-hydroxylase,CYP7B1)、结合型初级胆汁酸合成酶胆汁酰基辅酶A合成酶(bile acyl-CoA synthetase,BACS)和胆汁酸-辅酶A:氨基酸正酰基转移酶(bile acid-CoA:amino acidN-acyltransferase,BAAT) mRNA表达水平.结果 乳香醋炙后,对UC模型大鼠血浆中的初级胆汁酸总量和游离型、结合型初级胆汁酸总量的回调水平更接近正常组.乳香、醋乳香、醋炙前后差异成分ADHBA均可降低胆汁酸合成前体胆固醇水平,但以醋乳香及ADHBA的调控作用更加明显.醋乳香对游离型初级胆汁酸合成经典途径中CYP7A1的抑制和CYP8B1的上调作用以及对结合型初级胆汁酸合成酶BACS、BAAT的上调作用强于乳香组,乳香联合ADHBA组同醋乳香组上调作用一致.结论 乳香醋炙后对初级胆汁酸合成调控作用主要是基于其对游离型初级胆汁酸经典合成途径中限速酶CYP7A1的抑制,回调机体在UC病理状态下的初级胆汁酸水平,进而增强乳香对UC的改善作用;ADHBA可能为乳香醋炙增效的物质基础之一.

目的 比较乳香炮制前后挥发油中主要成分组成及含量的差异,研究不同炮制条件对挥发油成分的影响.方法 采用水蒸气蒸馏法提取挥发油,控制炮制温度和时间得到不同炮制条件醋乳香.利用气相色谱质谱联用技术(gas chromatography-mass spectrometry,GC-MS)分析乳香醋炙前后挥发油,通过质谱数据库检索和对照品对照确定化学成分结构,采用色谱峰峰面积归一化法计算各成分的相对百分含量.结果 乳香与醋乳香的主要成分均为乙酸辛酯、正辛醇、α-蒎烯、柠檬烯,乳香中主要成分的相对含量范围分别为51.92％～79.59％、2.09％～13.50％、0.16％～3.73％、0.10％～1.82％,醋乳香中主要成分的相对含量范围分别为66.72％～80.94％、4.33％～12.2％、0.39％～2.43％、0.25％～4.15％.萜类成分含量随醋炙温度的升高及醋炙时间的延长而增加,醇类及有机酸酯类成分随醋炙温度的升高及醋炙时间的延长而减少,大部分成分随醋炙程度加深而含量增加,而己酸乙酯、α-蒎烯等在乳香中含量较少且炮制后含量减少.结论 乳香挥发油含量较醋乳香高,各挥发油成分含量在炮制后发生改变.

BACKGROUND:Boswellic acids in Olibanum (known as frankincense) are potent anti-inflammatory properties in treating ulcerative colitis (UC), but its low bioavailability limited drug development. Evidence accumulated that vinegar processing of frankincense exerts positive effects on improving absorption of compositions. The underlying mechanism is unknown. In recent decades, spectacular growth and multidisciplinary integration of metabolic application were witnessed. The relationship between drug absorption and curative effect has been more or less established. However, it remains a knowledge gap in the field between drug absorption and endocrine metabolism.PURPOSE:To investigate the enhancement mechanism of vinegar processing in the absorption of boswellic acids via the aspect of bile acid metabolism.METHODS:The effects of raw frankincense (RF) and processed frankincense (PF) were compared by the UC model of rats. The plasma concentration of boswellic acids and the hepatic and colonic bile acids contents were quantified by UPLC-TQ-MS. The levels of mRNA and protein associated with bile acid metabolism were also compared.RESULTS:The results showed that PF exhibited re-markable mitigating effects on UC with the elevated plasma level of boswellic acid and upregulated expression of the absorption-related protein multidrug resistance-associated protein 2 (MRP2) and organic anion transporting polypeptide 1B3 (OATP1B3) in the liver and colon. It improved colonic lithocholic acid (LCA), which promoted the expression of bile acid nuclear receptors constitutive androstane receptor (CAR) and pregnane X receptor (PXR), resulting in the upregulation of MRP2 and OATP1B3.CONCLUSION:This paper revealed the mechanisms behind the absorption promotion effects of processing. Bile acids metabolism exhibits potential status in pharmaceutical development. The results shed light on the interdisciplinary collaboration between the metabolism and drug absorption fields.

目的 通过对比乳香醋炙前后11-羰基-β-乳香酸(11-keto-boswellic,KBA)、3-乙酰-11-羰基-β-乳香酸(3-acetyl-11-keto-boswellicacid,AKBA)、榄香醇酸、β-榄香酮酸、tsugaricacid A、9,11-去氢-α-乳香酸、9,11-去氢-β-乳香酸、α-乳香酸、β-乳香酸、3-乙酰-9,11-去氢-α-乳香酸、3-乙酰-9,11-去氢-β-乳香酸、3-乙酰α-乳香酸(3-acetyl-α-boswellic acid,α-ABA)和3-乙酰β-乳香酸(3-acetyl-β-boswellic acid,β-ABA)13个乳香酸成分含量变化,探究不同炮制条件对含量变化的影响,考察差异乳香酸成分抗炎活性,初步揭示乳香醋炙增效机制.方法 制备不同炮制温度和炮制时间醋乳香,基于超高效液相色谱仪串联三重四级杆质谱仪(UPLC-TQ-MS)建立13种乳香酸成分含量测定方法.采用AcquityUPLCBEHC18色谱柱(100 mm×2.1mm,1.7μm),以0.1％甲酸+5mmol/L乙酸铵水溶液-0.1％甲酸乙腈溶液为流动相,梯度洗脱,体积流量0.3mL/min,柱温40℃,质谱采用电喷雾负离子源,多反应监测(multi-reaction monitoring,MRM)模式;建立脂多糖诱导巨噬细胞的炎症细胞模型,通过ELISA法测定乳香、醋乳香、单体(3-乙酰-9,11-去氢-β-乳香酸)和乳香+单体(3-乙酰-9,11-去氢-β-乳香酸)对炎症细胞因子肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interleukin-1 β,IL-iβ)以及IL-6的影响.结果 随醋炙温度升高或炮制时间的延长,乳香中榄香醇酸、β-榄香酮酸、tsugaric acid A、9,11-去氢-α-乳香酸、9,11-去氢-β-乳香酸、3-乙酰-9,11-去氢-α-乳香酸、3-乙酰-9,11-去氢-β-乳香酸、α-ABA和β-ABA 9个乳香酸含量升高幅度增大,而KBA、AKBA、α-乳香酸、β-乳香酸4个成分含量降低幅度增大,其中3-乙酰-9,11-去氢-β-乳香酸含量变化幅度最大.炮制温度引起的成分变化强于炮制时间的影响.乳香酸类成分含量升高或降低与成分结构有关;乳香醋炙后抑制炎症因子分泌的作用显著增强,3-乙酰-9,11-去氢-β-乳香酸显示出明显的抑制炎症因子分泌活性.结论 温度是影响乳香中乳香酸成分变化的重要因素,含量升高或降低与结构有关.3-乙酰-9,11-去氢-β-乳香酸含量变化最大,该成分具有显著的抗炎活性,可能为乳香醋炙增效的物质基础之一.

The presence of hypertension (HTN) in type 2 diabetes mellitus (DM) is a common phenomenon in more than half of the diabetic patients. Since HTN constitutes a predictor of vascular complications and cardiovascular disease in type 2 DM patients, it is of significance to understand the molecular and cellular mechanisms of type 2 DM binding to HTN. This review attempts to understand the mechanism via the perspective of the metabolites. It reviewed the metabolic perturbations, the biological function of perturbated metabolites in two diseases, and the mechanism underlying metabolic perturbation that contributed to the connection of type 2 DM and HTN. DM-associated metabolic perturbations may be involved in the pathogenesis of HTN potentially in insulin, angiotensin II, sympathetic nervous system, and the energy reprogramming to address how perturbated metabolites in type 2 DM affect the pathogenesis of HTN. The recent integration of the metabolism field with microbiology and immunology may provide a wider perspective. Metabolism affects immune function and supports immune cell differentiation by the switch of energy. The diverse metabolites produced by bacteria modified the biological process in the inflammatory response of chronic metabolic diseases either. The rapidly evolving metabolomics has enabled to have a better understanding of the process of diseases, which is an important tool for providing some insight into the investigation of diseases mechanism. Metabolites served as direct modulators of biological processes were believed to assess the pathological mechanisms involved in diseases.

开展中医复方药效物质基础研究,对揭示方药配伍规律具有重大意义.而复方中种类繁多、含量悬殊的化学成分使得研究极具挑战,因此采用必要的分离手段至关重要.高速逆流色谱(high-speed counter-current chromatography,HSCCC)具有分离速度快、成分不变性且无损失的特点,为单体成分的分离、化学组分的分析以及活性成分的发现提供了有力的技术保障.本文针对近些年HSCCC技术在溶剂体系、洗脱方式、检测技术以及其他提取技术联合应用的相关报道进行综述,同时归纳了HSCCC技术在中药及天然产物化学成分研究中的应用实例,以期为中医复方药效物质基础研究提供参考.虽然HSCCC的广泛应用仍然受到制约,但相信随着中药及天然产物物质基础研究的逐步深入,HSCCC将为中医复方药效物质基础研究提供更加完备的技术支撑.

目的:研究黄精炮制前后多糖相对分子质量及其分布变化,以及其不同多糖级分对小鼠腹腔巨噬细胞免疫功能和炎症反应的影响.方法:采用高效凝胶渗透色谱法(HPGPC)测定黄精多糖(SC)和酒黄精多糖(JC)相对分子质量及其分布情况,并通过透析法得到不同相对分子质量多糖级分,将上述多糖级分分别作用于正常和经脂多糖(LPS)诱导的小鼠腹腔巨噬细胞,采用细胞增殖与活性检测试剂盒-8(CCK-8)法选取最佳给药浓度,酶联免疫吸附测定法(ELISA)测定细胞上清液中肿瘤坏死因子(TNF)-α,白细胞介素(IL)-1β的浓度,格里斯(Griess)试剂法检测一氧化氮(NO)的浓度.结果:SC和JC按相对分子质量及其分布范围可分为4个部分,第Ⅰ部分(14800～2273 kDa),第Ⅱ部分(2148～296 kDa),第Ⅲ部分(12～1 kDa),第Ⅳ部分(818～362 Da).而黄精经酒制后,第Ⅰ部分和第Ⅲ部分的多糖级分发生了改变,因此,选择将SC和JC按重均相对分子质量(Mw)＞50 kDa和＜50 kDa各分成2个部分.对于正常小鼠腹腔巨噬细胞,JC能显著促进细胞分泌TNF-α(P＜0.01),但SC无明显作用;4个多糖级分SD(Mw＞50 kDa的SC级分),JD(Mw＞50 kDa的JC级分),SX(Mw＜50 kDa的SC级分),JX(Mw＜50 kDa的JC级分)均能显著促进细胞分泌TNF-α(P＜0.01),但仅JX能明显促进细胞分泌NO (P＜0.05);另外,JX组促进细胞分泌TNF-α作用显著强于JD组(P＜0.01).对于LPS诱导巨噬细胞模型,JC和SC均能显著抑制细胞分泌TNF-α和IL-1β(P＜0.01),且JC作用更强;不同多糖级分间,JX抑制细胞分泌TNF-α和IL-1β的作用显著强于JD(P＜0.01),SX抑制细胞分泌TNF-α作用显著强于SD(P＜0.01).结论:黄精酒制前后多糖相对分子质量及其分布发生了改变,JC和SC增强免疫调节作用主要是通过抑制炎症反应,Mw＜50 kDa级分是其主要有效部位,且黄精在酒制后其多糖成分抑制炎症反应的作用增强.

中药质量影响中医复方功效的发挥,而中药有效成分含量的高低是评价中药质量优劣的重要指标.多糖作为中药的有效成分之一,由于其结构和组成极为复杂,其含量测定一直是难点问题.《中华人民共和国药典》收载的中药多糖含量测定方法基本采用紫外-可见分光光度法(UV-Visible spectrophotometer,UV-Vis)存在一些问题,方法的准确性亟待提高.本文通过系统查阅国内外文献,归纳了目前测定中药多糖含量的常用方法,主要分为多糖的直接测定和多糖水解后单糖的测定.前者主要采用UV-Vis法、高效凝胶渗透色谱法(high performance gel permeation chromatography,HPGPC)、近红外光谱法(near-infrared reflectance,NIR)和碘量法,后者主要采用高效液相色谱法(high performance liquid chromatography,HPLC)、气相色谱法(gas chromatography,GC)、离子色谱法(ion chromatography,IC).本文从对照品的选择、供试品溶液制备方法等方面对《中华人民共和国药典》收载的中药多糖含量测定方法进行了总结和讨论,通过归纳分析为中药多糖质量评价提供参考.

中医复方疗效与中药质量密切相关,采用科学方法评价中药质量十分必要.定性鉴别与指纹图谱是评价中药质量的主要指标.中药多糖多为不同分子量分布范围的混合物,其生物活性与其单糖组成、分子量大小与分布等结构因素密切相关.目前《中华人民共和国药典》仅采用理化鉴别法对中药中的多糖进行定性鉴别,未收载中药多糖的糖谱分析方法,无法体现各中药多糖的真实状态.本文通过系统查阅文献,归纳国内外中药多糖定性鉴别与糖谱的最新研究内容,主要从多糖分子量分布和多糖的单糖组成两方面开展研究.其中,依据多糖分子量分布的定性鉴别主要采用高效凝胶渗透色谱法(high performance gel permeation chromatography,HPGPC)、质谱法(mass spectrum,MS)和黏度法,依据多糖单糖组成的定性鉴别主要采用气相色谱法(gas chromatography,GC)、高效液相色谱法(high performance liquid chromatography,HPLC)、离子色谱法(ion chromatography,IC)、薄层色谱法(thin layer chromatography,TLC)和高效凝胶电泳法(high performance capillary electrophoresis,HPCE).糖谱是通过多批次样品分析而获取的共性信息建立的图谱,依据多糖整体结构特征的糖谱建立主要采用HPGPC法、红外光谱法(infrared ray,IR)和近红外光谱法(near infrared ray,NIR),依据多糖单糖组成的糖谱的建立主要采用HPLC法,通过归纳总结分析以期为科学合理地评价中药多糖质量,从而保障中医临床疗效提供依据.

经典名方作为中医药宝库的精华部分,其研发已成为中医药行业的热点,随着年代更迭,中药基原和产地均存在或多或少的变化,某些中药还存在代用品或易混品.因此,明确中药的基原和产地,对经典名方的安全性和有效性具有重要意义.本文在探讨药味考证的方法的基础上,从基原、产地、代用品和易混品4个方面,对国家中医药管理局发布的《古代经典名方目录(第一批)》中100首经典名方涉及药味进行分析.本草考证结果显示经典名方涉及的药味中多数存在基原和产地变迁问题,因此,全面考证经典名方涉及药味对经典名方研发过程中药味基原和产地的选取具有重要意义.

Boswellic acids (BAs), as the main components of frankincense, exhibit notable anti-inflammatory properties. However, their pharmaceutical development has been severely limited by their poor oral bioavailability. Traditional Chinese medicinal processing, called Pao Zhi, is believed to improve bioavailability, yet the mechanism is still completely unclear. Previous research suggested that the bioavailability of a drug can be influenced by physical properties. This paper was designed to investigate the physical properties of frankincense and processed frankincense, including the surface morphology, particle size, polydispersity index (PDI), zeta potential (ZP), specific surface area, porosity, and viscosity. The differences in the intestinal absorption characteristics and equilibrium solubilities between frankincense and processed frankincense were determined by an ultra-high-performance liquid chromatography coupled with a triple quadrupole electrospray tandem mass spectrometry (UHPLC-TQ-MS) analysis method. The results showed that vinegar processing can alter the surface morphology, decrease the particle size and PDI, raise the absolute values of the ZP, specific surface area and porosity, and drop the viscosity of frankincense. Meanwhile, the rates of absorption and dissolution of the main BAs were increased after the processing of frankincense. The present study proves that the physical properties were changed after processing, in which case the bioavailability of frankincense was enhanced.