Non‐targeted analyses of animal plasma: betaine and choline represent the nutritional and metabolic status

Simple liquid chromatography–mass spectrometry (LC‐MS) was applied to non‐targeted metabolic analyses to discover new metabolic markers in animal plasma. Principle component analysis (PCA) and partial least squares–discriminate analysis (PLS‐DA) were used to analyse LC‐MS multivariate data. PCA clearly generated two separate clusters for artificially induced diabetic mice and healthy control mice. PLS‐DA of time‐course changes in plasma metabolites of chicks after feeding generated three clusters (pre‐ and immediately after feeding, 0.5–3 h after feeding and 4 h after feeding). Two separate clusters were also generated for plasma metabolites of pregnant Angus heifers with differing live‐weight change profiles (gaining or losing). The accompanying PLS‐DA loading plot detailed the metabolites that contribute the most to the cluster separation. In each case, the same highly hydrophilic metabolite was strongly correlated to the group separation. The metabolite was identified as betaine by LC‐MS/MS. This result indicates that betaine and its metabolic precursor, choline, may be useful biomarkers to evaluate the nutritional and metabolic status of animals.     

基于LCMS技术对牦牛围产后期的血清动态代谢组学分析

旨在应用代谢组学的方法,阐明经产母牦牛围产后期动态代谢情况,探究营养代谢影响生殖激素的机制。本研究随机选取年龄（（7.13±0.78）岁）、体况（2.69±0.35）和胎次（（1.75±0.43）胎）相近的经产母牦牛8头作为试验动物,自分娩日开始每隔7 d收集血清至产后第28天,共5个时间点。应用液相质谱联用技术对5个时间点的血清样本进行检测,并通过生物信息学分析筛选关键代谢物,构建经产母牦牛围产后期动态代谢网络机制。试验在正、负离子模式下分别获得2 841和1 326种代谢物定性分析的结果,通过计算各时间点代谢物表达量的差异倍数、VIP值及每两个时间点T-test检验的P值,共筛选出117种差异代谢物。根据生物信息学分析,分别获得与繁殖性能、脂类代谢和氨基酸代谢相关的3种、7种和8种关键代谢物。基于关键代谢物构建动态代谢网络机制发现,糖代谢在围产后期的起始阶段作用效率较低,末期升高,脂类和氨基酸代谢在起始阶段作用效率较高,末期降低,推测经产母牦牛在产后发生能量负平衡。生殖激素的合成和分泌作用效率在围产后期一直维持较低水平。根据对经产母牦牛动态代谢谱的检测和网络机制的构建发现,经产母牦牛产后发生了能量负平衡,由于过度脂肪动员导致雌激素的合成和分泌受阻,这可能是导致经产母牦牛产后生殖机能不能及时恢复的主要原因,但具体机制还需要进一步验证。     

中兽药口服液中隐性药物筛查高分辨质谱方法的建立

建立了测定中兽药口服液中170种隐性药物的超高效液相色谱—四极杆静电场轨道阱质谱( HPLC- Q-Exactive Focus/MS) 高通量筛查方法。通过构建药物筛查数据库并与样品中未知物信息进行比对,从而实现快速筛查中兽药口服液中隐性药物的种类和含量。数据库包括磺胺类、喹诺酮类、β-受体激动剂类、硝基咪唑类、苯并咪唑类、蛋白同化激素类、糖皮质激素类、喹噁啉类、四环素类、抗病毒类、酰胺醇类和真菌毒素类等12类170种药物的筛查参数。样品中人为添加化学药物,经1%甲酸乙腈提取,QuEChERs净化,氮气吹干,0.2%甲酸水复溶上机,正负模式扫描,TraceFinder软件筛查定性,当药物添加浓度为400ng/g时,170种添加药物的检出率约为92.8%,随着药物浓度逐步提高至1 μg/g时,所有添加药物均可检出。结果表明,该方法可满足中兽药口服液中隐性药物高通量快速筛查的需要,为打击畜禽养殖投入品中非法使用化学药物的行为提高技术支持和数据支撑。     

高分辨四极杆飞行时间质谱仪判别复原乳和超高温灭菌乳

采用高分辨四极杆飞行时间质谱仪,结合化学计量学方法,利用非靶向代谢组学方法,对超高温灭菌乳和复原乳进行检测。牛乳样品经过前处理后,经过C18色谱柱分离,采用FullScan模式进行一级全扫描,扫描结果通过数据预处理后,导入SIMCA-P14.1软件中进行主成分分析和偏最小二乘方差判别分析。结果表明:正、负离子模式下共找到14种判别超高温灭菌乳和复原乳的表征因子,进一步通过这14种表征因子建立区分2种乳的判别模型,通过该判别模型能够准确地区分超高温灭菌乳和复原乳,为复原乳的判别提供理论依据。     

Energetic Requirements for Growth and Maintenance of the Cape Gannet (Aves; Sulidae)

Energetic requirements for growth and maintenance of the Cape gannet (Sula capensis) were studied by hand-rearing captive chicks and keeping juveniles in captivity at constant mass. Daily gain in mass was linear until 60 days of age; after 82 days the chicks lost mass prior to attaining fledging age (97 days). Daily relative growth decreased with age. Food intake increased to 70 days and thereafter decreased. Guano production increased to SO days. Relative food intake decreased with age. Energy content of guano and assimilation efficiency showed no clear changes with age. Mass recession prior to fledging may be related to the lack of post-parental care. Hand-reared gannets grew at an optimum rate when compared with field growth rates from the literature. Juvenile birds were maintained on a diet forming 12,4% of body mass at an assimilation efficiency of 74,2%. Aviary-existence maintenance was 2,4 times standard metabolic rate. The cost of free living in an adult gannet is roughly estimated to be 4,5 X SMR, corresponding to a food intake of 20% of body mass.     

β-受体激动剂质谱裂解特征研究

以克伦特罗、莱克多巴胺、沙丁胺醇为代表,阐明了β-受体激动剂在正离子软电离模式下的裂解机理。采用AB SCIEX 5500 QTRAP三重四级杆电喷雾质谱仪,运用IDA-EPI技术,对β-受体激动剂类化合物裂解规律进行研究解析。结果表明,在ESI正电离模式下β-受体激动剂容易获得H+而形成[M+H]+准分子离子峰,该结构易丢失水分子(H2O),继而与氨基结构相连的碳氮键(-C-N-)发生断裂,形成与苯环共轭的(Ph-CH=CH-NH+-)离子结构。本研究为同类化合物的结构解析提供了可靠依据以及跟踪新型β-受体激动剂提供了有力手段。     

Oxygen consumption, diffusing capacity and blood flow of the synovial membrane in osteoarthritic rabbit knee joints

In osteoarthritis the changes of the synovial membrane may seriously alter the oxygen transfer characteristics from the capillaries of the membrane to the synovial fluid and hence impede or deprive the joint cartilage of its sole source of oxygen. In the present study we have estimated the blood flow (Q), diffusing capacity (DO2) and oxygen consumption (VO2) of the synovial membrane in the chronic non effusive stage of experimental osteoarthritis. In 14 osteoarthritic knee joints we found a statistically significant increase in oxygen consumption, compared to previously reported results from normal joints, whereas the diffusing capacity and the blood flow were unchanged. The implication of this is that the partial pressure difference required to overcome the increased oxygen consumption was increased with a factor 4.3 and that the oxygenation of the joint cartilage was reduced with approx. 30 Torr.     

超高效液相色谱-串联质谱法测定鸡蛋中氯霉素的两种前处理方法比较

为选择氯霉素超高效液相色谱串联质谱分析前处理方法,分别对乙酸乙酯提取,正己烷液液萃取净化和乙腈提取,LC-Si固相萃取柱萃取净化(GB/T22338-2008方法),两种前处理方法进行对比分析。结果显示:乙酸乙酯提取法的检测限为0.1μg/kg,在0.5~10ng/mL范围内线性关系良好,r>0.99,回收率为98.2%~104.8%,RSD为1.5%~7.2%;乙腈提取法的检测限为0.05μg/kg,在0.25~10ng/mL范围内线性关系良好,r>0.995,回收率为100.0%~104.0%%,RSD值为0.5%~2.1%。结果表明:两种方法的线性范围、回收率、检测限和相对偏差均能满足畜产品检验要求。乙酸乙酯提取法步骤简单,样品处理效率高,乙腈提取法虽步骤繁琐,但结果的精密度和灵敏度更高。     

液相色谱-串联质谱测定饲料中7种硝基咪唑类药物

采用液相色谱-串联质谱法(LC-MS/MS)测定了饲料中甲硝唑(MNZ)、地美硝唑(DMZ)、洛硝唑(RNZ)、替硝唑(TNZ)、奥硝唑(ONZ)、异丙硝唑(IPZ)和塞克硝唑(SCZ)等7种硝基咪唑类药物。样品用乙酸乙酯提取,正己烷-磷酸溶液反萃取,MCX柱净化,以0.1%乙酸水溶液和乙腈作流动相进行梯度洗脱,电喷雾电离源正离子(ESI+)多反应监测(MRM)模式下检测。方法的检测限和定量限分别为0.03 mg/kg和0.1 mg/kg。添加浓度为0.1、1 mg/kg和10 mg/kg时的平均回收率为72.9%~98.8%,变异系数为4.9%~12.1%。     

高效液相色谱串联质谱法测定渔业水体及渔业饲料中的五氯苯酚

本文建立了一种简单、快速测定渔业水体和渔业饲料中五氯苯酚的液相色谱串联质谱分析方法。渔业水体采取固相萃取进行富集净化,渔业饲料样品经氨化乙腈提取,MAX固相萃取小柱净化,质谱采用选择反应监测负离子模式测定进行分析。结果表明:方法线性为0.5～50 ng/mL,相关系数大于0.9999;渔业水体的检出限(LOD)和定量限(LOQ)为0.05μg/L和0.1μg/L,渔业饲料的LOD和LOQ为0.8μg/kg和1.6μg/kg,该方法在水样和渔业饲料样品中的平均加标回收率为78.6%～92.1%。采用该方法对长沙市周边的湖泊和池塘地表水和渔业饲料样品进行检测,在水样中测得五氯苯酚浓度为0.64～1.22μg/L。