食品中棉酚的检测方法研究进展

棉酚以游离态和结合态的形式存在, 两者之和称为总棉酚。游离棉酚对人和动物具有急性毒性、肝毒性和生殖毒性。最近关于棉酚的功能研究也再次使消费者开始关注其健康作用和危害。鉴于棉酚可能存在的危害和功能, 建立可靠的食品中棉酚的检测方法显得尤为必要。本文简要综述了食品中棉酚的提取方法以及比色法、薄层色谱法、分光光度计法、高效液相色谱法、高效液相色谱-质谱联用法、近红外光谱法和酶联免疫吸附法等检测方法的最新进展和优缺点, 以期促进棉酚检测方法的研究和开发。     

棉油皂脚制备醋酸棉酚的研究

对棉油皂脚制备醋酸棉酚的工艺进行了研究。以棉油皂脚为原料,经酸化、水解、分液、结晶、过滤、干燥即可得到醋酸棉酚。通过单因素试验优化醋酸棉酚的制备工艺。结果表明:以棉油皂脚为原料,在浓硫酸添加量10%、加热时间90 min、冰醋酸与油相体积比1∶3条件下,一次结晶可得到纯度88%以上的醋酸棉酚,回收率大于60%;经过一次重结晶可得到纯度98%以上的醋酸棉酚,回收率大于45%。     

Effects of source of gossypol and supplemental iron on plasma gossypol in Holstein steers

Four experiments were conducted to evaluate factors influencing concentrations of plasma total gossypol (TG) in 30 Holstein steers fed cottonseed products. At the end of each 28-d experiment, steers were weighed and blood samples were collected and analyzed for plasma TG concentrations. During the entire study, steers did not show any overt signs of gossypol toxicity. In the 28 d before experiment 1, 30 steers with a body weight (BW) of 273 kg were fed a standardization diet with 15.0% Upland whole cottonseed (WCS) that resulted in a mean intake of 9.08 g/d of TG per steer/d and a plasma TG of 1.66 μg/mL. In experiment 1, 30 steers were fed 1 of 5 diets with 15.0% Upland WCS, but different levels of supplemental Fe [0, 150, 300, 450, and 600 mg/kg of diet dry matter (DM)]. Average daily gain was not affected by level of Fe in the diet, but DM intake, plasma TG, and plasma TG response decreased linearly as Fe in diets increased. In experiment 2, steers were fed diets with 15.0% Upland cottonseed as whole, cracked, roasted, cracked-roasted, or extruded. Analysis of the seed revealed that roasting or extrusion markedly reduced free gossypol (FG) content. Minor effects on animal performance were observed, but plasma TG decreased with roasting or extrusion of seeds, with the greatest reduction when the seed was cracked and then roasted. In experiment 3, steers were fed 2 levels of WCS (7.0 or 14.0% of DM) with 3 levels of cottonseed meal (2.8, 5.5, or 8.5% of DM) in the diet. Animal performance was not altered by diet, but plasma gossypol concentrations and responses were greater in steers fed diets with more WCS, because of the greater FG intake. In experiment 4, 24 steers were fed diets with 15.0% cottonseed (Upland or Pima) either as whole or cracked. Pima cottonseed increased TG and FG intakes, which resulted in greater plasma TG concentration and response. Animal response to processing of cottonseed tended to differ according to type of cottonseed. However, feeding Pima and cracking of cottonseed increased gossypol availability and plasma TG concentrations.     

Developmental competence of bovine embryos from heat-stressed ova

Because multiple ovulation embryo transfer procedures are occasionally performed in cows experiencing heat stress, the goal of this study was to assess the developmental competence of otherwise morphologically normal embryos from heat-stressed ova. To this end, the ability of compact morulae from heat-stressed and non-heat-stressed bovine ova to undergo blastocyst development after culture at 38.5 or 41.0°C was examined. It was hypothesized that heat-induced perturbations in the ooplasm carry over to increase the susceptibility of the preattachment embryo to heat stress. Initially, ova were matured at 38.5 or 41.0°C. The consequences of heat stress did not include altered cleavage, but did reduce the proportion of 8- to 16-cell-stage embryos (55.3 vs. 50.6%; SEM ± 1.9). Although proportionately fewer, compact morulae from heat-stressed ova were equivalent in quality to those from non-heat-stressed ova (2.1 and 2.1; SEM = 0.04). Culture of compact morulae from non-heat-stressed ova at 41.0°C did not affect blastocyst development (71.9 and 71.5%; SEM = 3.0). Furthermore, the development of compact morulae from heat-stressed ova was similar to that of non-heat-stressed ova after culture at 38.5°C (68.2 vs. 71.9 and 71.5%; SEM = 3.0). However, blastocyst development was reduced when compact morulae from heat-stressed ova were cultured at 41.0°C (62.3 vs. 71.9, 71.5 and 68.2; SEM = 3.1). In summary, reduced compaction rates of heat-stressed ova explained in part why fewer develop to the blastocyst stage after fertilization. The thermolability of the few embryos that develop from otherwise developmentally challenged ova emphasizes the importance of minimizing exposure to stressor(s) during oocyte maturation.     

棉酚在双液相溶剂中的分配系数

研究了双液相溶剂(甲醇相 石油醚)浸取破碎棉仁的工艺特性,测定了达到浸取平衡的时间,考察了温度、甲醇相浓度等因素对游离棉酚分配系数的影响.结果表明,甲醇相中加入一定量的柠檬酸可以提高游离棉酚在甲醇相中的分配系数,而且温度越高,甲醇相浓度越大,分配系数越大.并提出了一个经验方程用以关联分配系数与浸取温度、甲醇相浓度的关系,平均相对误差为4.65%.     

Type of cottonseed and level of gossypol in diets of lactating dairy cows: effects on lactation performance and plasma gossypol

Objectives were to determine the effects of feeding whole linted Upland (WUP) and cracked Pima cottonseed (BUPCP) to lactating cows on plasma gossypol concentrations and lactation performance. Holstein cows (n = 813) from three commercial dairies were assigned to one of two diets starting at 13+/-11 d in milk (DIM) for a 170-d experimental period. Cottonseed was included at 10% of the diet dry matter, and treatments consisted of replacing WUP with a 1:2 blend of BUPCP. Blood was collected from all cows at 61 and 91 DIM and from a subset of 56 cows on one dairy at 10, 33, 61, 91, 120, and 152 DIM. Intakes of free gossypol increased 32% in cows receiving the BUPCP diet, and concentrations of total gossypol (TG), as well as the proportion of TG as minus (-) isomer in plasma, were higher for cows fed BUPCP than WUP. Plasma TG increased linearly with increasing DIM, but cows fed the BUPCP, especially multiparous cows, had a higher increase in plasma TG over time. Yields of milk and milk components did not differ between the two treatments, although, yields of milk and milk components were greater in cows with higher plasma TG. Replacement of WUP cottonseed with BUPCP cottonseed increased plasma gossypol, but dry matter intake and yields of milk and milk components were not affected.     

棉粕脱毒工艺及游离棉酚对蛋鸡性能影响的研究进展

我国棉粕资源丰富,棉粕是优良的植物性蛋白饲料,粗蛋白含量高,基本都在40%以上,氨基酸种类丰富,但是棉粕中的游离棉酚(FG)严重限制了棉粕的利用。长期饲喂棉粕日粮会引起蛋鸡一系列的不良反应。对FG的毒性机理和脱除方法进行了阐述,总结了FG对产蛋性能、蛋品质的影响以及在体内各组织的残留规律,为棉粕的利用提供参考,并进一步指出棉粕在畜禽生产应用中需要解决的问题。     

四甲氧基棉酚对SD大鼠的毒性作用

目的 研究游离棉酚降解产物四甲氧基棉酚对SD大鼠的毒性作用。方法 将40只SD大鼠随机分为2组, 雌雄各半: 给药组(四甲氧基棉酚, 25 mg/kg ), 对照组(不含四甲氧基棉酚的棉籽油), 每组大鼠灌胃体积1.50 mL/100 g, 每天1次, 连续灌胃30 d。结合病理组织切片和血液及尿液生化指标评价四甲氧基棉酚的毒性。结果 给药组与对照组大鼠肝脏及脾脏系数存在显著性差异(P?0.01), 其他脏器与体重系数均无显著性差异(P>0.05); 与对照组相比, 给药组大鼠肝脏组织切片有明显病变, 其他器官组织切片无明显病理变化;与对照组相比, 给药组大鼠血液与尿液生化指标中: 天冬氨酸转氨酶(aspartate aminotransferase, AST)、尿氮素(urine nitrogen, BUN)、葡萄糖(glucose, GLU)、胆红素(bilirubin, Bb), 差异有统计学意义(P?0.05), 其他生化指标均无显著性差异(P>0.05), 其中给药组的BUN和AST含量较对照组低, Bb及GLU含量较对照组高。 结论 长期灌服四甲氧基棉酚能够抑制肝脏氨基酸转氨酶的活性和机体对含氮物质的分解、代谢和利用, 从而引起肝损伤。     

HPLC测定棉籽粕中游离棉酚含量

建立了棉籽粕中游离棉酚(FG)的高效液相色谱法(HPLC)测定。结果表明,HPLC测定FG的最佳色谱条件为:流动相V(甲醇+0.2%磷酸)∶V(V(乙腈)∶V(水)=1∶3)=85∶15(分别使用2个进样器),柱压16.2 MPa,柱温25℃,流速1.0 m L/min,进样量20μL,Waters XTerraMS C18不锈钢色谱柱(250 mm×4.6 mm,5μm),UV检测波长235.5 nm。在最佳条件下,能更好地检测棉籽粕中的游离棉酚,棉酚标准溶液的质量浓度与其峰面积呈正相关,相关系数为0.999 9,在0.15~5μg/m L之间呈良好线性关系,平均回收率在95.67%~98.99%之间,相对标准偏差在1.28%~2.03%之间。     

Type of cottonseed and level of gossypol in diets of lactating dairy cows: plasma gossypol,health, and reproductive performance

Objectives were to determine effects of altering gossypol intake by feeding whole linted Upland (WUP) or a 1:2 blend of WUP and cracked Pima (BUPCP) cottonseed on plasma gossypol (PG) concentrations, reproduction, and health of Holstein cows. Cows, 813, on three dairy farms were assigned to one of two diets starting at 13 +/- 11 d in milk for a 170-d experiment. Diets contained 717 and 951 mg of free gossypol/kg of dry matter from WUP and BUPCP, respectively. Concentrations of PG, as well as the proportion of total gossypol (TG) as the minus isomer were higher for cows fed BUPCP vs cows fed WUP. Conception rate at the first postpartum artificial insemination did not differ between treatments. However, cows consuming the higher gossypol diet had reduced subsequent conception rates and a lower pregnancy rate. Incidence of abortions increased in the higher gossypol diet, and cows that aborted or remained open had higher PG concentrations. Increasing PG concentrations resulted in reduced conception rates and extended days open. The probability of conception after the first artificial insemination declined at a decreasing rate as the plasma TG increased. Incidence of health disorders were unaffected by gossypol intake and PG concentrations. Similarly, gossypol intake and PG concentrations had no effect on culling or mortality. Six cows died in each diet, and none had postmortem signs compatible with gossypol toxicity. Consumption of a high gossypol diet for 170 d had no effect on health of lactating dairy cows, but it increased PG concentrations and impaired reproductive performance.     

挤压膨化温度对棉粕游离棉酚及营养成分的影响

本文研究了不同挤压膨化温度(90、100、110、120和130℃)对棉粕中游离棉酚和营养成分的影响。结果表明棉粕挤压膨化脱酚的最佳温度是120℃,超过这一温度后游离棉酚的降解速率不再随温度的升高而增加。随着挤压膨化温度升高棉粕蛋白氮溶解指数(NSI)迅速降低,棉粕中粗纤维、有效赖氨酸和总赖氨酸的含量也随温度升高而显著降低,但当温度达到120℃时,总氨基酸(AA)和必需氨基酸(EAA)的含量达到最大值。此外,120℃时,棉粕中支链氨基酸(BCAAs)的含量也最高。所以,挤压膨化温度在120℃时,既可以显著降低游离棉酚的含量又可较好的保持棉粕的营养价值。      

棉籽糖提取液的脱色与棉酚去除

用乙醇溶液从棉籽粕中提取的棉籽糖提取液,超滤后用活性炭、Al2O3和离子交换树脂进行脱色处理,经过对比选择了最佳脱色剂活性炭,并对其脱色工艺进行了优化.得到活性炭最佳脱色条件为活性炭添加量4%,温度50℃,pH4.5,时间2 h.在最佳脱色条件下,脱色率达到84%,且脱色液中检测不到游离棉酚的存在.     

棉仁中游离棉酚提取工艺的优化

为了获得高质量的棉籽油和棉粕,并实现副产物棉酚的精制,采用有机溶剂浸出技术研究棉仁中游离棉酚的提取工艺。在单因素试验的基础上,通过正交试验确定了70%丙酮提取游离棉酚的最优工艺条件。试验表明,对游离棉酚提取率影响的因素主次顺序为:提取次数,提取时间,料液比,提取温度;确定的最优提取工艺条件为:提取温度40℃,提取时间50 min,料液比1∶15,提取次数3次,搅拌速率850 r/min;在最优工艺条件下,游离棉酚提取率为89.58%,棉粕中游离棉酚含量仅为400 mg/kg。分别对棉酚标准样品和棉仁中提取的棉酚提取液进行40℃加热处理,结果表明加热会导致游离棉酚转化为其他物质,从0 h加热到8 h时,棉酚提取液中的游离棉酚含量从68.19μg/mL降至25.27μg/mL,损耗率达到62.94%。     

Disruption of bovine oocytes and preimplantation embryos by urea and acidic pH

Feeding cattle diets high in degradable crude protein (CP) or in excess of requirements can reduce fertility and lower uterine pH. Objectives were to determine direct effects of urea and acidic pH during oocyte maturation and embryonic development. For experiment 1, oocytes were matured in medium containing 0, 5, 7.5, or 10 mM urea (0, 14, 21, or 28 mg/dl urea nitrogen, respectively). Cleavage rate was not reduced by any concentration of urea. However, the proportion of oocytes developing to the blastocyst stage at d 8 after insemination was reduced by 7.5 mM urea. In addition, the proportion of cleaved oocytes becoming blastocysts was decreased by 5 and 7.5 mM urea. For experiment 2, putative zygotes were collected -9 h after insemination and cultured in modified Potassium Simplex Optimized Medium (KSOM). Urea did not reduce the proportion of oocytes developing to the blastocyst stage, although 10 mM urea reduced cleavage rate slightly. For experiment 3, dimethadione (DMD), a weak nonmetabolizable acid, was used to decrease culture medium pH. Putative zygotes were cultured in modified KSOM containing 0, 10, 15, or 20 mM DMD for 8 d. DMD reduced cleavage rate at 15 and 20 mM and development to the blastocyst stage at all concentrations. Results support the idea that feeding diets rich in highly degradable CP compromises fertility through direct actions of urea on the oocyte and through diet-induced alterations in uterine pH.     

降解游离棉酚的热带假丝酵母菌的发酵动力学研究

对降解游离棉酚的热带假丝酵母菌的发酵动力学进行了研究.通过Logistic方程,以干物质损失量表示生物量,建立了热带假丝酵母菌固态发酵过程中菌体生长、总糖消耗和游离棉酚含量的动力学模型,应用Origin 8.0对实验数据进行处理,得到了热带假丝酵母菌固态发酵的模型参数.动力学分析结果表明,动力学模型与实验数据能较好地拟合,基本上反映了热带假丝酵母菌固态发酵过程.     

中性蛋白酶降解棉粕中棉酚的研究

该实验旨在利用芽孢杆菌（Bacillus）和乳酸杆菌（Lactobacillus）发酵棉粕,研究对棉粕中游离棉酚降解率及饲料活菌数的影响。研究发现,芽孢杆菌BLCC1-0039在37 ℃发酵24 h后棉酚降解率达到85.89% （P＜0.05）,产中性蛋白酶活性达到2 811.40 U/g发酵料（P＜0.05）;发酵48 h时棉酚降解率达到91.47%,产中性蛋白酶活性达到2303.24 U/g。单独添加中性蛋白酶发酵24 h时,添加量为1 000 U/g的棉酚降解率已达到58.46%,2 000 U/g时棉酚降解率达到72.68%;发酵48 h时1 000 U/g的棉酚降解率已达到73.29%。乳酸菌发酵棉粕不能降解棉酚,但乳酸杆菌BLCC2-0092添加1 000 U/g的中性蛋白酶发酵24 h时,棉酚降解率达到67.13%,pH值降至4.71。     

食用植物油中游离棉酚测定方法的比较

食用植物油中游离棉酚测定方法有紫外分光光度法和苯胺法。通过对两种方法的比较得知:苯胺法虽操作较为繁琐,但检验结果准确,回收率达到83%～117%;紫外分光光度法虽是国家标准第一法,操作亦简单、快速,但测定结果偏高。建议首选苯胺法。     

Prediction of pregnancy viability in bovine in vitro-produced embryos and recipient plasma with Fourier transform infrared spectroscopy

We analyzed embryo culture medium (CM) and recipient blood plasma using Fourier transform infrared (FTIR) metabolomics to predict pregnancy outcome. Individually cultured, in vitro-produced (IVP) blastocysts were transferred to recipients as fresh and vitrified-warmed. Spent CM and plasma samples were evaluated using FTIR. The discrimination capability of the classifiers was assessed for accuracy, sensitivity (pregnancy), specificity (nonpregnancy), and area under the receiver operator characteristic curve (AUC). Within all IVP fresh embryos (birth rate = 52%), high AUC were obtained at birth, especially with expanded blastocysts (CM: 0.80 ± 0.053; plasma: 0.89 ± 0.034). The AUC of vitrified IVP embryos (birth rate = 31%) were 0.607 ± 0.038 (CM, expanded blastocysts) and 0.672 ± 0.023 (plasma, all stages). Recipient plasma generally predicted pregnancy outcome better than did embryo CM. Embryos and recipients with improved pregnancy viability were identified, which could increase the economic benefit to the breeding industry.     

Effects of gossypol from cottonseed on hematological responses and plasma alpha-tocopherol concentration of dairy cows

The effects of feeding whole cottonseed (WCS) and bovine somatotropin (bST) administration on hematological responses and plasma alpha-tocopherol concentrations of lactating dairy cattle were examined. After parturition, multi and primiparous Holstein cows (n = 159) were assigned randomly to a 2 x 2 factorial arrangement of treatments consisting of (0 or 15% WCS and 0 or 208 mg of bST injected every 2 wk starting within 7 d after calving. Blood samples were collected from a subset group of 64 cows at 14, 28, 42, and 56 d postpartum. Blood was collected from all cows (n = 159) at 75, 96 and 120 d postpartum. Blood samples were analyzed for alpha-tocopherol and total and (+)- and (-)-gossypol in plasma. Erythrocyte osmotic fragility, hemoglobin and hematocrit also were determined in blood. The mean concentrations of alpha-Tocopherol, total, and (+)- and (-)-gossypol were higher in cows fed WCS regardless of bST administration and plateaued by d 75 postpartum. Hematocrit and hemoglobin concentrations were not affected by treatments. Erythrocyte osmotic fragility was higher in cows fed WCS, but the increase was attenuated when bST was injected (diet x bST interaction). No clinical signs of gossypol toxicity were observed in the cows consuming the WCS.     

响应面优化单溶剂体系棉酚提取工艺研究

为了降低棉粕的毒性及最大限度地提取棉酚,以棉仁为原料,以甲醇为溶剂,采用溶剂浸出法对棉仁中游离棉酚的提取工艺进行研究。在单因素试验的基础上,通过响应面法对工艺条件进行优化。结果显示各因素影响游离棉酚提取率的主次顺序为:提取时间料液比提取温度搅拌速率;得到的最佳工艺条件为:料液比1∶24,提取温度35℃,提取时间42 min,搅拌速率2 600 r/min,提取次数2次(即先以料液比1∶12提取,提取21 min,再以料液比1∶12提取,提取21min);在最佳工艺条件下,游离棉酚的提取率为68.9%±0.81%,其纯度为81.9%±0.85%。此时,棉粕中游离棉酚残留量为839.7 mg/kg,小于饲料卫生标准(GB 13078—2001)中所规定的棉籽饼、粕中允许的游离棉酚最大残留量1 200 mg/kg。