肉制品中唾液酸含量和形态分析及在加工中的变化

本文主要运用了高效液相色谱法测量了我国牛肉、猪肉等常见肉类及常见熟食制品如火腿、香肠、卤牛肉、风干羊肉、香辣鸭脖中的两种唾液酸N-羟乙酰神经氨酸（N-Glycolylneuraminic acid,Neu5Gc）、N-乙酰神经氨酸（N-Acetylneuraminic acid,Neu5Ac）的含量、存在形式以及不同加工方式对其含量及存在形式的影响。结果表明红肉中Neu5Gc含量：牛肉46.57±2.53 μg/g>猪肉28.69±1.03 μg/g>羊肉29.09±2.32 μg/g,红肉中同时含有结合态Neu5Gc和游离态Neu5Gc,结合态Neu5Gc含量普遍高于游离态Neu5Gc。红肉中Neu5Ac含量：羊肉200.15±24.96 μg/g>猪肉110.89±5.71 μg/g>牛肉80.97±5.60 μg/g。白肉中不含Neu5Gc,鸭肉的Neu5Ac含量最高,为185.73±23.11 μg/g。熟食肉制品中,牛肉制品的Neu5Gc含量仍高于其他红肉熟食制品,并在白肉熟食制品中检测出了Neu5Gc的存在。对红肉进行蒸煮、油炸及腌制处理后Neu5Gc和Neu5Ac含量都有所下降,其中经过油炸处理的样品Neu5Gc和Neu5Ac含量下降最为明显,其次是腌制、蒸煮。本文还研究了蒸煮时间对猪肉中唾液酸含量的影响,即随着蒸煮时间的增加,Neu5Gc、Neu5Ac含量变化的总体趋势减少,蒸煮45 min后结合态唾液酸明显减少,游离态增多。即红肉中特异性含有Neu5Gc,白肉类熟食制品中也可能含有Neu5Gc,通过加工可改变肉制品中唾液酸的含量和形式,但是每种加工方式对于唾液酸含量及形式的改变不同。     

燕窝的精华——唾液酸

正1唾液酸简介唾液酸(Sialic Acid)广义地说是一系列含有氨基的九碳单糖衍生物,是所有神经氨酸及其衍生物的总称。但在人体当中只有一种,即N-乙酰神经氨酸(N-acetylneuraminic acid,Neu5Ac),它就是众所周知,大名鼎鼎的"燕窝酸"。因此,对我们人类来说,唾液酸就是特指"N-乙酰神经氨酸"。它最初由牛颌下腺粘蛋白中分离而出,也因此而得名唾液酸。唾液酸通常以低聚糖,糖脂或者糖蛋白的形式存在,主要以短链残基的形式     

不同加工工艺对燕窝产品唾液酸含量的影响

本文以毛燕为原料,研究浸泡、挑拣、高温灭菌等工艺流程处理后燕窝唾液酸含量的变化。采用LC-MS/MS检测分析燕窝中唾液酸种类,此外,测定了不同加工条件下燕窝产品唾液酸含量并分析加工条件对唾液酸含量的影响,进一步分析了唾液酸热稳定性。结果表明:燕窝中唾液酸仅由N-乙酰基神经氨酸(Neu5Ac)组成,经不同工艺条件处理后,Neu5Ac保留率均可高达95%以上,且Neu5Ac热稳定性好,不同情况下Neu5Ac含量无明显差异。说明燕窝经过上述加工处理后不会造成燕窝酸的大量流失,并且其受工艺条件变化影响小,燕窝加工产品可以很好地保存燕窝的营养价值,从而稳定发挥其营养功效,具有很高的消费价值。     

高效液相色谱法测定母乳中唾液酸含量

建立荧光高效液相色谱(fluorescence detector-high performance liquid chromatography,HPLC-FLD)测定母乳中唾液酸N-乙酰神经氨酸(N-acetylneuraminic acid,Neu5Ac)和N-羟乙酰神经氨酸(N-glycolyl neuraminic acid,Neu5Gc)含量的分析方法。利用酸水解法释放出母乳中的唾液酸,以4,5-亚甲二氧基-1,2-邻苯二胺盐(4,5-methylenedioxy-1,2-phenylenediamine dihydrochloride,DMB)为衍生化试剂,50℃避光衍生150min,采用荧光高效液相色谱仪检测。色谱条件：LiChrosorb RP-18柱(250mm×4mm,5μm),流动相为甲醇-乙腈-超纯水(7:8:85),流速0.9mL/min,进样体积10μL,柱温30℃,荧光检测器激发波长373nm,发射波长448nm。结果表明：唾液酸在50～400μmol/L范围内与唾液酸峰面积的线性关系良好,平均回收率为94.0%,精密度的相对标准偏差(relative standard deviation,RSD)为0.4%,稳定性RSD为1.0%,重复性RSD为0.8%,Neu5Ac的最低检出限为0.02μmol/L,Neu5Gc的最低检出限位0.03μmol/L。该方法简单、重复性好、灵敏度高,可广泛用于奶粉、牛奶及母乳中唾液酸含量测定。     

不同处理方式对红肉中N-羟乙酰神经氨酸解离的影响

应用不同方法处理红肉(猪肉与牛肉),研究能够有效解离红肉中N-羟乙酰神经氨酸(N-glycolylneuraminic acid,Neu5Gc)的预处理方式。将红肉通过水煮、微波加热和有机酸腌制等不同方式进行处理,利用酸水解法释放红肉中的两种唾液酸成分Neu5Gc和N-乙酰神经氨酸(N-acetylneuraminic acid,Neu5Ac),经1,2-二氨基-4,5-亚甲基二氧苯(1,2-diamino-4,5-methylenedioxybenzene,DMB)衍生化后用高效液相色谱检测其含量;另外还采用β-半乳糖苷酶水解红肉,再测定水解液中的Neu5Gc和Neu5Ac。结果表明:沸水浴处理红肉后其Neu5Gc和Neu5Ac都有一定程度的解离,水煮时间越长,红肉中Neu5Gc和Neu5Ac解离效果越好,且猪肉中的Neu5Gc相比牛肉的更容易水煮解离。采用微波炉高温处理红肉也能够解离Neu5Gc,但是解离率不超过70.0%,且处理时间影响较小;而Neu5Ac的解离率都低于Neu5Gc。红肉通过不同的弱有机酸腌制处理后Neu5Gc和Neu5Ac解离率差别不大,且醋酸腌制处理后Neu5Gc解离效果较好。另外,β-半乳糖苷酶能有效解离猪肉中Neu5Gc,水解时间越长,解离率越高。对于牛肉,Neu5Gc解离率最高为84.0%,解离效果不如猪肉。     

高效液相色谱-荧光检测法测定红肉及其加工肉中唾液酸含量

采用高效液相色谱-荧光检测法对红肉及其加工肉中两种唾液酸N-乙酰神经氨酸（N-acetylneuraminic acid,Neu5Ac）和N-羟乙酰神经氨酸（N-glycolylneuraminic acid,Neu5Gc）进行定性和定量分析。利用单因素试验对衍生化与样品酸解条件进行优化,并借助超声缩短酸解时间。结果表明,以盐酸作为酸解试剂,超声辅助酸解30 min,4,5-亚甲二氧基-1,2-邻苯二胺盐作为衍生化试剂,衍生化试剂浓度为13 mmol/L,样品与衍生化试剂比值为1∶1,衍生化时间120 min时,检测效果最佳。Neu5Ac和Neu5Gc在0.1～10 μg/mL范围内线性良好,回收率为91.2%～119.7%,检出限分别为0.003 mg/kg和0.01 mg/kg,重复性的相对标准偏差（relative standard deviation,RSD）为0.7%～1.8%,精密度RSD分别为1.4%和1.2%。本方法具有灵敏度高、分析时间短、重复性及准确性好等特点。     

牛免疫球蛋白G的体外人源唾液酸化及Fc片段的制备

建立将牛免疫球蛋白G（bovine immunoglobulin G,bIgG）糖链末端N-羟乙酰神经氨酸酶切并连接人源N-乙酰神经氨酸（N-acetylneuraminic acid,Neu5Ac）的方法,在实现bIgG转化为人源IgG（human IgG,hIgG）的基础上,研究hIgG可结晶（Fc）片段的制备。结果表明：以170 U/mL神经氨酸酶酶切bIgG（4.0 mg/mL）后,通过β-1,4-半乳糖基转移酶和α-2,6-唾液酸转移酶分别转移半乳糖（galactose,Gal）和Neu5Ac残基,可制备增加8.4 个Gal残基和42 个Neu5Ac残基的hIgG分子。此外,在10.0 mg/mL hIgG、m（木瓜蛋白酶）∶m（hIgG）＝0.05、10.0 mmol/L半胱氨酸激活剂、2.0 mmol/L EDTA溶液、pH 7.0条件下,酶解3 h可制得hIgG的较高纯度Fc片段,最终hIgG得率为71.7%,Fc片段得率为20.8%。本研究为bIgG的产品开发和营养价值评价提供科学依据。     

9种鱼卵中的游离态及结合态唾液酸含量的分析比较研究

从不同品种鱼卵中提取出游离态唾液酸和总唾液酸，采用DMB试剂对这2种唾液酸分别进行衍生化标记，再利用高效液相色谱(HPLC)系统性对比分析了9种鱼卵中N-乙酰神经氨(Neu5Ac)和N-羟乙酰神经氨酸(Neu5Gc)的存在形式及含量的差异。结果表明，游离态和结合态的Neu5Ac在鲫鱼鱼卵中含量最高，每克鱼卵干重中分别含1.61μg和4.07μg;Neu5Gc仅存在于黄花鱼、鳜鱼和鲂鱼的鱼卵中，其中鲂鱼鱼卵中游离态Neu5Gc含量最高，每克鱼卵干重中含0.57μg；而鳜鱼鱼卵中结合态Neu5Gc含量最高，每克鱼卵干重中含0.24μg。以上结果为鱼卵的深度开发利用提供理论依据。     

安琪"自然醇厚、回味持久"美味解决方案

1唾液酸简介唾液酸(Sialic Acid)广义地说是一系列含有氨基的九碳单糖衍生物,是所有神经氨酸及其衍生物的总称。但在人体当中只有一种,即N-乙酰神经氨酸(N-acetylneuraminic acid,Neu5Ac),它就是众所周知,大名鼎鼎的"燕窝酸"。因此,对我们人类来说,唾液酸就是特指"N-乙酰神经氨酸"。它最初由牛颌下腺粘蛋白中分离而出,也因此而得名唾液酸。唾液酸通常以低聚糖,糖脂或者糖蛋白的形式存在,主要以短链残基的形式     

4种禽蛋中氨基酸成分的测定及分析

采用全自动氨基酸分析仪测定4种禽蛋对应的蛋清、蛋黄中的16种氨基酸含量,对4种蛋类氨基酸组成及水平进行比较分析,进而评价4种蛋类的营养价值。试验结果表明：4种禽蛋中鹌鹑蛋的氨基酸总含量最高,其次是鸡蛋与鸭蛋,鹅蛋的氨基酸含量最低;将同种蛋的蛋黄与蛋清中氨基酸的含量进行比较发现,4种禽蛋的蛋黄氨基酸含量均高于蛋清的氨基酸含量;4种禽蛋蛋清、蛋黄都符合理想蛋白要求,均为优质蛋白质,并且通过比较发现,每种蛋类对应的蛋黄的必需氨基酸/总氨基酸更接近40%,说明蛋黄蛋白要优于蛋清蛋白;与联合国粮农组织（Food and Agriculture Organization of the United Nations,FAO）/世界卫生组织（World Health Organization,WHO）推荐的标准模式谱相比较,除鸡蛋中蛋氨酸占总氨基酸的质量分数（3.09）低于推荐值（3.5）,其余的比值均高于或接近推荐值。说明这4种禽蛋的必需氨基酸组成合理,具有较高的营养价值,是理想的膳食蛋白质来源。     

Comparison of spectrophotometric and HPLC methods for determining sialic acid in infant formulas

Two methods for determining sialic acid in infant formulas – spectrophotometry and HPLC with fluorescence detection – have been optimised and validated, the first one allows to determine total sialic acid while the second allows to differentiate the two main forms of sialic acid (N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc)). A common sample preparation procedure (hydrolysis and purification) for both methods has been proposed. The linearity (from 6 to 150 μg of total sialic acid in the assay for spectrophotometry, and from 12.5 to 250 ng and 1 to 5 ng of Neu5Ac and Neu5Gc, respectively, for HPLC) is adequate. The detection and quantification limits (0.29 and 0.97 mg of total sialic acid/L of reconstituted sample, respectively, for spectrophotometry, and 0.03 and 0.08 mg Neu5Ac/L; 0.003 and 0.009 mg Neu5Gc/L of reconstituted sample, respectively, for HPLC) are low enough for the determination of sialic acid in infant formulas. The precision of both methods, expressed as relative standard deviation, is less than 6%, and the accuracy evaluated by recovery assays show 104% recovery for spectrophotometry; 95% for Neu5Ac and 109% for Neu5Gc for HPLC. Samples analysed show no significant differences (α < 0.05) attributable to the method used; consequently, both of them could be applied after common sample preparation, the choice of technique depending on the facilities available in the laboratory.     

Rapid and simple method for the determination in sialic acid of yak milk fat globule membrane

A rapid and simple method for the determination of sialic acid (Neu5Ac) of yak milk fat globule membrane (MFGM) by HPLC with a diode array detector was developed and validated. Samples were cleaned up just by hydrolysis and derivatization before HPLC analysis. Separation was achieved with an Agilent TC-C18 column. The method showed a good linearity (r=0.997), the sensitivity results showed that the limits of detection and limits of quantification for sialic acid were 10.0 and 21.0 μg/mL, respectively. The recovery of Neu5Ac was 95–97%. The method proved very simple and rapid for Neu5Ac analysis since separation was completely achieved at 12 min.     

Determination of sialic acids in infant formula by chromatographic methods: a comparison of high-performance anion-exchange chromatography with pulsed amperometric detection and ultra-high-performance liquid chromatography methods

Sialic acid determination in an infant formula presents many challenges, including efficient sialic acid release from glycoconjugates, effective sample preparation, and rugged chromatography. This work compares 2 chromatographic assays developed for determination of sialic acids in infant formula. Prior to chromatography, both assays release sialic acids by acid hydrolysis and treat the hydrolysate with a subsequent anion-exchange sample preparation. Both high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and fluorescence ultra-high-performance liquid chromatography (UHPLC) sample analysis methods were evaluated to compare assay performance and convenience. Calibration ranges were chosen to encompass the expected amounts of 2 sialic acids in infant formula: N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc). Response was linear by either method with coefficients of determination of 1.00 by HPAEC-PAD between 5.0 and 100pmol of Neu5Ac and between 0.34 and 6.8 pmol of Neu5Gc and >0.99 by UHPLC between 5.0 and 260 pmol of Neu5Ac and between 0.20 and 9.8 pmol of Neu5Gc. Both methods had sufficient sensitivity to determine these sialic acids in infant formula. Three infant formulas were analyzed to evaluate accuracy and precision of the assays. The HPAEC-PAD assay was found to be faster overall and the UHPLC assay was more sensitive. Reaction efficiency, and therefore sensitivity, was dependent on the sample matrix. This work illustrates sample-specific complexity that must be considered in choosing an assay.     

Determining Neu5Ac in infant formula with ultra‐performance liquid chromatography–tandem mass spectrometry

The aim of this work was to measure N‐acetylneuraminic acid (Neu5Ac) in milk‐based infant formulas. The analysis was performed by ultra‐performance liquid chromatography–tandem mass spectrometry (UPLC‐MS/MS). The total Neu5Ac were released using trichloroacetic acid and hydrochloric acid and purified using a HLB column. The linearity from 0.05 to 5.0 μg/mg Neu5Ac was adequate. Sialic acid recoveries ranged from 91.8% to 112.4%. The detection and quantification limits (limit of detection, 0.01 μg Neu5Ac/mg; limit of quantitation, 1.08 μg Neu5Ac/mg) were low enough to determine the sialic acid in infant formulas. The validated method is highly reproducible and sensitive, and it is easy to perform.     

Dietary intervention with sialylated lactulose affects the immunomodulatory activities of mice

Four sialylated lactuloses [N-acetylneuraminic acid-α2,3-lactulose (Neu5Acα2,3lactulose), N-acetylneuraminic acid-α2,6-lactulose (Neu5Acα2,6lactulose), deaminoneuraminc acid-α2,3-lactulose (Kdnα2,3lactulose), and deaminoneuraminc acid-α-2,6-lactulose (Kdnα2,6lactulose)] were reported to modulate the immunity of mice. The influences of cytokine expression, cell immunity, humoral immunity, and nonspecific immunity were investigated in our study using several techniques. Analysis via ELISA showed that cytokine expression was induced by sialylated lactulose treatment consistently in the serum and spleen. Among the 4 tested sialylated lactuloses, Neu5Acα2,6lactulose performed the best, simultaneously and appropriately promoting the expression of proinflammatory and anti-inflammatory factors in the serum and spleen. Kdnα2,3lactulose showed the best antioxidant activity according to detection of the activity of superoxide dismutase, myeloperoxidase, peroxidase, and alkaline phosphatase. Flow cytometry revealed that only Kdnα2,3lactulose significantly boosted the CD3⁺ T lymphocyte ratio similarly to that of lactulose. Analysis of the hemolysin content to characterize humoral immunity revealed that Kdnα2,3lactulose notably increased hemolysin content compared with that in the control group. To evaluate the nonspecific immune effects of the 4 sialylated lactuloses, a fluorescence microsphere phagocytosis assay was used to analyze the phagocytosis of macrophages. Kdnα2,3lactulose still performed the best in enhancing the phagocytosis of macrophages, showing markedly increased phagocytic percentage and phagocytic index values compared with those in the control and lactulose groups. Comparing the differences of these 4 sialylated lactuloses in affecting immunity in mice revealed that Kdnα2,3lactulose had the best overall performance in influencing cytokine expression, cell immunity, humoral immunity, and nonspecific immunity. This study provides critical support for use of sialylated lactuloses as potential immunomodulators in foods.     

Use of Mucor miehei lipase to improve functional properties of yolk-contaminated egg whites

Egg yolk contamination of egg whites continues to be a serious problem in the egg industry. The ability of egg whites to form stable and voluminous foams is greatly inhibited by yolk contamination, even at very low levels, between 0.01% and 0.2% w/w yolk in white. Experiments were conducted to determine if Mucor miehei lipase could regenerate the functional properties of yolk-contaminated egg whites. Lipase from M. miehei and colipase from porcine pancreas were added to yolk-contaminated (0.2%, w/w) egg white samples to hydrolyze triglycerides originating from egg yolk. Enzymatic hydrolysis of triacylglycerols was confirmed using thin-layer chromatography. Treatment of yolk-contaminated samples with lipase and colipase yielded significant (P < 0.05) improvements in a number of the functional properties, including the final foam volume, foam capacity, and foaming power. These functional properties showed complete restoration to control levels. However, foam stability and foam drainage levels were not statistically different from yolk-contaminated samples that had not been enzymatically treated. Enzyme-treated yolk-contaminated egg whites were also tested in angel food cakes. Enzyme-treated, yolk-contaminated egg whites performed similarly to non-yolk-contaminated control, and much better than yolk-contaminated sample in angel food cakes. The results show that most negative effects of yolk contamination can be reversed by treatment with Mucor miehei lipase and colipase.     

气相色谱法同时测定多糖中的中性糖、糖醛酸、氨基糖和唾液酸

采用甲醇解和硅烷衍生化,用气相色谱法同时测定样品中的中性糖、糖醛酸、N-乙酰氨基糖和唾液酸,取得了很好的效果。3种标准单糖混合物(包含阿拉伯糖、鼠李糖、岩藻糖、木糖、甘露糖、半乳糖、葡萄糖、葡萄糖醛酸、半乳糖醛酸、N-乙酰半乳糖、N-乙酰葡萄糖、5-乙酰唾液酸)和2种多糖样品(豆腐渣多糖DFP、鸡腿菇多糖F32)用1 mol/L盐酸甲醇在85℃反应18～24 h,碳酸银中和,加入乙酸酐室温暗处反应24 h,使氨基糖重新引入N-乙酰基,除银盐,干燥,加入硅烷化试剂[V(吡啶)∶V(六甲基二硅氨烷)∶V(三甲基氯硅烷)=5∶1∶1),室温放置30 min,最后用气相色谱进行分析。