Simultaneous determination of lysine, lysinoalanine, furosine, and pyridosine in food and feed (author's transl)]

A method for the simultaneous separation and determination of lysinoalanine (LAL), lysine, furosine and pyridosine with an automatic amino acid analyzer using a single Column 4 buffer- and 3 temperature-programme is described. The procedure needs 160 Minutes analysis time and allows a separation and quantitative determination of LAL, when present in amounts of more than 100 ppm (semiquantitative above 10 ppm). All amino acids found in food proteins and all other ninhydrinpositive compounds tested until now do not interfere with the determination.     

A simple method for the preparation of furosine and pyridosine reference material

After heatingN -acetyllysine and glucose for 4 h at 90 °C in the dry state and subsequent acid hydrolysis with 7.8 N HCl, preparative fractionation of the dihydrochlorides of furosine and pyridosine was achieved by cation-exchange chromatography. The lysine derivatives could be prepared with high yield and sufficient purity for the use as reference material.

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Ein einfaches Verfahren zur Darstellung von Furosin- und Pyridosin-Standards

Zusammenfassung Nach dem Erhitzen vonN -Acetyllysin und Glucose für 4 h bei 90 °C in trockenem Zustand und nachfolgender Hydrolyse mit 7.8 N HCl erfolgte eine präparative Fraktionierung der Dihydrochloride von Furosin und Pyridosin durch Kationenaustauschchromatographie. Die Lysinderivate wurden in guter Ausbeute und in für Referenzmaterial ausreichender Reinheit erhalten.

     

赖氨酸Amadori和Heyns化合物热裂解产物对烟草风味的影响研究

  目的  比较赖氨酸Amadori和Heyns化合物的热解产物及对其烟草风味的影响。  方法  以赖氨酸和葡萄糖或果糖为原料,以偏重亚硫酸钠为催化剂、甲醇和冰醋酸为溶剂,分别合成赖氨酸的Amadori和Heyns化合物,采用在线热裂解-气相色谱/质谱联用技术(Py-GC/MS)对比了2种化合物的热解产物差异,并通过感官评吸实验比较了二者对卷烟风味的影响。  结果  （1）赖氨酸Amadori和Heyns化合物热解均能产生醛酮类、呋喃类、吡啶类、吡嗪类及吡咯类香味物质,同一类别产物的相对含量有较大差异。（2）赖氨酸Heyns化合物热解得到的风味成分数量更多。（3）赖氨酸Amadori化合物和Heyns化合物对卷烟的主要作用为增香、增浓和降低刺激,并使烟气更加流畅顺滑,赖氨酸Heyns化合物增加了卷烟白肋烟特征。  结论  赖氨酸Amadori化合物和Heyns化合物的热解产物不同,对卷烟风味的影响存在差异。      

灭菌乳产品不同生产工艺及储存条件下糠氨酸的研究

目的研究不同生产工艺及储存条件对灭菌乳产品糠氨酸的影响,建立对应生产过程的管控要求。方法利用高效液相色谱法,对不同生产工艺、储存温度、储存时间条件下的灭菌乳产品中的糠氨酸含量进行了检测与比较。结果用巴氏杀菌降膜系统的产品糠氨酸含量均高于使用巴氏杀菌闪蒸系统的产品,带有无菌罐的产品糠氨酸含量低于不带无菌罐的产品;储存温度越高、时间越长,乳产品中糠氨酸的含量更高。结论灭菌乳类产品糠氨酸含量受不同生产工艺、储存时间和储存温度的影响,乳产品在生产环节应尽量减小热处理程度,同时控制产品储存过程温度和时间。     

Effect of carbohydrate DE on blocked lysine and furosine in a liquid nutritional product

A controlled, quantitative study of Maillard browning vs. dextrose equivalents (DE) was performed on a liquid nutritional product. The early stage Maillard markers furosine and available lysine were determined in retort-sterilised, pilot scale batches formulated with carbohydrate systems with DE variations of 2, 4, 10, and 20. Both markers varied proportionately with DE; every DE increase of 2 units resulted in the blockage (glycation) of an additional 1% of total lysine. When DE 20 maltodextrin was replaced with an 80/20 blend of DE 5 maltodextrin and sucrose (blend DE = 4), lysine blockage decreased by 831 mg/100 g protein, which was 8.25% of the total lysine, and the mole equivalent of 1.75 g of fructoselysine. The quantitative browning/DE relationships enable reliable projections of the nutritional benefits which may be attained through the use of low-DE maltodextrins.     

奶及奶制品中糠氨酸检测方法的研究进展

糠氨酸源于美拉德反应的早期产物,并最终经酸水解产生.它间接反映了美拉德反应的程度、赖氨酸营养功能的损失程度和食品受热程度等,被广泛应用于食品的品质评价.虽然糠氨酸在酸水解时的产率仅有30％～40％,但在相同的检测条件下,该产率是稳定的.近40年来,糠氨酸的检测方法研究主要集中于色谱分离系统和检测器的选择与优化,以及酸水...     

Kinetics of hydroxymethylfurfural,lactulose and furosine formation in milk with different fat content

In the context of the general applicability of hydroxymethylfurfural (HMF), lactulose and furosine as time-temperature integrators (TTIs) for thermal processing of milk, the influence of milk fat content was studied. Formation kinetics were analysed for milk with fat content of 4.0 +/- <0.1%. In previous experiments, it was observed that, under isothermal and non-isothermal heating conditions, formation of the three chemical compounds could be described by pseudo-zero order kinetics. Since the kinetic model was known, the experimental design could be simplified. Data were analysed by a non-linear regression procedure and results were evaluated by construction of joint confidence regions and temperature time tolerance (TTT-) diagrams. Formation kinetics of HMF and lactulose was not affected by milk fat content. Regarding furosine, significant differences were observed between kinetic parameters in whole, semi-skimmed and skimmed milk. The observed differences however were negligible in the context of process impact evaluation.     

赖氨酸在Maillard 反应中形成有害产物AGEs的研究

模拟食品加工条件,构建赖氨酸—还原糖和精氨酸—赖氨酸—还原糖体系,分析各影响因素在体系发生Maillard反应中产生有害AGEs(晚期糖基化终末产物)的作用,比较两个体系添加和不添加黄酮条件下对AGEs形成量的影响。用荧光光谱法(λex/λem=370/440nm)测定AGEs的含量,考察不同因素如还原糖种类、浓度、金属离子、温度、pH和染料木素对有害AGEs形成的影响,以及赖氨酸/精氨酸并存对AGEs形成量的影响。结果表明:在赖氨酸发生Maillard反应过程中,温度越高、还原糖中核糖,糖浓度越大、pH 9.2,形成的AGEs越多;不同的金属离子对AGEs的作用各异,Mg~(2+)和Fe~(2+)能促进AGEs的形成,Ca~(2+)抑制AGEs的形成;抑制剂染料木素浓度越大(9mmol/L),AGEs抑制效果越好;赖氨酸/精氨酸混合体系中AGEs形成量主要由赖氨酸来决定。各影响因素对赖氨酸Maillard反应过程中产生AGEs均有一定作用,时间是最主要因素,其次是pH和温度;而赖氨酸/精氨酸混合体系中AGEs的形成量取决于赖氨酸的含量。     

Evaluation of the furosine and homoarginine methods for determining reactive lysine in rumen-undegraded protein

Three samples of soybean meal (SBM), 3 samples of expeller SBM (SoyPlus, West Central Cooperative, Ralston, IA), 5 samples of distillers dried grains with solubles (DDGS), and 5 samples of fish meal were used to evaluate the furosine and homoarginine procedures to estimate reactive Lys in the rumen-undegraded protein fraction (RUP-Lys). One sample each of SBM, expeller SBM, and DDGS were subjected to additional heat treatment in the lab to ensure there was a wide range in reactive RUP-Lys content among the samples. Furosine is a secondary product of the initial stages of the Maillard reaction and can be used to calculate blocked Lys. Homoarginine is formed via the reaction of reactive Lys with O-methylisourea and can be used to calculate the concentration of reactive Lys. In previous experiments, each sample was ruminally incubated in situ for 16 h, and standardized RUP-Lys digestibility of the samples was determined in cecectomized roosters. All rumen-undegraded residue (RUR) samples were analyzed for furosine and Lys; however, only 9 of the 16 samples contained furosine, and only the 4 unheated DDGS samples contained appreciable amounts of furosine. Blocked RUP-Lys was calculated from the furosine and Lys concentrations of the RUR. Both the intact feed and RUR samples were evaluated using the homoarginine method. All samples were incubated with an O-methylisourea/BaOH solution for 72 h and analyzed for Lys and homoarginine concentrations. Reactive Lys concentrations of the intact feeds and RUR were calculated. Results of the experiment indicate that blocked RUP-Lys determined via the furosine method was negatively correlated with standardized RUP-Lys digestibility, and reactive RUP-Lys determined via the guanidination method was positively correlated with standardized RUP-Lys digestibility. Reactive Lys concentrations of the intact samples were also highly correlated with RUP-Lys digestibility. In conclusion, the furosine assay is useful in predicting RUP-Lys digestibility of DDGS samples, and the guanidination procedure can be used to predict RUP-Lys digestibility of SBM, expeller SBM, DDGS, and fish meal samples.     

赖氨酸/葡萄糖美拉德反应产物对丙烯酰胺生成的抑制作用

建立天冬酰胺/葡萄糖模式反应体系,运用高效液相色谱法、自由基清除法等手段研究赖氨酸/葡萄糖美拉德反应产物对该模式体系中丙烯酰胺生成量及体系抗氧化性的影响。结果显示:高、低分子量的赖氨酸/葡萄糖美拉德反应产物(H-LGP和L-LGP)均能显著降低模式反应体系中的丙烯酰胺生成量,其中H-LGP和L-LGP的添加量为10 mg/mL时,对丙烯酰胺生成的抑制率最大,分别为32.0%和31.4%。H-LGP和L-LGP(均为10mg/mL)的添加还可显著提高反应体系清除自由基DPPH和ABTS的能力。结论:赖氨酸/葡萄糖美拉德反应产物(LGP)能够显著抑制天冬酰胺/葡萄糖模式反应体系中丙烯酰胺的生成,且高浓度的LGP能够显著提高反应体系的抗氧化性。     

Effect of Amadori rearrangement products on the non-enzymic browning in model systems

The Amadori product was prepared from D-glucose and L-alanine after Hashiba. The browning process was investigated by determining the absorbance at 520 nm, in aqueous solutions at 110 °C for 5–8 h. Because of the limited access of oxygen and its low solubility in the reaction medium, the browning proceeded after zeroth order kinetics with the maximum browning rate at pH = 8–9. The browning rate remained unaffected by additions of sodium sulphite, rutin. propyl gallate (except when present at high levels), iron(III)chloride or copper(II)chloride but decreased in presence of L-cysteine or iron(II)chloride. Hydrogen peroxide bleached the pigment but did not inhibit the subsequent browning of reaction products. Under experimental conditions the solution of Amadori product did not darken with substantially greater rate than the solution of D-glucose and L-alanine. Lower additions of D-glucose to the solution of Amadori product moderately increased the reaction rate while additions of L-alanine or L-hydroxyproline moderately decreased the browning rate.     

Studies on the formation of lysinomethylalanine and histidinomethylalanine in milk products

From reaction mixtures consisting ofN-acetyldehydroaminobutyric acid methyl ester andN -acetyl-l-lysine orN -acetyl-l-histidine, respectively, distinct amounts of the cross-link amino acidsN -(2-amino-2-carboxy-l-methylethyl)-l-lysine (lysinomethylalanine, LMeAL) andN -(2-amino-2-carboxy-1-methyl-ethyl)-l-histidine (histidinomethylalanine, HMeAL) were isolated via preparative ion-exchange chromatography and identified by¹H- and¹³C-nuclear magnetic resonance. In the amino acid chromatogram, both compounds eluted clearly separated from other basic amino acids. However, neither LMeAL nor HMeAL could be detected in numerous acid hydrolysates of a range of milk products. In model studies, threonine showed a significantly lower tendency for an alkali-induced -elimination reaction compared to serine. The reactivity of the resulting dehydroaminobutyric acid towards nucleophiles was more than tenfold lower as compared to dehydroalanine. Thus, the formation of LMeAL as well as of HMeAL during food processing is negligible.

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Studien zur Bildung von Lysinomethylalanin und Histidinomethylalanin in Milchprodukten

Zusammenfassung Aus Reaktionsansätzen bestehend ausN-Acetyldehydroaminobuttersäurenthylester undN -Acetyl-l-lysin beziehungsweiseN -Acetyl-l-histidin konnten die Crosslink-AminosäurenN -(2-Amino-2-carboxy-1-methyl-ethyl)-l-lysin (Lysinomethylalanin, LMeAL) undN -(2 amino-2-carboxy-2 -methyl-ethyl)-l-histidin (Histidinomethylalanin, HMeAL) durch präparative Ionenaustauschchromatographie isoliert und anschließend durch 1H-und¹³C-NMR identifiziert werden. Das Aminosäurechromatogramm zeigte eine eindeutige Trennung beider Aminosäurederivate von anderen basischen Aminosäuren. Dennoch konnten in einer großen Zahl von Säurehydrolysaten der verschiedensten Milchprodukte weder LMeAL noch HMeAL nachgewiesen werden. In Modelluntersuchungen erfolgte die -Elimination an Threonin im Vergleich zu Serin in einem deutlich geringeren Maße. Die Reaktivität der dabei entstehenden Dehydroaminobuttersäure gegenüber Nucleophilen war mehr als zehnfach geringer als die von Dehydroalanin. Damit kann die Bildung sowohl von LMeAL als auch von HMeAL bei der Verarbeitung von Nahrungsmitteln als vernachlässigbar gering eingeschätzt werden.

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Dedicated to Prof. Dr. W. Baltes on the occasion of his 65th birthday     

盐酸处理对壳聚糖的降解作用研究

研究以壳聚糖为原料,针对壳聚糖因水溶性差,反应物浓度小的难题,采用醋酸溶解和添加2%的盐酸,于90℃保温搅拌6h,可以使此壳聚糖体系的黏度从3.00Pa.s降至0.266Pa.s,溶液流动性好,同时可使壳聚糖浓度提高到10%。该处理方法不仅使壳聚糖浓度增加,而且又避免了采用高温加热法易产生单糖的缺陷。     

Effects of different heat treatments on the furosine content in fresh filled pasta

The effect of different heat treatments on the furosine content in fresh filled pasta was studied and the rate of furosine increase after treatment has been found to be influenced by the initial furosine content in the mixture and by the treatment design. A mathematical model to describe the experimental data has been developed. The reaction of furosine formation appears to follow a pseudo-zero kinetic order. The estimated value of activation the energy is roughly 111 KJ mole⁻¹ and the z value has been estimated around 22.9 °C. The “furosine increase” (If) parameter has been calculated from processing data in order to compare samples obtained with different time–temperature processing combinations. Calculated values were correlated with F₇₀¹⁰ values (pasteurising effect) and results indicate a good linear relationship between the furosine increase and F₇₀¹⁰ values (r²=0.963; P<0.01). The comparison of the pasteurisation curve with the furosine increase curve shows that an optimal process design for thermal processing of fresh filled pasta under our experimental conditions is obtained at temperatures between 95 and 99 °C for times from 6 to 2 min.     

鲣鱼双酶酶解工艺优化及其产物动态变化研究

以鲣鱼为对象,优化了胰蛋白酶(trypsin)与木瓜蛋白酶(papain)复合酶解鱼肉蛋白的条件,研究了酶解过程中酶解液中游离氨基酸组成和含量、肽分子质量分布以及呈味核苷酸二钠含量的动态变化规律。结果表明:木瓜蛋白酶和胰蛋白酶双酶酶解鲣鱼最优条件为:最适比例1:3,总酶添加量3 000 U/g,底物浓度为25%。酶解过程中,酶解液中游离氨基酸总量不断增加,酶解6 h后,呈苦味氨基酸含量有所减少,而呈鲜味氨基酸大幅增加;对氨基酸呈味强度值(taste activity value,TAV)的分析表明,Glu、Met、Ile和His等几种氨基酸对酶解液的呈味特性起了关键作用。随着酶解时间的增加,大分子质量蛋白和肽类组分逐渐减少而小分子质量肽类组分逐渐增多,酶解6 h后分子质量小于0.5 ku的组分含量达到80.17%。呈味核苷酸二钠含量随酶解时间缓慢增加。     

Analysis of dithiocarbamate fungicides. Reaction products of the thiuram disulphide fungicide thiram (TMTD) during acid hydrolysis

The acid hydrolysis products (CS₂, COS, and H₂S) of thiram (tetramethylthiram disulphide, TMTD) absorbed in a methanolic amine reagent (ethylenediamine, piperidine) were investigated by second derivative UV spectroscopy. When the hydrolysis temperatures are below the boiling point the formation of the side products COS and H₂S at the expense of CS₂ is unavoidable. Failure to carefully scrub the liberated gases with a lead acetate solution leads to the absorption of H₂S also in the amine reagent and, like COS, causes an absorption at 230 nm that is erroneously attributed to COS. With the second-derivative technique it has been undoubtedly proved that, under the conditions of the Deutsche Forschungsgemeinschaft method S 15, thiram liberates CS₂ and COS in a molar ratio of 1.92 to 0.06 (average of eight determinations) but not in a molar ratio of 1.5 to 0.5, as previously reported by some workers. Under these conditions it is impossible, contrary to earlier claims, to differentiate thiram from other dithiocarbamates based on the ratio of the hydrolysis products in residue analysis.

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Analytik der Dithiocarbamat-Fungicide. Reaktionsprodukte des Thiuramdisulfid-Fungicids Thiram (TMTD) während der Säurehydrolyse

Zusammenfassung Mittels Derivativ-UV-Spektroskopie wurden die Produkte der Säurehydrolyse (CS₂, COS und H₂S) des Fungicids Thiram (TMTD) nach Absorption in einem methanolischen Aminreagens (Ethylendiamin, Piperidin) ermittelt. Bei Hydrolysetemperaturen unter dem Siedepunkt ist die Bildung der Nebenprodukte COS und H₂S auf Kosten von CS₂ unvermeidlich. Erfolgt kein sorgfältiges Waschen der hydrolytisch freigesetzten Gase mit einer Bleiacetatlösung, so wird auch H₂S im Aminreagens absorbiert, das wie COS eine UV-Absorption bei 230 nm verursacht und die COS-Bestimmung verfälscht. Mit der Derivativspektroskopie konnte eindeutig gezeigt werden, daß Thiram unter den Bedingungen der DFG-Methode S 15 zu CS₂ and COS im molaren Verhältnis von 1,92 zu 0,06 (8 Messungen) hydrolysiert wird und nicht, wie von anderen Autoren angenommen, im molaren Verhältnis von 1,5 zu 0,5. Daher ist es, den gehegten Hoffnungen entgegen, nicht möglich, auf der Basis der Verteilung der Hydrolyseprodukte Thiram von anderen Dithiocarbamaten rückstandsanalytisch zu unterscheiden.

     

植物油脂水解工艺及不饱和脂肪酸组成研究

研究了碱性水解小桐子油脂制备脂肪酸的较佳工艺条件：油水比（g/g）为2：1,乙醇-水比（v/v）为2：1,皂化时间1.5h、温度70℃,在该条件下脂肪酸得率、酸值和碘值的平均值分别为：98.25%、203.67mgKOH/g和109.89g/100g。采用桐油、光皮油、棕榈油和大豆油等植物油脂考察了该工艺的原料适用程度,该水解工艺条件下各种油脂制备所得的脂肪酸得率和酸值都接近98%和199mgKOH/g。采用GC-MS分析五种植物油脂脂肪酸组成和含量,其不饱和脂肪酸含量依次为：桐油（94.88%）〉大豆油（84.07%）〉光皮油（81.90%）〉小桐子油（77.84%）〉棕榈油（77.19%）;其中多价不饱和脂肪酸含量次序为桐油（85.36%）〉大豆油（58.01%）〉棕榈油（56.81%）〉光皮油（55.31%）〉小桐子油（42.16%）。大豆油、棕榈油、光皮油和小桐子油四种植物油脂脂肪酸成分基本相同,主要是亚油酸、7,10-十八碳二烯酸、油酸。桐油中脂肪酸组成与其他植物油脂有较大区别,含大量三价不饱和脂肪酸,其主要成分有油酸（9-十八烯酸）13.83%、8,11-十八碳二烯酸24.37%、亚麻酸（9,12,15-十八碳三烯酸和6,9,15-十八碳三烯酸）60.01%。     

Investigation of the formation of Strecker aldehydes from the reaction of Amadori rearrangement products with α-amino acids in low moisture model systems

 The formation of Strecker aldehydes in low moisture model systems (based on microcrystalline cellulose) containing the Amadori rearrangement product (ARP) fructose-leucine (Fru-Leu), the amino acid isoleucine (Ile) and the amine o-phenylenediamine (OPD) respectively in varying molar ratios was investigated. The model systems were heated under constant conditions and the formation of Strecker aldehydes analysed using headspace-GC. The experimental results obtained provide evidence that the formation of Strecker aldehydes does not only occur via ordinary Strecker degradation but also via alternative reaction pathways branching off directly from the ARP. Alternative reaction pathways are proposed. Received: 2 November 1999 / Revised version: 3 February 2000