Effect of Freeze-Thaw Cycles on the Oxidation of Protein and Fat and Its Relationship with the Formation of Heterocyclic Aromatic Amines and Advanced Glycation End Products in Raw Meat

The aim of this research was to investigate the effect of the number of freeze–thaw cycles (0, 1, 3, 5, and 7) on porcine longissimus protein and lipid oxidation, as well as changes in heterocyclic aromatic amines (HAAs) and advanced glycation end products (AGEs) and their precursors. We analyzed the relationship among HAAs, AGEs, oxidation, and precursors and found the following results after seven freeze–thaw cycles. The HAAs, Norharman and Harman, were 20.33% and 16.67% higher, respectively. The AGEs, Nε-carboxyethyllysine (CEL) and Nε-carboxymethyllysine (CML), were 11.81% and 14.02% higher, respectively. Glucose, creatine, and creatinine were reduced by 33.92%, 5.93%, and 1.12%, respectively after seven freeze–thaw cycles. Norharman was significantly correlated with thiobarbituric acid reactive substances (TBARS; r² = 0.910) and glucose (r² = −0.914). Harman was significantly correlated to TBARS (r² = 0.951), carbonyl (r² = 0.990), and glucose (r² = −0.920). CEL was correlated to TBARS (r² = 0.992) and carbonyl (r² = 0.933). These changes suggest that oxidation and the Maillard reaction during freeze–thaw cycles promote HAA and AGE production in raw pork.     

基于紫外及荧光光谱方法的中药抗氧化应激作用研究

糖尿病引起患者全身多部位发生慢性并发症,严重危害患者的生命安全,成为其致死致残的主要原因。糖尿病患者体内的高血糖环境引发的组织内氧化应激反应增加是糖尿病并发症的重要发病机制之一,而蛋白非酶糖基化反应和脂质过氧化反应过程与氧化应激密切相关。从糖尿病并发症的发病机理出发,以氧化应激为基点,运用光谱法对17种中药提取物抗氧化应激反应的活性进行综合考察。利用大鼠肝脏体外脂质自氧化反应结合紫外光谱法对中药提取物抑制脂质过氧化反应的能力进行评价,中药黄芩和槐米具有较强的抑制活性,富含黄酮类、生物碱类及木脂素类提取物表现出较好的脂质过氧化反应抑制作用,而皂苷类成分的活性相对较弱。以牛血清白蛋白/葡萄糖(果糖)体系为糖基化筛选模型,利用荧光光谱法对体外避光孵育反应体系进行研究,中药槐米、黄芩和知母表现出了较强的非酶糖基化反应抑制能力。由17种中药提取物抗氧化应激反应的结果分析,富含黄酮类成分中药的抗氧化应激作用强于生物碱类、木脂素类和萜类的,皂苷类中药的活性相对较弱;而体外综合活性较好的中药槐米和黄芩可作为下一步研究的主要对象。本研究为进一步的药理活性研究提供了一定的理论指导,为开发防治糖尿病并发症的中药新药奠定基础。     

皮肤晚期糖基化终末产物的自体荧光光谱特性

在分析晚期糖基化终末产物(Advanced glycation end products,AGEs)标准品三维荧光光谱和皮肤中AGEs自体荧光光谱的基础上,确定了皮肤中AGEs的最佳激发波长为370 nm.使用自行设计的皮肤荧光光谱检测装置对正常人和糖尿病患者皮肤AGEs的荧光光谱进行了检测,结果表明:在418,450...     

Inhibition of resveratrol glucosides (REs) on advanced glycation endproducts (AGEs) formation: inhibitory mechanism and structure-activity relationship

Abstract

The study on inhibitory effects of resveratrol glucosides (REs) on advanced glycation endproducts (AGEs) formation is still unmet. Herein, for the first time, the antiglycation activities of five REs in the fetal bovine serum proteins (FBS)/fructose system were evaluated, and its structure-activity relationship and antiglycation mechanism were further explored. These REs showed remarkable inhibition toward AGEs formation. Among them, Piceatannol-3'-O-glucoside (PG) exhibited highest antiglycation activity as reflected in approximately 80% inhibition of fluorescent AGEs at the concentration of 1.0?mM. The structure-activity relationship analysis indicated that glucoside attached to the B ring of resveratrol displays a superior antiglycation activity. Moreover, the results of antiglycation mechanism showed that the antiglycation activity of REs was proportional to their antioxidant capacity and methylglyoxal (MGO) trapping capacity. Therefore, the REs are promising candidates worthy of further exploration for preventing AGEs accumulation in vivo, thereby treating AGEs-associated diseases.     

Non-enzymatic modifications of proteins under high-pressure treatment

High-pressure treatment, e.g. used as an alternative method for food preservation, affects protein cross-linking and glycation reactions. These reactions were monitored by using mainly milk proteins in the absence and presence of different saccharides or dicarbonyl compounds. Without carbohydrates, protein cross-linking of casein is enhanced by pressure through the formation of dehydroalanine-derived lysinoalanine. A similar effect can be observed in wool, where pressure accelerates the formation of lanthionine. In contrast, saccharide or dicarbonyl compound-induced cross-linking is constrained by high pressures. Despite this, pressure is able to accelerate the degradation of sugar or carbonyl compounds, as could be seen by the measurement of their residual contents in the examined test preparations.     

Accurate mass measurements by Fourier transform mass spectrometry in the study of advanced glycation end products/peptides

The Maillard reaction occurring between sugars and amino groups is important in living systems. When amino groups belonging to protein chains are involved, the Maillard reaction has been invoked as responsible for protein cross-linking and the production of 'toxic' compounds. The reaction leads to the production of a heterogeneous group of substances, usually called advanced glycation end products (AGEs). Classical analytical approaches, such as spectroscopic (ultraviolet, fluorescence) and mass spectrometric (matrix-assisted laser desorption/ionization, liquid chromatography/electrospray ionization mass spectrometry) methods, have shown that the digestion mixture is highly complex. However, there are clear differences between the digestion mixtures of glycated and unglycated human serum albumin (HSA). In the former case, possible glycated peptides belonging to the AGE peptide class may be identified. Tandem mass spectrometric experiments on selected species seemed to be promising as regards structural information, but it was thought of interest to undertake the present investigation, based on liquid chromatography/electrospray ionization Fourier transform mass spectrometry, in order to obtain definitive results on their elemental composition. Using this approach, about 20 glycated peptides were detected and their possible structures were postulated by examining the known sequence of HSA.     

Antihyperglucolipidaemic and anticarbonyl stress properties in green,yellow and red sweet bell peppers (Capsicum annuum L.)

Effect of aqueous methanol extract of different colour sweet bell peppers (Capsicum annuum L.) on parameters of diabesity and carbonyl stress was analysed in vitro. Yellow pepper displayed significantly (p < 0.001) higher intestinal α-glucosidase inhibitory activity than green and red pepper. Porcine pancreatic lipase inhibitory activity was significantly (p < 0.01) high in yellow and red pepper than in green pepper. Green and red pepper inhibited vesperlysine-type advanced glycation end products (AGEs) more potently than yellow pepper; however, pentosidine-type AGEs were similarly inhibited by all three peppers. Yellow and red pepper inhibited lipid peroxidation more potently (p < 0.01) than green pepper. Total polyphenol content and free radicals scavenging activities in yellow and red bell peppers were higher than in green pepper. Total flavonoid content was high in green pepper than that present in yellow and red peppers. Green pepper displayed presence of proanthocyanins; however, oligomeric anthocyanins were detected in yellow and red peppers.     

Reactions of a modified lysine with aldehydic and diketonic dicarbonyl compounds: an electrospray mass spectrometry structure/activity study

The phenomenon known as non-enzymatic glycation is described as the reaction of reducing sugars with basic amino groups of proteins and nucleic acids, as well as with simple amines, without enzyme mediation. Non-enzymatic model glycation reactions that make use of low-molecular-weight compounds make an important contribution in the elucidation of glicated processes in vitro and in vivo. Four alpha-dicarbonyl compounds, aldehydic (glyoxal, methylglyoxal and phenylglyoxal) and ketonic (diacetyl), were reacted with the modified amino acid N(alpha)-acetyl-L-lysine (AcLys) in an attempt to establish structure/activity relationships for the reactivity of alpha-dicarbonyls with the amine compound. Electrospray ionization mass spectrometry (ESI-MS) combined with tandem mass spectrometry (MS/MS) and collision-induced dissociation (CID) was used to identify and characterize reagents, intermediates and reaction products. The formation of dicarbonyl-derived lysine dimers was observed exclusively. Especially, attention is drawn to alkyl- (asymmetrical dicarbonyl systems) and carboxyl- (glyoxal system) substituted imidazolium ions, at ring position 2.The main differences observed in the reactions studied were related to the reactivity with the diimine intermediate. This intermediate can react either with a non-hydrated dicarbonyl molecule at the aldehydic carbonyl, or with a mono-hydrated one at the ketonic carbonyl, particularly for asymmetrical dicarbonyls. For 2-carboxyl-substituted imidazolium ion (glyoxal reaction), besides the usual keto-enol rearrangement from the diol group, an alternative reaction pathway (proton abstraction) appears to contribute also for the imidazolium ring-closure process. Moreover, the formation of imidazolium ring structures can depend on several factors, namely, the presence (or absence) of electron donor substituents at the formed diol, the degree of stability of the new electrophile generated and/or the equilibrium concentration of the non- and mono-hydrated dicarbonyl forms in solution, the last being particularly important for asymmetrical dicarbonyls.The results reported reveal the complexity of reactivity as well as the diversity of imidazolium molecular structures.     

Rapid and simple method for determination of Nepsilon-(carboxymethyl)lysine and Nepsilon-(carboxyethyl)lysine in urine using gas chromatography/mass spectrometry

A new procedure was developed to determine in urine the concentrations of N(epsilon)-(carboxymethyl)lysine (CML) and N(epsilon)-(carboxyethyl)lysine (CEL), the major products of oxidative modification of glycated proteins, to assess levels of oxidative stress in physiological systems. The urine samples were acetonitrile-deproteinized, then derivatized by ethylchloroformate, and N(O,S)-ethoxycarbonyl ethyl esters of amino acids were analysed by isotope dilution gas chromatography/mass spectrometry. Recovery averaged 89%. Linearity was excellent (r = 0.998-0.999) in the 0.5-25 micromol/L range for CML and CEL. The limit of detection of this assay was 0.1 micromol/L (corresponding to 0.20 pmol of CML or CEL on column). Intra-day and inter-day precisions were likewise excellent, with relative standard deviations <4.63 and <6.15%, respectively. Accuracy of CML and CEL determination (15 micromol/L) was 2.9 and 5.9% of the estimated theoretical value. The time from obtaining the urine sample to determination of the concentration from the chromatographic peak was 80 min or less. This method is sensitive, reproducible, accurate, relatively cheap and very simple. It can be useful for laboratories involved in the diagnosis and monitoring of age-related chronic diseases.