Assessment of Autoxidation in Freeze-Dried Meats by a Fluorescence Assay

A simple and sensitive fluorescence-measuring technique was developed to assess extent of lipid oxidation in freeze-dried meats. Solvent extracts of reconstituted stored samples were assayed by fluorimetry. Spectra of “oxidized” meats show maximum excitation and emission wavelengths of λex = 350 and λem = 440 nm, respectively. At λem of 440 nm, “unoxidized” meats show three peaks in excitation spectrum at λex₁= 308, λex₂= 318 (max.), and λex₃= 350 nm. However, at λex of 350 nm, these samples show a peak at λem = 476 nm. The intensity ratio of λex₃ or λem over λex₂ are useful as sensitive and reliable “internal standards” of lipid oxidation. Presence of 100 ppm TBHQ (monotertiary butylhydroquinone), absence of oxygen, and compression of meat before freeze-drying, which protect against oxidation also result in corresponding reductions of these ratios.     

Effects of Vacuum Packaging, Glazing, and Erythorbic Acid on the Shelf-Life of Frozen White Hake and Mackerel

The quality changes during frozen storage of two underutilized species of fish: mackerel (Scomber scombrus), a fatty species, and white hake (Urophycis tenuis) a nonfatty, gadoid species, with or without a bag, vacuum, and/or erythorbic acid, were measured using the dimethy-lamine (DMA) test for hake and the thiobarbituric acid (TBA) test for mackerel, and by Instron deformation, expressible moisture, thaw drip, cook loss, and sensory evaluation for both species. Texture deterioration and lipid oxidation limited the shelf-life of hake and mackerel, respectively. Air (oxygen) prolonged the shelf-life of hake but lessened that of mackerel. Erythorbic acid accelerated the rate of texture deterioration in hake but inhibited the rate of lipid oxidation in mackerel.     

Influence of formaldehyde in the formation of fluorescence related to fish deterioration

 In previous studies fluorescence detection at different excitation/emission maxima during common fish processing has been used. A bathochromic shift towards higher wavelength maxima was observed and measured as the ratio between absorption at two of the maxima tested. This fluorescence ratio (δF) value correlates positively with fish damage. In the present work, the influence of formaldehyde (FA) on the value δF was studied. A model system was set up in which FA reacted at 30°C for 25 days with propylamine and fish muscle. It was observed that FA was less able to produce fluorescent compounds compared with common fish oxidation products that were also tested, i. e. propanal and hexanal. However, in the presence of both lipid oxidation aldehydes, the FA-containing mixtures led to a higher δF value. Model systems consisting of FA and fatty fish (sardine) muscle produced more fluorescence than FA and lean fish (cod), because of the formation of lipid oxidation compounds under the reaction conditions of the former systems. It is thus concluded that the presence of FA in a reacting medium enhances fluorescence formation, such that δF be can used as an accurate measure of fish damage. It is thought that measurement of δF in processes such as the freezing of gadoid fish, in which both FA and lipid oxidation are produced, could be of benefit. Received: 23 June 1997     

Lipid changes during long-term storage of canned tuna (Thunnus alalunga)

 In previous studies fluorescence detection at different excitation/emission maxima during common fish processing has been used. A bathochromic shift towards higher wavelength maxima was observed and measured as the ratio between absorption at two of the maxima tested. This fluorescence ratio (δF) value correlates positively with fish damage. In the present work, the influence of formaldehyde (FA) on the value δF was studied. A model system was set up in which FA reacted at 30°C for 25 days with propylamine and fish muscle. It was observed that FA was less able to produce fluorescent compounds compared with common fish oxidation products that were also tested, i. e. propanal and hexanal. However, in the presence of both lipid oxidation aldehydes, the FA-containing mixtures led to a higher δF value. Model systems consisting of FA and fatty fish (sardine) muscle produced more fluorescence than FA and lean fish (cod), because of the formation of lipid oxidation compounds under the reaction conditions of the former systems. It is thus concluded that the presence of FA in a reacting medium enhances fluorescence formation, such that δF be can used as an accurate measure of fish damage. It is thought that measurement of δF in processes such as the freezing of gadoid fish, in which both FA and lipid oxidation are produced, could be of benefit. Received: 23 June 1997     

Combined Application of Fluorescence Spectroscopy and Chemometrics Analysis in Oxidative Deterioration of Edible Oils

Combined methods of fluorescence spectrometry with chemometrics were used to monitor oxidation deterioration of edible oil. Synchronous and three dimensional fluorescence spectroscopy techniques were proposed for monitoring palm oil, camellia oil, sunflower oil and perilla oil during oven accelerated oxidation. Principal component analysis plot of fluorescence intensity (λex = 320–700 nm) clearly showed oxidative evolution of oils over heating time. High saturated or monounsaturated oils exhibited high regression coefficients between peroxide values and fluorescence intensity (R ²  = 0.973 for 400 nm in palm oil; R ²  = 0.956 for 370 nm in camellia oil). High diunsaturated oil exhibited high regression coefficient between nonpolar carbonyl compounds and fluorescence intensity (R ²  = 0.970 for 370 nm in sunflower oil). High triunsaturated oil exhibited high regression coefficient between p-anisidine value and fluorescence intensity (R ²  = 0.938 for 665 nm in perilla oil). In conclusion, Fluorescence spectroscopy is a rapid and green nondestructive method for oxidation monitoring. Differences of fatty acid compositions played key rules in formation of oxidation products and evolution of fluorescence spectra.     

Influence of storage time and temperature on lipid deterioration during cod (Gadus morhua) and haddock (Melanogrammus aeglefinus) frozen storage

Lean fish deterioration during frozen storage (−30 and −10 °C) for up to 1 year was studied by the assessment of lipid changes. Comparison between a formaldehyde (FA)-forming species (cod) and a non-FA-forming one (haddock) was carried out. Lipid damages were measured on the basis of free fatty acids (FFA), peroxide value (PV), thiobarbituric acid index (TBA-i) and fluorescent compounds. In both species at −30 °C, most lipid damage indices showed significant correlations with the storage time. However, at −10 °C, only the FFA and fluorescence detections provided significant correlations with the storage time. Comparison between the fish species showed higher lipid oxidation (PV and TBA-i) and hydrolysis (FFA content) in haddock than in cod at −10 °C; however, a higher fluorescence development was observed in cod at the same temperature. At −30 °C, little differences in lipid damage indices were detected between the two species. © 1999 Society of Chemical Industry     

Fluorescence due to interactions of oxidizing soybean oil and soy proteins

Soy proteins with soybean oil (9:1,w/w) were stored at 60°C to investigate the changes in intrinsic fluorescence during oxidation. The front-surface fluorescence of the oxidized samples showed excitation and emission maxima at 355 and 440 nm respectively. The fluorescent compounds were soluble in the organic layer of the chloroform–methanol (2:1,v/v). The solution fluorescence showed an excitation maximum at 365 nm and an emission maximum at 450 nm, and the intensity increased during storage. The interactions of oxidizing soybean oil and soy proteins also resulted in decreases of protein solubility, soluble protein hydrophobicity, and free amino groups of proteins. With an antioxidant (BHT) addition, the changes in fluorescence and in protein properties were inhibited. The intensity of the solution fluorescence showed high correlation with TBA value (r=0.968) and protein solubility (r=−0.979), which could serve as an indicator for oxidative deterioration of soy proteins and soybean oil systems.     

高效液相法检测几种米醋中的桔霉素含量

实验采用岛津ODS柱(Hypersil ODS 2,5μm,250mm×4.6),乙腈(色谱纯)∶超纯水为30∶70为流动相。流速0.8mL/min荧光检测波长为λex=331nm,λem=500nm。在0.01～100μg/mL浓度范围内,桔霉素浓度与荧光检测器响应值成良好线性关系,Y=78253x+37970(R2=0.9991)。样品加标回收率达到91.09%～101.2%。结果表明:米醋类食品较为安全,桔霉素含量最高的是米醋1,含有1.3361μg/mL桔霉素,含量最低的是米醋5,0.0158μg/mL。     

Inhibition of lipid damage in refrigerated salmon (Oncorhynchus kisutch) by a combined treatment of CO2 packaging and high-pressure processing

The effect of a previous combined treatment (CO₂-enriched modified atmosphere packaging, MAP and high-pressure processing, HPP, 150 MPa/5 min) on lipid stability of refrigerated (10 days/4 °C) salmon (Oncorhynchus kisutch) was studied. The following processing conditions were compared: B-0 (fish without MAP or HPP), B-1 (fish packaged under MAP and without HPP), B-2 (fish subjected to HPP without MAP) and B-3 (fish subjected to MAP and HPP). An inhibitory effect (P < 0.05) on lipid hydrolysis and oxidation was obtained by the presence of CO₂ in the packaging medium; values detected at day 10 for B-0 and B-1 fish were 80.72 and 49.61 (g free fatty acids, FFA, kg⁻¹ lipids), 6.14 and 2.81 (meq active oxygen kg⁻¹ lipids; peroxide value), 5.05 and 3.10 (mg malondialdehyde kg⁻¹ muscle), and 5.56 and 2.70 (fluorescence ratio), respectively. Furthermore, inhibition of lipid damage was observed for HPP alone; values detected at day 10 for B-2 fish were 76.24 (g FFA kg⁻¹ lipids) and 5.28 (meq active oxygen kg⁻¹ lipids). The lowest average values for lipid hydrolysis and oxidation were obtained in samples corresponding to the combined treatment (B-3 batch), differences being significant (P < 0.05) at day 10 for FFA (41.43 g kg⁻¹ lipids), peroxide (1.84 meq kg⁻¹ lipids) and fluorescence (2.50) values.     

High-Performance Liquid Chromatography with Fluorescence Detection for Simultaneous Analysis of Retinoids (Retinyl Palmitate,Retinyl Acetate,and Free Retinol) and α-, β-, γ-, and δ-Tocopherols in Foods

A method based on high-performance liquid chromatography with fluorescence detection for the simultaneous analysis of retinoids (vitamin A) and tocopherols (vitamin E) was developed. This method consists of an isocratic solution using hexane/ethyl acetate (85:15, v/v) as the mobile phase and fluorescence detection using a time program that sets the excitation (Ex) and emission (Em) wavelengths at adequate elution times for retinoids (Ex 342 nm, Em 476 nm) and tocopherols (Ex 298 nm, Em 325 nm), respectively. The separation of three retinoids (retinyl palmitate, retinyl acetate, and free retinol) and four tocopherol homologs was achieved with sufficient reproducibility and quantitative ability. Additionally, the necessity of saponification was considered. As a result, saponification was not used in this method because of the complexity of the procedure and the loss of free retinol. The retinoid and tocopherol contents of various foods were evaluated using the developed method. Our method could evaluate the retinoid and tocopherol contents of fish (eel, Anguilla japonica, and amberjack, Seriola dumerili) muscle and liver, roasted soybean (Glycine max) flour, and Japanese torreya seed (Torreya nucifera). Additionally, our method could be applied to the determination of retinoids and tocopherols not only in foods but also in supplements and cosmetics.     

Critical evaluation of non-thermal plasma as an innovative accelerated lipid oxidation technique in fish oil

Food products enriched with healthier unsaturated fatty acids are more sensitive to lipid oxidation, leading to an overall quality deterioration and the development of unwanted aroma properties. To evaluate the oxidative stability a wide range of techniques has been described in literature, of which most are thermally based. These unrealistic test conditions result in the induction of deviating oxidation chemistry compared to that observed during ambient storage. Non-thermal plasma technology is capable to generate a wide range of highly reactive oxidative species (e.g. atomic oxygen, hydroxyl radicals, singlet oxygen) while maintaining ambient temperatures. For the first time, a DBD-plasma jet (Ar/0.6% O₂) is used on fish oil samples as a faster and more realistic accelerated lipid oxidation method. This paper critically evaluates both a thermal as a non-thermal plasma based accelerated oxidation protocol using naturally aged fish oil as reference. Experiments were done using both virgin, as alpha-tocopherol-enriched fish oil samples. Secondary lipid oxidation volatiles were measured using HS-SPME–GC–MS. Both accelerated oxidation techniques induced the formation of typical lipid oxidation markers (e.g. 2-propenal, (E)-2-pentenal, heptanal), however in both cases significant differences were observed compared to the naturally aged fish oil. On the other side, non-thermal plasma correctly predicted an antioxidative effect when 1000 μg/g alpha-tocopherol was added to the fish oil, while thermally based tests resulted in the induction of prooxidative chemistry. Despite the differences with naturally aged fish oil, several non-thermal plasma characteristics (reactor configuration, gas feed mixture, power source, …) can be fine-tuned to evolve towards a technology that is capable to accelerate lipid oxidation in a highly realistic manner.     

荧光光度法测定鱼肉中的氧氟沙星残留

采用荧光光度法测定了鲫鱼、鳗鱼、鲤鱼和罗非鱼肌肉组织中的氧氟沙星药物残留量。样品用酸性乙腈提取,正己烷脱脂,水相在B-R(pH4.0)缓冲溶液中,激发波长λex294nm,荧光波长λem496nm下用荧光光度法测定其中的氧氟沙星含量,氧氟沙星的线性范围0.03～0.20μg/ml,相关系数r=0.9929,检出限为0.005μg/ml。回收率为76.0%～94.5%。日内变异系数为2.14%～6.50%,日间变异系数为3.02%～9.78%。     

Synchronous Front-Face Fluorescence Spectroscopy Coupled with Parallel Factors (PARAFAC) Analysis to Study the Effects of Cooking Time on Meat

ABSTRACT:  In this study, the potential of synchronous front-face fluorescence coupled with chemometrics has been investigated for the analysis of cooked meat. Bovine meat samples (thin slices of 5 cm diameter) taken from  Longissimus dorsi  muscle were cooked at 237 °C for 0, 1, 2, 5, 7, and 10 min under control conditions. Synchronous front-face fluorescence spectra were collected on meat samples in the excitation wavelength range of 250 to 550 nm using offsets (Δλ) of 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, and 160 nm between excitation and emission wavelengths. The synchronous fluorescence landscape containing 360 spectra was analyzed using PARAFAC. The best PARAFAC model presented 2 components since core consistency values for the first 2 components were 100% and the explained variance was 67.98%. The loading profiles of 1st and 2nd components had an optimal Δλ of 70 and 40 nm, respectively, allowing to determine the excitation (exc.) and emission (em.) maxima wavelengths of 1st (fluorescence band at about exc.: 340 to 400/em.: 410 to 470 nm, and peak at exc.: 468/em.: 538 nm) and 2nd (exc.: 294 nm/em.: 334 nm) components. As the loading profile of the 1st component of PARAFAC was assigned to Maillard-reaction products formed during cooking, the profile of the 2nd component corresponded with the fluorescence characteristics of tryptophan residues in proteins. Loadings and scores of the PARAFAC model developed from the synchronous fluorescence spectra enabled to get information regarding the changes occurring in meat fluorophores during cooking of meat at 237 °C from 0 to 10 min.     

Residues of Ethoxyquin and Ethoxyquin Dimer in Ocean-Farmed Salmonids Determined by High-Pressure Liquid Chromatography

The occurrence of residues of ethoxyquin (EQ) and ethoxyquin dimer (DM) from fish feeds in the liver and muscle of farmed salmon and steelhead was studied. For quantitative analysis of DM, the tissue was partially hydrolyzed with 50% NaOH, and then the lipid and antioxidant residues were extracted with hexane. After solvent removal and recovery into acetonitrile, DM was determined by reversed‐phase high‐pressure liquid chromatography (HPLC) with a fluorescence detector, with the excitation wavelength set at 360 nm and the emission wavelength at 440 nm. The mobile phase was acetonitrile:0.01M ammonium acetate (80:20, v:v). The EQ levels were very low, but the DM levels were higher. The recoveries from the spiked samples were 88 ± 4% for DM. Sample site determinations indicated that the DM residue level could be associated with depot fat except in the liver, where the tissue content of DM was 60% to 70% less. Similar results appear satisfactory for ethoxyquin extracted from fish oils.     

Browning of salted sun-dried fish

The lipid of traditional salted, sun-dried fish is highly susceptible to oxidation during processing and storage at tropical ambient temperatures (25–30°C), leading to browning and potential loss of nutritional and economic value of the product. Determinations of extractable fluorescence and soluble brown colour have been found to be relevant indicators of the high degree of lipid oxidation in such fish. Studies on model systems consisting of aerated fish oil and a range of components natural to fish confirmed that, at 25°C, the products of lipid oxidation reacted with phospholipids and amino acids to produce significant fluorescence. Similarly, proteins and amino acids interacted with lipid oxidation products to produce browning, although at 25°C, this only occurred in the presence of water. Temperatures above 50°C are required for the development of browning of aerated fish oil alone. The level of free amino acids in salted, sun-dried fish was found to decrease during storage which correlates with amino acid involvement in fluorescence and colour production. The fluorescence/colour can be related mechanistically to the development of lipid oxidation products and hence provides a realistic basis for their acting as indicators of extensive lipid oxidation.     

影响酪蛋白糖基化的因素分析

目的：通过建立酪蛋白-乳糖、酪蛋白-丙酮醛、酪蛋白-乙二醛3 种模拟体系,考察影响该体系晚期糖基化终末产物（advanced glycation end products,AGEs）形成的因素。方法：荧光光谱法（λex/λem=370 nm/440 nm）检测还原糖种类、温度、pH值、金属离子、时间和染料木黄酮对AGEs形成的影响;并分析各个影响因素的作用效果。结果：还原糖为核糖、pH 7.4、加工温度121 ℃和贮藏温度37 ℃条件下荧光性AGEs的产生量最多,金属离子Fe2＋和Ca2＋、9 mmol/L抑制剂染料木黄酮对荧光性AGEs的形成有抑制作用。结论：还原糖种类、pH值、温度、金属离子和抑制剂染料木黄酮均对3 种模拟体系中荧光性AGEs的形成有影响,其中,还原糖是影响酪蛋白糖基化生成荧光性AGEs最重要的影响因素,其次为时间、温度。     

Utilisation of a rapid technique based on front-face fluorescence spectroscopy for differentiating between fresh and frozen–thawed fish fillets

The potential of front-face fluorescence spectroscopy combined with chemometric tools was investigated to differentiate frozen–thawed from fresh fish. A total of 24 fish fillets, i.e., 12 fresh samples and 12 frozen–thawed samples, were investigated. Regarding the frozen–thawed samples, two speeds of freezing and thawing were tested (fast and slow) and for each condition, three whiting fillets were analysed. The fluorescence emission spectra of tryptophan (excitation: 290 nm, emission: 305–400 nm) and nicotinamide adenine dinucleotide (NADH) (excitation: 340 nm, emission: 360–570 nm) were recorded directly on samples. In a first step, the principal component analysis (PCA) was applied separately to the tryptophan or NADH fluorescence spectra. From the NADH spectra, PCA results showed a good discrimination between fresh and frozen–thawed fish samples. But it was not the case with the tryptophan fluorescence spectra. In a second step, factorial discriminant analysis (FDA) was applied to the first five principal components (PCs) of the PCA performed on the two data sets. Considering tryptophan fluorescence spectra, correct classification was observed for 62.5% and 70.8% of the calibration and validation spectra, respectively. A better classification was obtained from NADH fluorescence spectra since 100% correct classifications were obtained for the calibration and validation spectra. It was concluded that NADH fluorescence spectra may be considered as a promising probe for the reliable differentiation between frozen–thawed and fresh fish.     

Lipid nanoparticles based on omega-3 fatty acids as effective carriers for lutein delivery. Preparation and in vitro characterization studies

This study aimed at exploring the behavior of fish oil enriched with ω-3 fatty acids in order to obtain stable lipid nanocarriers (NLCs) with improved characteristics as effective delivery systems for lutein. The particle size of optimized lutein-NLCs was below 200 nm. The less ordered arrangement of lipid core revealed by scanning calorimetry and the high entrapment efficiency of 88.5% clearly indicated the appropriate role of fish oil in obtaining effective lipid nanocarriers. The evaluation of in vitro antioxidant activity has demonstrated a significant blocking effect of NLCs, scavenging up to 98% oxygen free radicals. The in vitro release profile has shown that NLCs are able to ensure a better, in vitro sustained release of lutein as compared to conventional nanoemulsions.     

Development of a rapid method based on front-face fluorescence spectroscopy for the monitoring of fish freshness

The intrinsic fluorescence of fish muscle was evaluated as a possible rapid and non-destructive method for the monitoring of fish freshness. The fluorescence emission spectra of aromatic-amino-acids+nucleic-acids (excitation: 250 nm, emission: 280–480 nm), tryptophan residues (excitation: 290 nm, emission: 305–400 nm) of proteins and NADH (excitation: 336 nm, emission: 360–600 nm) were recorded on cod, mackerel, salmon and whiting fillets at 1, 5, 8 and 13 days of storage. Principal component analysis and Mahalanobis distance method were applied to the spectral data sets. Considering the mackerel, the similarity map defined by the principal components 1 and 2 showed a discrimination of the aromatic-amino-acids+nucleic-acids spectra recorded after 1 day of storage versus the ones recorded after 5 and 8 days of storage according to the principal component 1. Similar trends were observed for all the fish species and all the fluorophores investigated. It appears that the intrinsic fluorescence spectra could be considered as fingerprints that may allow the discrimination between fresh and aged fish fillets.     

Classification and characterisation of Spanish red wines according to their appellation of origin based on chromatographic profiles and chemometric data analysis

Chromatographic profiles of wines have been used as a fingerprint for the discrimination of Spanish wines based on oenological practices. In order to extract information of different families of phenolic compounds, profiles of different UV-vis absorption wavelengths (280, 310, 370 and 520nm) and fluorescence (ex=260nm; em=360nm) were analysed. A total of thirteen phenolic compounds which allowed the discrimination of wines of three different Spanish appellations (Penedes, Rioja and Ribera del Duero) were selected by means of principal component analysis (PCA). Afterwards, these compounds were used to build partial least squares discriminant analysis (PLS1-DA and PLS2-DA) models which allowed the discrimination of wines according to their appellation with classification rates for independent test sets higher than 96% and 93% for PLS1-DA and PLS2-DA models respectively. Finally, characteristic compounds of each appellation were tentatively identified by means of liquid chromatography-mass spectrometry (LC-MS) analysis. Thus, ten out of thirteen compounds (i.e., gallic acid for Penedes, trans-coumaroyltartaric and trans-caffeoyltartaric acids for Rioja and myricetin for Ribera del Duero wines) have been proposed.