鱼源特定腐败菌及其应用研究进展

特定腐败菌(specific spoilage organisms,SSOs)是影响鱼肉腐败的重要因素之一。不同条件下鱼源SSOs不同,本文重点就SSOs的确定,即鱼源优势腐败菌筛选及其腐败能力分析的研究现状进行概述。在此基础上,对鱼源SSOs生长预测,靶向控制以延长鱼类鲜品货架期等应用研究进行了总结,旨在说明鱼源SSOs的研究价值,并对SSOs未来研究方向进行了展望。     

Nuclear DNA barcodes for cod identification in mildly-treated and processed food products

Gadoids are a group of fish with historical importance in the fishing industry. The high demand for cod is one of the reasons why cod products are often mislabelled, and numerous observations have been made on the replacement of Atlantic cod (Gadus morhua) by cheaper species or its illegal capture in contravention of fish quotas. Fish species identification is traditionally based on morphological features, but this may be difficult in case of heat-treated or processed products, or where the species look similar, as in the Gadoid group. DNA-based approaches (using either nuclear or mitochondrial DNA) are most commonly used in this case, due to their high specificity and to the high resilience of the target molecules to food processing techniques. In this article, we identified, using an automated screening approach, novel barcode regions and their associated primers in the nuclear genome, to be used for the efficient identification of Gadoids. The barcode regions were tested on official and commercial samples, raw or mildly treated products, like frozen, or salted, as well as pre-cooked complex mixtures and processed samples, using next-generation sequencing (NGS) technique. The method proposed could complement existing fish identification strategies in establishing an efficient framework to detect and prevent frauds along the food chain.     

Recent advances in shelf life prediction models for monitoring food quality

Each year, 1.3 billion tons of food is lost due to spoilage or loss in the supply chain, accounting for approximately one third of global food production. This requires a manufacturer to provide accurate information on the shelf life of the food in each stage. Various models for monitoring food quality have been developed and applied to predict food shelf life. This review classified shelf life models and detailed the application background and characteristics of commonly used models to better understand the different uses and aspects of the commonly used models. In particular, the structural framework, application mechanisms, and numerical relationships of commonly used models were elaborated. In addition, the study focused on the application of commonly used models in the food field. Besides predicting the freshness index and remaining shelf life of food, the study addressed aspects such as food classification (maturity and damage) and content prediction. Finally, further promotion of shelf life models in the food field, use of multivariate analysis methods, and development of new models were foreseen. More reliable transportation, processing, and packaging methods could be screened out based on real-time food quality monitoring.     

LSPR-based colorimetric biosensing for food quality and safety

Ensuring consistently high quality and safety is paramount to food producers and consumers alike. Wet chemistry and microbiological methods provide accurate results, but those methods are not conducive to rapid, onsite testing needs. Hence, many efforts have focused on rapid testing for food quality and safety, including the development of various biosensors. Herein, we focus on a group of biosensors, which provide visually recognizable colorimetric signals within minutes and can be used onsite. Although there are different ways to achieve visual color-change signals, we restrict our focus on sensors that exploit the localized surface plasmon resonance (LSPR) phenomenon of metal nanoparticles, primarily gold and silver nanoparticles. The typical approach in the design of LSPR biosensors is to conjugate biorecognition ligands on the surface of metal nanoparticles and allow the ligands to specifically recognize and bind the target analyte. This ligand–target binding reaction leads to a change in color of the test sample and a concomitant shift in the ultraviolet-visual absorption peak. Various designs applying this and other signal generation schemes are reviewed with an emphasis on those applied for evaluating factors that compromise the quality and safety of food and agricultural products. The LSPR-based colorimetric biosensing platform is a promising technology for enhancing food quality and safety. Aided by the advances in nanotechnology, this sensing technique lends itself easily for further development on field-deployable platforms such as smartphones for onsite and end-user applications.     

GCMS/MS 同时测定鱼样中39 种多溴联苯醚

目的 采用快速溶剂萃取(ASE)、系列固相萃取(SPE, 包括分散性固相萃取(DSPE)和串联SPE柱)和GCMS/MS, 建立一个便捷、高效测定鱼样中39种多溴联苯醚(PBDEs)的检测方法。方法 代表性鲈鱼样品(脂肪含量11%)经冷冻干燥后ASE提取, 然后采用DSPE去除脂肪和串联SPE柱净化, 最后采用GCMS/MS检测, 同位素内标法定量。结果 本方法的检测限(MDLs)为0.16~3.29 pg/g; 具有良好的线性范围(R2>0.998)、准确度(除BDE30为53%以外, 其他组分回收率为62%~1139%)和精密度(RSD<20%); 测定参考物质的结果均在标准值范围内, 与气相色谱磁场电场双聚焦高分辨质谱(GC/HRMS)测定结果一致。结论 本方法为鱼样中PBDEs检测提供了一个快速、简便和可靠的方法, 适用于对痕量PBDEs残留水平的食用鱼进行人体健康风险评价研究。     

Persistent organic pollutants in fish: biomonitoring and cocktail effect with implications for food safety

The impact of anthropogenic wastes of persistent organic pollutants (POPs) on the marine environment has increased in the last decades. POPs include polychlorinated biphenyls (PCBs) and organochlorine pesticides (OCs). To assess the levels of these POPs in the wild fish population, pelagic and benthopelagic predator fish species were selected as biomonitors. For detection and quantification of POPs in muscular tissues, a simple extraction through Accelerated-Solvent-Extraction (ASE) with an ‘in-line’ clean up purification approach was applied, followed by a GC–MS/MS analysis. Concentrations of sum DDT, sum HCH and endrin correlated with all PCB concentrations. Significant differences among fish species were found for all OCs and all PCBs except PCB 31 and 101. Blackspot seabream had the highest PCB concentrations; OCs were highest in tuna. Due to major concerns regarding fish population losses and the possible human chronic exposure to contaminated fish, studies addressing combined effects of multiple POPs (‘cocktail effect’) should be implemented. Our data motivate further experimental and observational studies in fish to define adequate baseline levels for cumulative human exposure and potential role of these contaminants for food safety.     

Tetrodotoxin detection and species identification of pufferfish in retail roasted fish fillet by DNA barcoding in China

This study identifies the pufferfish species and detects tetrodotoxin (TTX) in roasted fish fillet samples collected in Beijing, Qingdao and Xiamen, China. The cytochrome c oxidase I (COI) gene was used as the target gene for identification of the pufferfish species in the samples. Enzyme-linked immunosorbent assay (ELISA) screened the TTX levels in samples that had been detected as containing pufferfish by DNA barcode. A total of 125 samples were identified by DNA barcodes; 32 (26%) samples contained pufferfish composition and, among them, 26 (81%) were the highly toxic species Lagocephalus lunaris. All 32 samples containing the pufferfish composition were positive for TTX with levels ranging from 100 to 63 800 ng g^–1. Most of the 32 samples contained the highly toxic L. lunaris. Based on the results, we suggest that the monitoring of roasted fish fillet should be strengthened and the processing procedures should be standardised to minimise TTX poisoning caused by pufferfish.     

Comparison of PCR‐DGGE and PCR‐SSCP analysis for bacterial flora of Japanese traditional fermented fish products,aji‐narezushi and iwashi‐nukazuke

BACKGROUND: The bacterial flora of two Japanese traditional fermented fish products, aji‐narezushi (salted and long‐fermented horse mackerel (Trachurus japonicas) with rice) and iwashi‐nukazuke (salted and long‐fermented sardine (Sardinops melanostica) with rice bran), was analysed using non‐culture‐based polymerase chain reaction (PCR) denaturing gradient gel electrophoresis (DGGE) and culture‐based PCR single‐strand conformation polymorphism (SSCP) methods. RESULTS: Viable plate counts in aji‐narezushi and iwashi‐nukazuke were about 6.3–6.6 and 5.7–6.9 log colony‐forming units g^?1 respectively. In the PCR‐DGGE analysis, Lactobacillus acidipiscis was detected as the predominant bacterium in two of three aji‐narezushi samples, while Lactobacillus versmoldensis was predominant in the third sample. By the PCR‐SSCP method, Lb. acidipiscis and Lactobacillus plantarum were isolated as the predominant bacteria, while Lb. versmoldensis was not detected. The predominant bacterium in two of three iwashi‐nukazuke samples was Tetragenococcus muriaticus, while Tetragenococcus halophilus was predominant in the third sample. CONCLUSION: The results suggest that the detection of some predominant lactic acid bacteria species in fermented fish by cultivation methods is difficult. Copyright © 2010 Society of Chemical Industry     

A novel real-time polymerase chain reaction method for the qualitative detection of pistachio in food

In order to provide an appropriate method for the detection of pistachio (Pistacia vera) in food products, a novel real-time PCR was developed. The pistachio-specific primers and the TaqMan fluorogenic probe were designed to target the internal transcribed spacer between 18S ribosomal RNA and 5.8S ribosomal RNA genes. Using dilutions of the pistachio DNA, the intrinsic detection limit of the method was determined to be 0.012 pg. At specificity testing, the method was positive for 11 pistachio varieties and negative for 26 plant and animal species used in food industry. A detection limit of 0.0004% (w/w) was determined for pistachio nuts in model pastry. Practical applicability of the elaborated method was tested by the analysis of 44 food samples, out of which 7 food products were identified as containing undeclared pistachio. The developed real-time PCR may be utilized for sensitive and selective detection of pistachio in food products.     

A novel real-time polymerase chain reaction method for the detection of pecan nuts in food

A real-time PCR-based method for the detection of the pecan (Carya illinoiensis) component in food is described. The method consists of DNA isolation by chaotropic solid phase extraction and the subsequent PCR with pecan-specific primers and a TaqMan fluorescent probe. The primers and the probe are targeted to the putative gene for allergenic vicilin-like seed storage protein of pecan. The method was positive for 10 pecan varieties and negative for all other tested plant materials used in food industry, including walnut. The intrinsic detection limit of the method was 1 pg pecan DNA which corresponds to 1.2 haploid genome copies. Using a series of model pastry samples with defined pecan contents, a practical detection limit of 0.01% (w/w) pecan was estimated. Practical applicability of the PCR method was tested by the analysis of 13 food samples; no discrepancies between the declared and detected pecan contents were found. The presented PCR method is useful for sensitive and selective detection of pecans in food samples and can be performed in one working day.     

A novel real-time polymerase chain reaction method for the detection of macadamia nuts in food

A real-time PCR-based method for the detection of macadamia nuts (fruits of Macadamia integrifolia or M. tetraphylla or their hybrids) in food products is described. The method consists of DNA isolation by chaotropic solid phase extraction and subsequent PCR with macadamia-specific primers and a TaqMan fluorescent probe. The primers and the probe were targeted to the gene encoding for vicilin precursor. The method was positive for M. integrifolia and M. tetraphylla and negative for 16 other plant species used in food industry, including peanuts, walnuts, hazelnuts, almonds, pistachio nuts, cashew nuts, Brazil nuts, and chestnuts. The DNA-based detection limit of the method was 1.45 pg. Using a series of model samples with defined macadamia nut contents, a practical detection limit of 0.02% (w/w) macadamia nuts was determined. Practical applicability of the PCR method was tested by the analysis of 14 confectionery samples. For all of the samples, results conforming to the labeling were obtained. The presented PCR method is useful for relatively fast, highly selective, and moderately sensitive detection of macadamia nuts in food samples.     

Optimization of the procedure for the determination of polycyclic aromatic hydrocarbons and their derivatives in fish tissue: Estimation of measurements uncertainty

Three alternative procedures were employed for the isolation of polycyclic aromatic hydrocarbons (PAHs; 15 of 16 US EPA priority pollutants and benzo[e]pyrene), their methyl-derivatives and sulphur analogues from fish tissue: (1) Soxhlet extraction, (2) batch extraction enhanced by sonication, and (3) saponification of the sample followed by re-extraction of analytes into hexane. Soxhlet extraction using hexane-acetone (1:1, v/v) was the most efficient extraction technique, with analyte recoveries in the range 70-108%. Within optimization of the clean-up step, several types of gel permeation chromatography (GPC) systems were tested: two types of polystyrene divinylbenzene copolymer gels (PSDVB), both 'soft' gel type (Bio-Beads S-X3) and 'rigid' gels type (PL gel and Envirogel) in combination with various mobile phases were compared. Bio-Beads S-X3 and mobile phase chloroform were the most appropriate for purifying of crude extracts before the final determinative step. High-performance liquid chromatography with fluorimetric detection (HPLC/FLD) was used for identification and quantification of PAHs in purified fish extracts. The uncertainties of PAHs measurements were estimated by employing two alternative approaches. Both provided similar results: the expanded uncertainties obtained for individual PAHs by the 'top-down' approach were in the range 9-53%, their values resulting from application of the 'bottom-up' approach were in the range 16-52%.     

A novel real-time polymerase chain reaction (PCR) method for the detection of hazelnuts in food

A real-time PCR (polymerase chain reaction)-based method for the detection of hazelnuts (nuts of Corylus avellana or C. maxima) in confectionery and bakery products is described. The method consists of DNA isolation by chaotropic solid phase extraction and the subsequent PCR with hazelnut-specific primers and a TaqMan fluorescent probe. The primers and the probe are targeted to the hsp1 gene encoding for a low molecular weight heat-shock protein. The method was positive for five hazelnut varieties approved in Slovakia and negative for all other tested plant materials used in food industry including peanuts, walnuts, almonds, pistachio nuts, cashews and chestnuts. The intrinsic detection limit of the method was 13 pg hazelnut DNA, which corresponds to approximately 27 genome equivalents (1C). Using a series of model pastry samples with defined hazelnut contents, a practical detection limit of 0.01% (w/w) hazelnut was determined. Practical applicability of the PCR method was tested by the analysis of 20 food samples (confectionery and bakery products) along with ELISA. For all of the food samples, identical results were obtained by both methods, which conformed to the labelling. The presented PCR method is useful for sensitive and selective detection of hazelnuts in food samples and can be performed in one working day.     

A novel real-time polymerase chain reaction (PCR) method for the detection of walnuts in food

A real-time polymerase chain reaction (PCR)-based method for the detection of the walnut (Juglans regia) component in food is described here. The method consists of DNA isolation by chaotropic solid phase extraction and the subsequent PCR with walnut-specific primers and a TaqMan fluorescent probe. The primers and the probe are targeted to the jug r2, a major allergen gene of walnut. The method was positive for 8 varieties of walnut and negative for all other tested plant materials used in food industry, including pecan nuts. The intrinsic detection limit of the method was 0.24 ng walnut DNA. Using a series of model pastry samples with defined walnut contents, a practical detection limit of 0.01% walnut content was estimated. Practical applicability of the PCR method was tested by the analysis of 13 food samples (bakery and confectionery products), out of which two cakes were found to contain walnuts although they were not adequately labelled. The presented PCR method is useful for sensitive and selective detection of walnuts in food samples and can be performed in one working day.     

New monitoring approach for metabolic dynamics in microbial ecosystems using stable-isotope-labeling technologies

We have developed a new approach for monitoring the metabolic dynamics in microbial ecosystems using a combination of DNA fingerprinting and metabolome analysis based on stable-isotope-labeling technologies. Stable-isotope probing of DNA (DNA-SIP) has been used previously for the evaluation of cross-feeding in microbial communities. For the development and validation of our monitoring approach, fecal microbiota were analyzed with stable-isotope-labeled glucose used as the sole carbon source. In order to link the metabolic information and the microbial variability, we performed metabolic–microbial correlation analysis based on nuclear magnetic resonance (NMR) profiles and denaturing gradient gel electrophoresis (DGGE) fingerprints, which successfully identified the glucose-utilizing bacteria and their related extracellular metabolites. Moreover, our approach revealed information regarding the carbon flux, in that the “first” wave of extracellular metabolites secreted by the glucose-utilizing bacteria were incorporated into the “secondary” group of substrate-utilizing bacteria, and that this “secondary” group further produced their own secondary metabolized substrates. Thus, this approach is a powerful tool for monitoring the metabolic dynamics in microbial ecosystems and allows for the tracking of the carbon flux within a microbial community.     

Macroporous resin purification of grass carp fish (Ctenopharyngodonidella) scale peptides with in vitro angiotensin-I converting enzyme (ACE) inhibitory ability

The adsorption dynamics and thermodynamics of grass carp fish scale peptides (FSPs) onto non-polar macroporous resins (MARs), DA201-C, have been investigated. The adsorption of FSPs was affected by time, pH and peptide concentration. The adsorption process followed the Langmuir adsorption isotherm, and was endothermic (ΔH < 43 kJ/mol). The predominant force in adsorption of FSPs onto DA201-C was hydrophobic. Depending on this force, the dynamic adsorption and gradient desorption results showed that DA201-C resins were good at desalting and enriching peptides with higher contents of hydrophobic amino acids, and these peptides had higher ACE inhibitory capabilities in vitro. The lowest concentration at which the eluted fraction possessed half of its original ACE activity (IC₅₀) was 0.13 mg/ml. The results indicated that fish scale peptides produced showed good ACE-inhibitory effect in vitro and fish scales are a good source of peptides with in vitro ACE inhibitory activity.     

A sensitive solid-phase microextraction/gas chromatography-based procedure for determining pentachlorophenol in food

A new method for the determination of pentachlorophenol (PCP) in different foods was developed using capillary gas chromatography (GC) and microwave induced-plasma atomic emission spectrometry (MIP-AED). The analyte is first derivatized and then extracted and pre-concentrated by solid-phase microextraction (SPME) in headspace (HS) mode. A clear matrix effect was found for the different samples investigated, so that standard addition was required for quantification. Detection limits of 0.03-6.0 ng g^-1 were obtained, depending on the sample analysed. The method gave recoveries of 81-109% from spiked samples. Concentration levels of PCP ranged from 0.3 to 1.5 ng g^-1 were found in honey, but no PCP was detected in other samples.     

Dioxins and polychlorinated biphenyls (PCBs) in fish oil dietary supplements: Occurrence and human exposure in the UK

Commercially available fish oil supplements sourced from retail outlets in the UK, as well as by mail order, were surveyed in 2000-02 for dioxin (PCDD/Fs) and polychlorinated biphenyl (PCB) content. Sampled products were representative of market share. The WHO-TEQ values for these products ranged from 0.18 to 8.4 ng kg^-1 for ΣPCDD/F and from 1.1 to 41 ng kg^-1 for Σ dioxin-like PCBs. The results suggest a downward trend in the levels of dioxins in fish oil supplements over the last decade, since levels for similar products ranged from 0.3 to 10 ng kg^-1 for ΣPCDD/F WHO-TEQ in 1996. Levels of ICES (International Council for the Exploration of the Seas) 7 PCBs in the current study ranged from 8.3 to 267 µg kg^-1. Subsequent to this survey, European Union legislation has been introduced that includes a maximum limit of 2 ng kg^-1 WHO-TEQ for dioxins in fish oil products for human consumption. Twelve of the 33 products reported here would have exceeded this limit. Negotiations are in progress to incorporate dioxin-like PCBs into the European Union regulations. When manufacturer-recommended doses were applied to the observed levels, the estimated upper bound human exposure to dioxins and dioxin-like PCBs from dietary intake of these products ranged from 0.02 to 7.1 pg WHO-TEQ kg^-1 body weight day^-1 for adults and from 0.02 to 10 pg WHO-TEQ kg^-1 body weight day^-1 for schoolchildren. This level rises to 1.8-8.9 pg WHO-TEQ kg^-1 body weight day^-1 for adults and 1.4-14 pg WHO-TEQ kg^-1 body weight day^-1 for schoolchildren when combined with the average exposure from the whole diet in 1997. Again, subsequent to this survey, the Committee on Toxicity of Chemicals in Food, Consumer Products and the Environment (COT) revised the UK tolerable daily intake (TDI) for mixtures of dioxins and dioxin-like PCBs from 10 to 2 pg WHO-TEQ kg^-1 body weight day^-1. This is in line with the tolerable weekly intake (TWI) of 14 pg WHO-TEQ kg^-1 body weight set by the Scientific Committee on Food (SCF).