Zwitterionic hydrophilic interaction solid-phase extraction and multi-dimensional mass spectrometry for shotgun lipidomic study of Hypophthalmichthys nobilis.

Zwitterionic hydrophilic interaction liquid chromatography (ZIC-HILIC) material was used as solid-phase extraction sorbent for purification of phospholipids from Hypophthalmichthys nobilis. The conditions were optimized to be pH 6, flow rate 2.0mL·min(-1), loading breakthrough volume ⩽5mL, and eluting solvent 5mL. Afterwards, the extracts were analyzed by multi-dimensional mass spectrometry (MDMS) based shotgun lipidomics; 20 species of phosphatidylcholine (PC), 22 species of phosphatidylethanoamine (PE), 15 species of phosphatidylserine (PS), and 5 species of phosphatidylinositol (PI) were identified, with content 224.1, 124.1, 27.4, and 34.7µg·g(-1), respectively. The MDMS method was validated in terms of linearity (0.9963-0.9988), LOD (3.7ng·mL(-1)), LOQ (9.8ng·mL(-1)), intra-day precision (<3.64%), inter-day precision (<5.31%), and recovery (78.8-85.6%). ZIC-HILIC and MDMS shotgun lipidomics are efficient for studying phospholipids in H. nobilis.

Protein-peptide nutritional material prepared from surimi wash-water using immobilized chymotrypsin-trypsin.

BACKGROUND: In the production process of surimi, large quantities of wastewater are produced. Thus it would be interesting to develop an efficient protocol for the recovery of protein from hairtail surimi wash-water. RESULTS: A technique involving the use of immobilized chymotrypsin-trypsin (I-CT) was developed, providing a practical method for the preparation of protein-peptide nutritional material (PPNM). Under optimized reaction conditions, the recovery rate of nitrogen of surimi wash-water was measured as 98.3 ± 2.9%. Nutritional evaluation of the protein-peptide fraction demonstrated that it contained all essential amino acids (EAA) for humans, accounting for 44.1% of the total amino acid (TAA) content, which was determined to be 78.2 g per 100 g dry matter. The essential amino acid index (EAAI) and biological value (BV) were 101.7 (>95) and 76.7 respectively. A wide range of volatile flavor compounds (>50), including aldehydes, ketones, alcohols, hydrocarbons and heterocyclic compounds, were identified in PPNM by gas chromatography/mass spectrometry (GC/MS) analysis. CONCLUSION: An efficient and practical protocol for the recovery of protein from hairtail surimi wash-water has been developed. The PPNM prepared in this work could be used as a nutraceutical and as an ingredient of functional foods. © 2016 Society of Chemical Industry.

Lipidomic profiling of dried seahorses by hydrophilic interaction chromatography coupled to mass spectrometry.

Dried seahorse is a precious raw food material for cooking soups. In this study, a lipidomics strategy using the techniques of solid-phase extraction (SPE) and hydrophilic interaction chromatography-tandem mass spectrometry (HILIC-QTOF/MS) was developed for extraction, visualization, and quantification of phospholipids in dried seahorses. The parameters of SPE were optimized, and 1 mL of sample and chloroform/methanol (1:2, v/v) were found to be the best loading volume and eluting solvent, respectively. Afterwards, each phospholipid class was successfully separated on a HILIC column and analyzed by mass spectrometry. A total of 50 phospholipid molecular species were identified and determined, including 15 phosphatidylcholines (PCs), 14 phosphatidylethanolamines (PEs), 12 phosphatidylinositols (PIs) and 9 phosphatidylserines (PSs). In comparison to previously methods, this strategy was robust and efficient in extraction, characterization, and determination of phospholipids. The dried seahorse was found to contain large amounts of polyunsaturated fatty acyl phospholipids which are beneficial to human health.

Differentiation and classification of bacteria using vancomycin functionalized silver nanorods array based surface-enhanced Raman spectroscopy and chemometric analysis.

Twenty seven different bacteria isolates from 12 species were analyzed using intrinsic surface-enhanced Raman scattering (SERS) spectra with recently developed vancomycin coated silver nanorod (VAN AgNR) substrates. The VAN AgNR substrates could generate reproducible SERS spectra of the bacteria with little to no interference from the environment or bacterial by-products as compared to the pristine substrates. By taking advantage of the structural composition of the cellular wall which varies from species to species, the differentiation of bacterial species is demonstrated by using chemometric analyses on those spectra. A second chemometric analysis step within the species cluster is able to differentiate serotypes and strains. The spectral features used for serotype differentiation arises from the surface proteins, while Raman peaks from adenine dominate the differentiation of strains. In addition, due to the intrinsic structural differences in the cell walls, the SERS spectra can distinguish Gram-positive from Gram-negative bacteria with high sensitivity and specificity, as well as 100% accuracy on predicting test samples. Our results provide important insights for using SERS as a bacterial diagnostic tool and further guide the design of a SERS-based detection platform.

Detection of polycyclic aromatic hydrocarbons from cooking oil using ultra-thin layer chromatography and surface enhanced Raman spectroscopy.

The combinatory use of on-chip ultra-thin layer chromatography (UTLC) and surface enhanced Raman spectroscopy (SERS) has been demonstrated to detect mixtures of hazardous food contaminants, polycyclic aromatic hydrocarbons (PAHs), from cooking oil samples. After a simple acetonitrile extraction step, the organic phase was directly applied to the chemically modified SERS-active silver nanorod (AgNR) substrate without further cleanup, and subjected to UTLC separation. The spectral interference from co-extracted oil residues was mitigated by UTLC, and the SERS detection limits were found to be equivalent to or lower than those found in standard PAH solutions. The interference from the oil matrix was also quantitatively assessed, and the PAH extraction procedure was optimized for UTLC-SERS. This UTLC-SERS strategy provides a simple but effective method for on-chip sample cleanup and sensing from complex sample matrices.

Detection of metronidazole and ronidazole from environmental samples by surface enhanced Raman spectroscopy.

In this study, the surface enhanced Raman spectra (SERS) of two prohibited veterinary drugs, metronidazole (MNZ) and ronidazole (RNZ), have been acquired, and compared to the theoretically calculated spectra using density function theory (DFT). The experimental Raman and SERS spectra of MNZ and RNZ exhibit high resemblance with the DFT calculations. SERS detection of MNZ and RNZ from standard solutions as well as real environmental samples (tap, lake, swamp waters and soil) was performed on highly sensitive and reproducible silver nanorod array substrates. The limits of detection for MNZ and RNZ are 10 and 1 µg/mL in methanol and ultra-pure water, respectively, and 10-50 µg/mL in the environmental samples. The SERS-based method demonstrates its potential as a rapid, simple, and inexpensive means for the onsite screening of banned antibiotics from the aquatic and sediment environments, with minimal requirement for sample pretreatment.

Detection and differentiation of foodborne pathogenic bacteria in mung bean sprouts using field deployable label-free SERS devices.

Vancomycin functionalized silver nanorod arrays substrates were used to obtain the surface enhanced Raman scattering (SERS) signals of six foodborne pathogenic bacteria in mung bean sprouts samples using both a portable and a handheld Raman system. The silver nanorod arrays substrates were optimized to facilitate quantitative, rapid, and sensitive detection of Salmonella enterica serotype Anatum, Salmonella enterica serotype Cubana, Salmonella enterica serotype Stanley, Salmonella enteritidis, Escherichia coli O157:H7, and Staphylococcus epidermidis. Substrate optimization was achieved by varying the nanorod length and vancomycin incubation concentration. By combining these substrates with a two-step filtration process we found that the foodborne pathogenic bacteria used in this study can be identified in mung bean sprouts with a limit of detection as low as 100 CFU ml(-1) in less than 4 h using both portable and handheld Raman systems. The results show that SERS spectra can be used to differentiate between bacterial species and serotypes when chemometric methods are employed. The low detection limit and rapid detection time of this biosensing platform for foodborne pathogenic bacteria could be a valuable field detection method for the fresh produce and food processing industries.

Influence of alanine uptake on Staphylococcus aureus surface charge and its susceptibility to two cationic antibacterial agents, nisin and low molecular weight chitosan.

Low molecular weight chitosan (LMWC) and nisin, recognized as cationic antibacterial agents (CAAs), inhibit bacterial growth by interacting with the anionically charged cell wall. In this study, alanine uptake significantly reduced the anionic cell surface charge, as determined by the zeta potential (ZP) measurements, of Staphylococcus aureus, resulting from the incorporation of d-alanine into the cell wall. Minimum inhibitory concentration (MIC) tests and growth inhibition curves revealed that LMWC and nisin possessed inverse antibacterial activity against three strains of S. aureus, depending on the strains' net charge. A twofold reduction in the MIC value of nisin was obtained against S. aureus, inoculated in a 1.0% d- or l-alanine-augmented trypticase soy broth medium. A flocculation test demonstrated that neutralizing the anionic surface charge using d-alanine reduced the adsorption of S. aureus onto LMWC. Furthermore, the reduced surface net charge could enhance the colonization capacity of S. aureus on glass.

The surface-enhanced Raman spectra of aflatoxins: spectral analysis, density functional theory calculation, detection and differentiation.

High-quality surface-enhanced Raman scattering (SERS) spectra of aflatoxin (AF) B(1), B(2), G(1) and G(2) have been acquired using silver nanorod (AgNR) array substrates fabricated by oblique angle deposition method. Significant vibrational peaks are identified on the argon plasma-cleaned substrates, and those peaks agree very well with the Raman spectra calculated by density function theory (DFT). The concentration-dependent SERS detection is also explored. The relationship between the concentration (C) of different AFs and the SERS intensity (I) of the Raman peak at Δν = 1592 cm(-1) is found to follow the general relationship I = AC(α), with α ranging from 0.32 to 0.46 for the four AFs. The limits of detection (LODs) reach 5 × 10(-5) mol L(-1) for AFB(1), 1 × 10(-4) mol L(-1) for AFB(2), and 5 × 10(-6) mol L(-1) for both AFG(1) and AFG(2) in bulk solution, or 6.17 × 10(-16) mol/1.93 × 10(-4) ng of AFB(1), 1.23 × 10(-15) mol/3.88 × 10(-4) ng for AFB(2), 6.17 × 10(-17) mol/2.03 × 10(-5) ng for AFG(1), and 6.17 × 10(-17) mol/2.04 × 10(-5) ng for AFG(2) per laser spot. Principal component analysis (PCA) is used to successfully differentiate these four different kinds of AFs at different concentrations up to their detection limits. The LODs obtained from PCA agree with the LODs obtained by using peak fitting method. With such a low detection limit and outstanding differentiation ability, we prove the possibility of utilizing the SERS detection system as a platform for highly sensitive mycotoxin detection.

On-chip ultra-thin layer chromatography and surface enhanced Raman spectroscopy.

We demonstrate that silver nanorod (AgNR) array substrates can be used for on-chip separation and detection of chemical mixtures by combining ultra-thin layer chromatography (UTLC) and surface enhanced Raman spectroscopy (SERS). The UTLC-SERS plate consists of an AgNR array fabricated by oblique angle deposition. The capability of the AgNR substrates to separate the different compounds in a mixture was explored using a mixture of four dyes and a mixture of melamine and Rhodamine 6G at varied concentrations with different mobile phase solvents. After UTLC separation, spatially-resolved SERS spectra were collected along the mobile phase development direction and the intensities of specific SERS peaks from each component were used to generate chromatograms. The AgNR substrates demonstrate the potential for separating the test dyes with plate heights as low as 9.6 µm. The limits of detection are between 10(-5)-10(-6) M. Furthermore, we show that the coupling of UTLC with SERS improves the SERS detection specificity, as small amounts of target analytes can be separated from the interfering background components.

Identification of causative agents and species in shrimp implicated in a food poisoning case in Taiwan.

The possible causative agent and shrimp species involved in a bait shrimp poisoning case that occurred in northern Taiwan was determined. Because the patient's symptoms were similar to those caused by boric acid and slightly similar to those caused by sulfite, the concentrations of boric acid and sulfite (as sulfur dioxide) in the patient's vomitus and in shrimp collected from bait stores and markets were analyzed. The concentration of boric acid was 36.8 to 37.1 mg/g in the patient's vomitus, 1.4 to 3.8 mg/g in shrimp meats obtained from bait stores, and not detectable (less than 0.001 mg/g) in shrimp meat obtained from commercial markets. No significant differences in sulfur dioxide concentrations (0.067 to 0.088 mg/g) were found in patient's vomitus and the shrimp meat from both bait stores and commercial markets. A fragment of the cytochrome b gene (∼406 bp) was amplified by PCR using a pair of primers (UCYTB151F and UCYTB270R) from shrimp meat of two species and the vomitus. The vomited shrimp was identified as Parapenaeus fissuroides on the basis of gene sequencing and restriction fragment length polymorphism patterns after treatment with endonuclease Alu I. Based on the patient's symptoms and analytical data, we concluded that boric acid at toxic levels had been illegally added to the bait shrimp P. fissuroides.

Fate of Staphylococcus aureus, Salmonella enterica serovar typhimurium, and vibrio vulnificus in raw oysters treated with chitosan.

The fate of Staphylococcus aureus, Salmonella enterica serovar Typhimurium, and Vibrio vulnificus in oysters treated with chitosan was investigated. Three concentrations (0.5, 1.0, and 2.0%) of chitosan in 0.5% hydrochloric acid were prepared and coated onto raw oysters, which were then stored at 4 degrees C for 12 days. Untreated oysters and oysters coated with 0.5% hydrochloric acid without chitosan were used as controls. S. aureus cells were most sensitive to 2.0% chitosan followed by 0.5 and 1.0%. In general, chitosan treatment of oysters produced a decline in the population of S. aureus by 1 to 4 log CFU/ml compared with the untreated control. Chitosan treatment had no influence on the reduction of Salmonella Typhimurium over the 12-day storage period; inhibition of Salmonella Typhimurium growth was similar in both the control samples and the chitosan-treated samples. However, time of storage had a major effect on the survival of Salmonella Typhimurium on oysters. Neither time nor chitosan concentration had a significant effect on the growth of V. vulnificus during storage. All treatments were similar in inhibiting V. vulnificus growth.

Influence of hydroxypropyl methylcellulose edible coating on fresh-keeping and storability of tomato.

The effect of application of cellulose-based edible coating, hydroxypropyl methylcellulose (HPMC) to mature-green tomatoes on the firmness and color was investigated. Tomatoes were stored at 20 degrees C for up to 18 days. Firmness decreased as storage time increased in all treatments. However, application of HPMC edible coating delayed softening of tomatoes during 18 days of storage at 20 degrees C. At days 7, 13 and 18, the firmness of tomatoes coated with HPMC was significantly (P < or = 0.05) greater than the firmness of uncoated tomatoes. The study also confirmed that HPMC coatings could significantly (P < or = 0.05) delay the changes in color of tomatoes stored at 20 degrees C. The ripening of tomatoes from the pink stage to the red stage was successfully retarded. HPMC coating could extend the shelf life of fresh tomatoes. The retardation of the rate of loss of firmness could reduce the economic loss that would result from spoilage by mechanical injury during transportation of tomatoes.

Changes in Dieldrin and p,p'-DOE Residues Following Cooking of Channel Catfish.

The influence of cooking (frying, baking, and smoking) on dieldrin and 1,1-dichloro-2,2-bis[4-chlorophenyl]ethylene (DDE) residues in treated channel catfish ( Ictalarus punctatus ) was determined. Dieldrin and DDE were significantly reduced (P < 0.05) during cooking of catfish by 50 to 65% (dry basis) and 50 to 80%, respectively. Smoking resulted in maximum reduction (82%) of DDE residues, while baking resulted in the least reductions for both dieldrin (50%) and DDE (50%) when compared to the other preparation methods.

Toxin Production by Clostridium botulinum Type E in Packaged Channel Catfish.

Channel catfish were inoculated with 3 to 4 log spores/g of a mixed pool of four strains of C. botulinum type E (Beluga, Minnesota, G21-5, and 070) and were packaged with an oxygen-permeable overwrap, in an oxygen-barrier bag with a modified atmosphere of CO2-N2 (80:20) or in a master bag with the same modified atmosphere. Packaged fish were stored at either 4°C and sampled at intervals over 30 days or at 10°C and sampled at intervals over 12 days. An additional master bag treatment in which overwrap-packaged catfish was stored first at 4°C, then removed from the master bags and stored at 10°C, was sampled at intervals over 18 days. Toxin production was evaluated using the mouse bioassay. Aerobic psychrotrophic and anaerobic populations were enumerated, and product spoilage characteristics were noted. Under abusive storage conditions of 10°C, there was no difference among the potential for toxin production in the packaged fish, with botulinum toxin detected on fish from each package type by day 6. At 4°C, toxin production was detected on day 9 in the overwrapped packages, while it was on day 18 in the modified atmosphere packaging. No toxin was found in the master bags held continually at 4°C. Toxin was detected on day 18 from samples initially held at 4°C in the master bag and subsequently held at 10°C. Spoilage preceded toxin production for samples stored at 4°C for each type of packaging. At 10°C, spoilage and toxin detection times coincided.

Growth and Production of Toxin of Clostridium botulinum Type E in Rainbow Trout under Various Storage Conditions.

Rainbow trout ( Oncorhynchus mykiss ) were inoculated with 3 to 4 1og10 spores per g of fish of a mixed pool of four strains of Clostridium botulinum type E (Beluga, Minnesota, G21-5, and 070). The trout were vacuum-skin packaged with either oxygen-barrier or oxygen-permeable films. Trout packaged with oxygen-permeable film were stored at 4°C for 21 days, while trout packaged with oxygen-barrier film were stored either at 4°C for 21 days or at 10°C for 15 days. Storage at 10°C was used to simulate commercial temperature abuse. Clostridium botulinum outgrowth was determined by a most probable-number (MPN) method using (tryptone peptone yeast extract glucose trypsin) anaerobic broth. Toxin production was evaluated using a mouse bioassay. Psychrotrophic and anaerobic populations increased with time regardless of packaging type. After 6 days at l0°C, botulinum toxin was detected in the packaged trout; however, the fish was noticeably spoiled before that time. No botulinum toxin was detected in trout packaged with either barrier or permeable films and stored at 4°C for 21 days, although the product was considered spoiled by day 12.

Bacterial Pathogens and Indicators in Catfish and Pond Environments.

Channel catfish ( Ictalurus punctatus ) fed a diet containing 26 or 38% protein with restricted and satiety feeding methods were examined for microorganisms on the fish surface and viscera. Water, sediment, and fish samples from the ponds were tested for fecal streptococci, fecal coliforms, Aeromonas hydrophila , and Pseudomonas aeruginosa , while fish samples were also analyzed for presumptive Listeria spp. (count on m-VJ agar) and psychrotrophic bacteria. There were no significant differences (P<0.05) in the fecal streptococci and fecal coliform counts for the water, sediment, and fish visceral samples. However, the aeromonad count for the visceral samples (4.20 log CFU/g wet weight) was significantly higher (P<0.05) than that of the water and sediments (2.40 log CFU/ml and 3.78 log CFU/g wet weight, respectively). Similarly, the P. aeruginosa count for the fish visceral samples was significantly higher (P<0.05) than that of the water and sediments. The mean presumptive Listeria spp. count for the fish visceral samples was 1.99 log CFU/g wet weight. Because of the higher bacterial concentrations in the fish viscera, it was concluded that cross-contamination of fish samples could occur during evisceration. Finally, the feed protein level and feeding method used in the ponds influenced the bacterial concentrations in selected sample types (i.e., water, sediment, fish surface rinse, or fish viscera).

Fate of Listeria monocytogenes and Aeromonas hydrophila on Packaged Channel Catfish Fillets Stored at 4°C.

The fate of Listeria monocytogenes and Aeromonas hydrophila on packaged channel catfish fillets stored at 4°C was determined. Fish fillets were inoculated with L. monocytogenes or A. hydrophila or both and packaged by film overwrapping or vacuum skin packaging. They were stored at 4°C for 16 d. The L. monocytogenes population did not increase on the samples while the A. hydrophila population increased from 3 log CFU/cm2 on day 1 to 6 log CFU/cm2 on day 16 with a faster growth rate on overwrapped packaged fillets. Inoculation of both bacteria on the product markedly suppressed each other's growth.

Fate of Listeria monocytogenes on Packaged, Refrigerated, and Frozen Seafood.

The fate of Listeria monocytogenes on packaged, refrigerated, and frozen seafood was determined. Fish and shrimp were inoculated with L. monocytogenes , packaged by one of three treatments (film overwrap or 2 vacuum packaging treatments), and stored either on ice for 21 d or at -20°C for 3 months. No increase in the L. monocytogenes population on iced products was noted regardless of the type of packaging. Populations on frozen products decreased by less than 1 log after 3 months. Vacuum packaging of chilled fish and seafood in the material evaluated may be an alternative packaging treatment.

Thermal Death Times for Listeria monocytogenes (Scott A) in Crabmeat.

Thermal death times for Listeria monocytogenes (Scott A) in blue crabmeat were determined. Blue crabmeat was inoculated with 107 cells of L. monocytogenes strain Scott A/g prior to distributing 7.5 g into sausage casings (1.6 cm × 4 cm). Ten to 12 sausages, one with a thermocouple connected to a recorder, were placed into a preheated, recirculating water bath at either 50, 55, or 60°C. At designated times, the L. monocytogenes populations was determined by plating serial dilutions onto both trypticase soy agar (TSA) and modified Vogel-Johnson agar. After incubation, presumptive L. monocytogenes colonies were counted. D-values based on enumeration of colonies from TSA were 40.43, 12.00, and 2.61 min at 50, 55, and 60°C, respectively. Heat resistance as demonstrated by using modified Vogel-Johnson agar as the plating medium was less with D-values of 34.48, 9.18, and 1.31 min at each of the same heating temperatures. Z-values of 8.40 and 6.99°C were derived from the TSA and modified Vogel-Johnson agar data, respectively. Based on these findings, the current parameters used to commercially pasteurize crabmeat are adequate to inactivate L. monocytogenes strain Scott A.