Effect of Chitosan and Alginate-Based Edible Membranes with Oregano Essential Oil and Olive Oil in the Microbiological, Physicochemical and Organoleptic Characteristics of Mutton.

Edible chitosan or alginate coatings and their combinations with oregano essential oil or olive oil, have been examined for their effect on the microbiological, physicochemical and organoleptic characteristics of mutton. The results indicated that these edible coatings can contribute to maintaining good quality characteristics and extending mutton shelf-life. The total mesophilic counts in mutton ranged from 3.48 to 8.00 log10 CFU/g, the total psychrophilic counts from 4.00 to 9.50 log10 CFU/g, the B. thermosphacta counts from 2.30 to 7.77 log10 CFU/g and the lactic acid bacteria counts from 2.00 to 5.85 log10 CFU/g. Chitosan coatings significantly (p < 0.05) reduced the total mesophilic, the total psychrophilic (1-2 log10 cfu/g), the B. thermosphacta and the lactic acid bacteria counts in mutton. Alginate exhibited a lower L* value and a higher a* value and chroma compared with the control and chitosan lots. No significant differences were observed in the chemical composition of meat pieces among the experimental groups. Oregano oil positively affected the sensory attributes of meat. The most favourable combination, based on the microbiological counts, the organoleptic characteristics and the shelf-life extension of mutton, was that of chitosan with oregano essential oil.

A Comparative Study on Microbiological and Chemical Characteristics of Small Ruminant Carcasses from Abattoirs in Greece.

Meat quality dictates consumer preferences with hygiene forming a key component, especially in meat types with declining popularity, such as sheep and goat meat. Aiming to increase the marketability of sheep and goat meat, we examined 370 sheep and goat carcasses from two abattoirs in Greece. Tests included enumeration of the total mesophilic viable count, total psychrophilic viable count and coliform count, and detection of Salmonella spp., Listeria monocytogenes and presumptive ESBL Escherichia coli. Moreover, designated samples of meat were used to measure pH, moisture, total fat and protein content. Goat carcasses had significantly higher microbial counts compared to sheep carcasses. Lamb and kid carcasses had larger TMVC, TPVC and coliform counts compared to carcasses from adult animals. One strain of L. monocytogenes (0.8%), typed as serovar 1/2a (3a), was isolated from one adult sheep carcass. Twelve strains of ESBL Escherichia coli (25%) were isolated; there were not any strains of Salmonella spp. The average values of pH, moisture, total fat and total protein were 5.83%, 67.76%, 7.21% and 21.31%, respectively, for sheep carcasses and 5.70%, 68.2%, 5.69% and 24.10%, respectively, for goat carcasses. The results showed a small deviation in assessed parameters, implying the uniformity of the conditions concerning rearing and slaughtering.

Dry Fermented Sausages with Total Replacement of Fat by Extra Virgin Olive Oil Emulsion and Indigenous Lactic Acid Bacteria.

RESEARCH BACKGROUND: Formulations based on vegetable or fish oil and modifications in the production technology of dry fermented sausages have emerged in recent years aiming to achieve the desirable target of reducing the fat content of these meat products. However, previous efforts have confronted many difficulties, such as high mass loss and unacceptable appearance due to intensely wrinkled surfaces and case hardening. The objective of this study is to produce and evaluate dry fermented sausages by utilising a meat protein-olive oil emulsion as fat substitute and indigenous lactic acid bacteria (LAB) with probiotic properties isolated from traditional Greek meat products. EXPERIMENTAL APPROACH: A novel formulation with extra virgin olive oil and turkey protein was developed to totally replace the conventionally added pork fat. Probiotic and safety characteristics of autochthonous LAB isolates from spontaneously fermented sausages were evaluated and three LAB isolates were finally selected as starter cultures. Physicochemical, microbiological and sensory analyses were carried out in all treatments (control, Lactobacillus acidophilus, L. casei, L. sakei and Pediococcus pentosaceus) during fermentation. RESULTS AND CONCLUSIONS: Ready-to-eat sausages were found to be microbiologically stable. The olive oil-based formulation produced in this study generated a mosaic pattern visible in the sliced product simulating the fat in conventional fermented sausages and was regarded as an ideal fat substitute for the production of fermented sausages. An autochthonous isolate of Lactobacillus casei adapted the best to the final products as it was molecularly identified to be present in the highest counts among the LAB isolates used as starter cultures. NOVELTY AND SCIENTIFIC CONTRIBUTION: Α novel and high-quality dry fermented meat product was produced by replacing the added pork fat with a fat substitute based on a meat protein-olive oil emulsion. Autochthonous LAB with in vitro probiotic properties could have a potential use in large-scale novel dry fermented sausage production. Such isolates could be used as starters in an effort to standardise the production process and retain the typical organoleptic and sensory characteristics. Moreover, isolates like L. casei 62 that survived in high counts in the final products can increase the safety of fermented sausages by competing not only with pathogens but also with the indigenous microbiota and could have a potential functional value for the consumer.

Effect of olive leaf (Olea europea L.) extracts on protein and lipid oxidation of long-term frozen n-3 fatty acids-enriched pork patties.

Our previous study has demonstrated the protective effects of olive leaf extracts on the oxidation of pork patties from n-3 fatty acid-enriched meat during refrigerated storage. The target of the present study was to examine these effects during frozen storage. Results showed that frozen storage accelerated (P=0.05) both lipid and protein oxidation in pork patties, but an addition of olive leaf extract at 200mg gallic acid equivalent/kg improved sensory attributes by delaying oxidation of lipids (reduction (P=0.05) of conjugated dienes, hydroperoxides and malondialdehyde), and of proteins (reduction (P=0.05) of protein carbonyls and inhibition (P=0.05) of the decrease of protein sulfhydryls).

Effect of olive leaf (Olea europea L.) extracts on protein and lipid oxidation in cooked pork meat patties enriched with n-3 fatty acids.

BACKGROUND: The effect of olive leaf extracts on lipid and protein oxidation of cooked pork patties refrigerated stored for 9 days was evaluated. Patties were prepared from longissimus dorsi muscle of pigs, and dietary supplemented with linseed oil. RESULTS: Results showed that dietary linseed oil modified the fatty acid composition of pork patties by increasing (P ≤ 0.05) n-3 (α-linolenic acid) and decreasing (P ≤ 0.05) n-6 (linoleic acid) fatty acids. Olive leaf extracts at supplementation levels of 200 and, especially, of 300 mg gallic acid equivalents kg⁻¹ meat, delayed lipid oxidation by reducing (P ≤ 0.05) both primary (conjugated dienes and hydroperoxides) and secondary (malondialdehyde) oxidation products. They also inhibited protein oxidation in a concentration-dependent manner by reducing (P ≤ 0.05) protein carbonyls and increasing (P ≤ 0.05) protein sulfhydryls. In addition, they improved sensory attributes of the n-3 enriched patties. CONCLUSION: Results suggested that olive leaf extracts might be useful to the meat industry as an efficient alternative to synthetic antioxidants.

Olive leaves (Olea europaea L.) versus α-tocopheryl acetate as dietary supplements for enhancing the oxidative stability of eggs enriched with very-long-chain n-3 fatty acids.

BACKGROUND: Ninety-six brown Lohmann laying hens were equally assigned into four groups with six replicates. Hens within the control group were given a corn/soybean-based diet supplemented with 30 g kg(-1) fish oil. Two other groups were given the same diet further supplemented with olive leaves at 5 (OL5) and 10 (OL10) g kg(-1) respectively, while the diet of the fourth group was supplemented with α-tocopheryl acetate (TOC) at 200 mg kg(-1). Eggs were analysed for lipid hydroperoxide and malondialdehyde (MDA) contents, fatty acid profile, α-tocopherol content and susceptibility to iron-induced lipid oxidation. RESULTS: Neither OL nor TOC supplementation affected (P>0.05) the fatty acid composition. Dietary supplementation with OL10 or TOC reduced (P≤0.05) the lipid hydroperoxide content but exerted no (P>0.05) effect on the MDA content of fresh eggs compared with controls. Eggs submitted to iron-induced lipid oxidation from the OL5 group presented higher (P≤0.05) MDA levels than the control but lower (P≤0.05) than the OL10 group. Eggs from the TOC group presented lower (P≤0.05) MDA levels compared with all groups at all incubation time points. CONCLUSION: The results of this study suggested that dietary supplementation with both OL10 and TOC could protect n-3 fatty acids in eggs from deterioration.

Levels of biogenic amines in retail market fermented meat products.

The qualitative and quantitative profile of biogenic amines (BA) in 50 samples of dry fermented sausages sold in Greek retail markets were determined by HPLC. Putrescine, cadaverine, tryptamine, ß-phenylethylamine spermidine, spermine were analysed by UV detection after pre-column derivatization with benzoyl chloride, whereas tyramine and histamine were analysed by fluorescence detection after post-column derivatization with o-phthaldialdehyde (OPA). With the exception of spermidine and spermine a wide variation of BA levels was observed among the samples. Of the BA examined, tyramine, putrescine, histamine and cadaverine showed high concentrations ranging from: 0 to 510 mg/kg (median: 197.7 mg/kg), 0 to 505 mg/kg (median: 96.5mg/kg), 0 to 515 mg/kg (median: 7.0mg/kg) and 0 to 690 mg/kg (median: 3.6 mg/kg), respectively. The histamine content of 28% of the samples exceeded the toxicity limit of 100mg/kg set for histamine in some fish species. Levels of tryptamine and ß-phenyl-ethylamine never exceeded 50 and 29 mg/kg, respectively. Results of the present study suggest that the amounts of BA in dry fermented sausages, sold in Greek retail markets, may pose a potential health risk for sensitive individuals or for those undergoing classical monoamine oxidase inhibiting (MAOI) drug therapy.

Lipid and protein oxidation of α-linolenic acid-enriched pork during refrigerated storage as influenced by diet supplementation with olive leaves (Olea europea L.) or α-tocopheryl acetate.

The objective of this study was to evaluate the effect of diet supplementation with olive leaves or α-tocopheryl acetate on lipid and protein oxidation of raw and cooked n-3 enriched-pork during refrigerated storage. Enrichment of pork with α-linolenic acid through diet supplementation with linseed oil enhanced (p≤0.05) lipid oxidation in both raw and cooked chops but had no effect (p>0.05) on protein oxidation during refrigerated storage while decreasing (p≤0.05) the sensory attributes of cooked pork. Diet supplementation with olive leaves or α-tocopheryl acetate had no effect (p>0.05) on the fatty acid composition of pork but decreased (p≤0.05) lipid oxidation while exerting no effect (p>0.05) on protein oxidation in both raw and cooked α-linolenic acid-enriched chops stored and chilled for 9 days. Moreover, olive leaves and α-tocopheryl acetate supplemented at 10 g/kg and 200mg/kg diet, respectively, exerted (p≤0.05) a beneficial effect on the sensory attributes of cooked α-linolenic acid-enriched pork chops.

Rapid quantifiable assessment of nutritional parameters influencing pediocin production by Pediococcus acidilactici NRRL B5627.

A direct plate bioassay procedure was applied for rapid and quantifiable assessment of the influence of various nutritional parameters on pediocin production by Pediococcus acidilactici NRRL B5627. Solid-state cultivation of the microorganism was done on MRS-based media over 3-and 6-hours incubation periods. Nutritional parameters assessed included the carbon source (glucose, sucrose, fructose, galactose, glycerol), and various salts (NH(4)PO(4), CaCl(2), KH(2)PO(4), MnSO(4).H(2)O). Glucose was found to be the optimal carbon source while glycerol exhibited the most suppressive effect. Using glucose as the carbon source, addition of various salts, in amounts used in liquid media commonly applied in the cultivation of the pediococci, was assessed with respect to bacteriocin production on a per cell basis. Experimental data obtained showed that several nutritional parameters repress pediocin production by P. acidilactici, while the direct plate assay proved to be a good pilot assay prior to conducting more intensive kinetic analysis in liquid cultivation.

Pediocin SA-1, an antimicrobial peptide from Pediococcus acidilactici NRRL B5627: production conditions, purification and characterization.

Fermentation broths of Pediococcus acidilactici NRRL B5627 exhibited a certain antimicrobial activity due to a bacteriocin produced during early growth and until the stationary phase of growth was reached (at optimum of 60% dissolved oxygen saturation). Its size was determined by electrospray ionization mass spectrometric analysis as 3.660 kDa. N-terminal sequencing showed that the bacteriocin had 19 amino acid residues in the order KYYGXNGVXTXGKHSXVDX. The purified bacteriocin is similar to pediocins isolated by various Pediococci and therefore, it belongs to the class IIa of bacteriocins and is thus designated pediocin SA-1. Sensitivity of the purified pediocin to various storage temperatures and enzyme treatments was examined. Purified pediocin SA-1 is heat stable for up to 60 min at 121 degrees C. Pediocin SA-1 is inhibitory to several food-borne pathogens and food spoilage bacteria. It appears to be significantly more effective against Listeria spp. compared to pediocin PD-1 produced by P. damnosus. The mode of action of the purified bacteriocin appears to be bactericidal.

Mould growth on traditional greek sausages and penicillin production by Penicillium isolates.

Visible moulds were isolated and identified from traditional Greek sausages from Northern Greece. Penicillium species were isolated from 90.8% of visibly mouldy sausages. Penicillium solitum, P. nalgiovense and P. commune species made up 60.6% of the total number of isolates. The most frequently occurring species was P. solitum (26.1%). P. nalgiovense and P. olsonii were found to be positive to penicillin production in an agar assay and further examination for antibiotic production in liquid culture with complex media designed for penicillin production, confirmed their ability for penicillin biosynthesis. Penicillin production by P. olsonii is reported for the first time in this study.

Effect of rosemary extract, chitosan and α-tocopherol on lipid oxidation and colour stability during frozen storage of beef burgers.

The effect of rosemary extract, chitosan and α-tocopherol, added individually or in combination, on lipid oxidation and colour stability of frozen (-18°C) beef burgers stored for 180 days was investigated. The burgers' lipid oxidation and appearance were evaluated through measurement of primary (conjugated dienes and peroxide value) and secondary (malondialdehyde) oxidation products, together with visual assessment and instrumental measurement of colour. Chitosan alone and in combination with either rosemary or α-tocopherol had the best antioxidative effect (P⩽0.05) compared to individual use of rosemary or α-tocopherol, while the best results were obtained with the combination of chitosan and rosemary. The differences of antioxidative effects, however, between individual use of rosemary or α-tocopherol as compared to the controls were also significant (P⩽0.05). Chitosan added individually or in combination with either rosemary or α-tocopherol had also a noteworthy effect on the burgers' appearance as it contributed to red colour retention for a much longer period (P⩽0.05) compared all other treatments and the controls. In conclusion, the best antioxidative effects were obtained with the combination of chitosan and rosemary extract.

Effect of rosemary extract, chitosan and α-tocopherol on microbiological parameters and lipid oxidation of fresh pork sausages stored at 4°C.

The effect of rosemary extract, chitosan and α-tocopherol, added individually or in combination, on microbiological parameters and lipid oxidation of fresh pork sausages stored for 20days at 4°C was investigated. Microbiological determinations included the enumeration of Enterobacteriaceae, Pseudomonas spp., total viable count, yeasts and moulds and lactic acid bacteria, while pH values were also determined. Lipid oxidation was evaluated through measurement of primary (peroxide value) and secondary (malondialdehyde concentration) oxidation products. Chitosan addition resulted in significant (P⩽0.05) inhibition of microbial growth, while the lowest microbial counts were obtained in the samples containing both chitosan and rosemary, indicating a possible synergistic effect. Chitosan and its combinations with either α-tocopherol or rosemary, and especially the latter combination, showed also the most intense antioxidative effect (P⩽0.05), at least when compared to α-tocopherol alone or the controls, while use of rosemary extract alone had a comparable (P>0.05) antioxidative effect with chitosan and its combinations. Shelf life of samples containing chitosan was almost doubled compared to the remaining samples. In conclusion, the best antimicrobial and antioxidative effects were obtained from the combination of chitosan with the rosemary extract.

Determination of bacteriocin activity with bioassays carried out on solid and liquid substrates: assessing the factor "indicator microorganism".

BACKGROUND: Successful application of growth inhibition techniques for quantitative determination of bacteriocins relies on the sensitivity of the applied indicator microorganism to the bacteriocin to which is exposed. However, information on indicator microorganisms' performance and comparisons in bacteriocin determination with bioassays is almost non-existing in the literature. The aim of the present work was to evaluate the parameter "indicator microorganism" in bioassays carried out on solid -agar diffusion assay- and liquid -turbidometric assay- substrates, applied in the quantification of the most studied bacteriocin nisin. RESULTS: The performance of characterized microorganisms of known sources, belonging to the genera of Lactobacillus, Pediococcus, Micrococcus and Leuconostoc, has been assessed in this work in the assays of plate agar diffusion and turbidometry. Dose responses and sensitivities were examined and compared over a range of assay variables in standard bacteriocin solutions, fermentation broth filtrates and processed food samples. Measurements on inhibition zones produced on agar plates were made by means of digital image analysis. The data produced were analyzed statistically using the ANOVA technique and pairwise comparisons tests. Sensitivity limits and linearity of responses to bacteriocin varied significantly among different test-microorganisms in both applied methods, the lower sensitivity limits depending on both the test-microorganism and the applied method. In both methods, however, only two of the nine tested microorganisms (Lactobacillus curvatus ATCC 51436 and Pediococcus acidilactici ATCC 25740) were sensitive to very low concentrations of the bacteriocin and produced a linear-type of response in all kinds of samples used in this work. In all cases, very low bacteriocin concentrations, e.g. 1 IU/ml nisin, were more accurately determined in the turbidometric assay. CONCLUSION: The present work shows that in growth inhibition techniques used in bacteriocin quantification, the choice of the indicator microorganism is critical. Evaluation of sensitivity levels and type of produced responses showed that they can vary widely among different test-microorganisms and different applied methods, indicating that not all microorganisms can be used successfully as indicators and that measurements of growth inhibition in liquid media produce more reliable results.

Relation of biogenic amines' formation with microbiological and sensory attributes in Lactobacillus-inoculated vacuum-packed rainbow trout (Oncorhynchus mykiss) fillets.

The biogenic amine (BA) content of vacuum-packed filleted rainbow trout (Oncorhynchus mykiss) inoculated or not with two different Lactobacillus strains, individually or in combination, was monitored during refrigerated storage for 20 days and related to respective bacteriological and sensory changes occurring during the same period. Eight amines, namely putrescine, cadaverine, tyramine, tryptamine, beta-phenylethylamine, histamine, spermine, and spermidine, were determined, whereas agmatine was not detected in any of the samples. In all cases, BA concentration was higher (P < or = 0.05) in the controls compared to all inoculated treatments, whereas the trend with regard to the bacterial populations (Enterobacteriaceae, pseudomonads, and H2S-producing bacteria) and the off-odor scores was similar. Inoculation with Lactobacillus sakei CECT 4808 showed the best preservative effect among inoculated treatments. Concentrations of putrescine and cadaverine, the main BAs formed, correlated well with both spoilage bacterial counts and off-odor scores and can be useful indicators of shelf life. Spermine and spermidine contents decreased during storage, while levels of the other determined BAs remained below 10 mg/kg even after sensory rejection.