Anastasios Zotos

Dispersive liquid–liquid microextraction for the determination of organochlorine pesticides residues in honey by gas chromatography-electron capture and ion trap mass spectrometric detection

A simple dispersive liquid-liquid microextraction (DLLME) protocol for the determination of 15 organochlorine pesticides residues in honey is proposed. The selected pesticides were separated using gas chromatography and detected by electron capture (ECD) or ion trap mass spectrometry (GC-IT/MS). Several parameters affecting the extraction efficiency namely type and volume of organic extraction solvent, type and volume of disperser solvent, sample pH, ionic strength, extraction time and centrifugation speed were systematically investigated. The final DLLME protocol involved the addition of 750 μL acetonitrile (disperser) and 50 μL chloroform (extraction solvent) into a 5 mL aqueous honey solution followed by centrifugation. The sedimented organic phase (chloroform) were analysed directly by GC-IT/MS or evaporated and reconstituted in acetonitrile prior to the GC-ECD analysis. The analytical performance of the GC-ECD and GC-IT/MS methods was compared and discussed. Under the selected experimental conditions, the enrichment factors varied between of 36 and 114. The limits of detection (LOD) were in the range of 0.02-0.15 μg L(-1) (0.4-3 ng g(-1)) for GC-ECD and 0.01-0.2 μg L(-1) (0.2-4 ng g(-1)) for GC-IT/MS which is adequate to verify compliance of products to legal tolerances. The proposed method was applied to the analysis of the selected organochlorine pesticides residues in various honey samples obtained from Greek region. Mean recoveries were ranged from 75% to 119% while the precision was better than 20% in both methodologies.

Seasonal changes in composition, fatty acid, cholesterol and mineral content of six highly commercial fish species of Greece:

Changes in lipid, protein, ash and moisture, quantitative distribution of fatty acids, cholesterol and mineral content (Cd, Co, Cr, Cu, Fe, Mg, Mn, Ni and Zn) of six common fish species from Greece were studied. The proximate composition of all samples was significantly influenced by the fishing period, even the cultured ones. The sum of C20:5ω–3 and C22:6ω–3 ranged from 1.18 to 2.76 for sardine, from 0.37 to 1.99 for bogue, from 1.1 to 1.52 for mackerel, from 1.23 to 1.46 for sea bass, from 1.00 to 1.24 for trout and from 0.26 to 0.45 g/100 g edible portion for hake samples. Besides the ratio of ω–3/ω–6 polyunsaturated fatty acids ranged from 6.80 to 19.00 for the wild fish samples and from 1.01 to 3.67 for the cultured ones. Cholesterol of sardine, bogue, mackerel, trout and sea bass ranged from 37 up to 76 mg/100 g edible flesh, while it was varied from 72 to 124 mg/100 g in the flesh of hake. The concentration of minerals was detected at acceptable levels. All fish samples seemed to be quite good sources of Mg, Zn and Fe. Co was not detected in the samples studied.

Effect of baking of sardine (Sardina pilchardus) and frying of anchovy (Engraulis encrasicholus) in olive and sunflower oil on their quality

Quality changes due to oven-baking of sardine for 20, 40, 50 and 60 min and due to deep frying of anchovy for 2, 3, 4 and 5 min in olive and sunflower oil were studied. Linear increase in total losses with the time of processing was observed. A linear inverse relationship was observed between moisture/lipid and moisture/protein due to time of baking of sardines and time of frying of anchovies (wet matter). However, no changes were detected in sardine samples due to time of baking (dry matter), while a reduction in proteins and ash followed by an increase in lipids was detected in fried anchovies due to time of frying (dry matter). The fatty acid profiles indicated that a rich in EPA + DHA (33.16%) and in ω-3/ω-6 ratio (9.40) baked sardines can be produced in 20 min at 200 °C. The fatty acid profiles of fried anchovies tremendously changed, indicating entirely different products. Olive oil is probably a better medium to fry fish products, since either the two beneficial fatty acids (EPA and DHA) detected at higher concentrations in anchovies fried in olive oil or the ω-3/ω-6 ratio remained at higher values (0.71–2.56). An increase of cholesterol and squalene content with increasing the time of baking was detected in sardine samples, probably due to decline of moisture content. On the contrary, cholesterol significantly reduced due to frying of anchovy in olive oil. Simultaneously squalene concentration significantly and linearly increased, from 3.87 mg/100 g in the unprocessed anchovies to 73.25 mg/100 g in the samples fried for 5 min, indicating its existence at beneficial levels, besides low cholesterol concentration detected in fried olive oil and squalene concentration gradually and linearly decreased, confirming the absorption from the anchovy samples. Similar was the changes of cholesterol and squalene in anchovies samples fried in sunflower oil.

The Effect of Washing, Microbial Transglutaminase, Salts and Starch Addition on the Functional Properties of Sardine (Sardina Pilchardus) Kamaboko Gels

The functional properties of kamaboko gels, produced from sardine surimi (SS), were evaluated based on three factors: (a) washing conditions, (b) absence or presence of microbial transglutaminase (MTGase), and (c) addition of CaCl2, MgCl 2, and NH4Cl. Fish gels prepared from sardine mince washed at pH 5.5 showed the highest L* values and whiteness index (WI). A similar trend was also observed for firmness. Addition of MTGase had a beneficial effect on the L* values and WI as well as on firmness and cohesiveness of kamaboko gels (p<0.05). Fish gels containing CaCl2 or MgCl 2 were lighter and firmer compared to those containing NH4Cl (p<0.05). Three transitions, using a differential scanning calorimeter (DSC), were observed during heating of surimi containing 50g/kg of unmodified wheat starch (UWS), occurring at 45.2 °C, 63.3 °C, and 71.4 °C and were substantially the same when UWS was added at higher concentrations (p≥0.05). The thermal gelation of SS, studied with Dynamic Mechanical Analysis (DMA), occurred at three or four distinct stages named: gel softening, low temperature gelation, gel resolution, and high temperature gelation. The cooling process was found to be crucial in surimi gelation since it strongly contributed in the development of gel rigidity.

Improvement in the production of smoked trout fillets ("Salmo gairdnerii") steamed with liquid smoke

A fish smoking process was applied using a combination of liquid smoke and steaming at pressures up to 1 bar above atmospheric pressure. Drying and brining prior to smoking have not shown any significant effect on the quality of the trout fillets. The nondried and dried for 4 h trout fillets prior to processing were assessed as slightly more acceptable products, processing yield, sensory analysis, instrumental color, pH, available lysine, polycyclic aromatic hydrocarbons (PAHs) and preservation time were estimated. The losses due to processing of the nondried samples were 23.7%, while 28.8% for those dried for 4h. The fillets processed at 1.5 and 2 bar steam pressure were assessed as highly acceptable. Lightness (L*) showed a decreasing trend due to pressure while redness (a*), and yellowness (b*) an increasing one. The destruction of available lysine was low (13.20% ± 1.01) and it was slightly dependent upon the process. No PAHs were detected. The preservation time, studied via total viable count, lactic acid bacteria, existence of Listeria monocytogenes, and sensory analysis, extended to more than 98 days at 4 ± 1 °C. The drying prior to processing seems to affect the preservation time. This processing technique is much simpler, faster, environmental effective, and leads to high quality smoked products.

Studies on the roles and identification of the small molecule factors on phenolase activation in Norway lobster (Nephrops norvegiens)

The direct effect of small molecule factor(s) on the activation of phenolase in the suggested multiple component process of blackspot development in Norway lobster (Nephrops norvegicus) was studied. When acetone-precipitated phenolase was treated with the small molecule filtrate, the phenolase activation profile was lower compared to the control, particularly at pH 6.7. However, when purified form I phenolase was treated with small molecule filtrate no activation was observed, the assayed phenolase activity being constant but lower than the control. A similar effect was observed when form I phenolase was treated with 1.5 mM dihydroxyphenylalanine (DOPA) (approximately the concentration found in the small molecule filtrate), indicating that this lower pattern resulted from the competition of the phenolase substrates (catechol, DOPA, and possibly tyrosine) in the small molecule filtrate. When the activity of the three proteases in the acetone-precipitated phenolase was studied, it was found that there was a high recovery of proteases I and II in the precipitate but little protease III. This also confirms that the effect of the small molecule filtrate on phenolase activity in acetone-precipitated phenolase was due to its indirect effect on the proteases present. The modification of the compounds that absorb at 270 nm was studied by a high-performance liquid chromatographic (HPLC) separation method and the quantitative analysis of tyrosine and DOPA in the small molecule filtrate was achieved. Some of the small molecules were identified, by an HPLC method, as tyrosine, DOPA, oxidised DOPA, vitamin C in oxidised form or breakdown product of vitamin C. Oxidised DOPA, DOPA, tyrosine and N-acetyldopamine inhibit the proteases to varying degrees. Vitamin C also inhibits all three proteases at pH 6.4; however, at pH 8.2 it markedly activates protease III and slightly activates the two thiol proteases (I and II).

Studies on the influence of small molecule factor(s) on protease activities in Norway lobster (Nephrops norvegicus)

The interactions of small molecule(s) and proteases in the suggested multiple component process of phenolase activation in Norway lobster (Nephrops norvegicus) were studied. It was observed that protease I is mainly a visceral protease, whilst proteases II and III exist in approximately equal amounts in viscera and flesh, respectively. It was found that the small molecule factor(s) (molecular weight less than 10 000) inhibited proteases I, II and III at pH 6.7 and 8.2, except when it was freshly prepared and then it activated only protease III and only at pH 8.2. This indicates that the small molecule(s) may be modified on storage. It was found that the small molecule factor(s) is primarily water-soluble and heat-stable, with heating resulting in a similar change to storage in its effect on proteases. Treatment of the small molecule filtrate with a cation-exchanger also increased the inhibition effect on proteases. However, whilst treatment with an anion-exchanger at pH 8.2 reduced inhibition, treatment at pH 4.0 restored activation of protease III. These results also infer that the small molecule factor(s) is modified on storage or exists in two forms.

Optimization of a steaming with liquid smoke smoking process of Mediterranean mussel (Mytilus galloprovincialis)

Mussel samples were brined in 0%, 10% and 20% sodium chloride solutions and steamed with 2%, 5% and 8% liquid smoke at 1 (atmospheric pressure), 1.5 and 2 bar pressures. Sodium chloride and moisture content and instrumental color were analyzed. They were also objectively and hedonically assessed by 35 and 70 panellists, respectively, for their smoked flavor, saltiness, juiciness and color. The sodium chloride content was influenced by the brine concentration and brightness by the smoke and brine concentrations. The smoked flavor significantly and linearly (p = 0.018) influenced by the liquid smoke and brine concentration. A linear effect (p < 0.001) of brine concentration (p < 0.001) on sensory saltiness was expectedly observed. Brine concentration affected not only the saltiness but also the juiciness of the mussel smoked products. All factors of the study produced a significant linear effect (p = 0.008) on the intensity of color. Brine concentration was the only process variable affecting all the hedonic sensory variables in a curvilinear mode. The optimal hedonic conditions of the mussel products were achieved at smoke concentrations 3.8–8%, brine level from 8.5% to 13.5% at pressure 1 bar and from 11.5% to 16.5% at pressure 1.5 bar. All hedonic variables reached their maximum likeliness between 9 and 13. The optimal objective saltiness and juiciness varied between moderate and adequate salty and juicy product. The physicochemical variables employed in the study were adequately perceived by the panellists’ sensory objective assessment as redundancy analysis revealed. Sodium chloride and moisture were the most important chemical variables (R2 = 42% and 13%, respectively).

Physicochemical, textural and sensory properties of white soft cheese made from buffalo and cow milk mixtures

The physicochemical, textural and sensory properties of white soft cheeses made from three different buffalo and cow milk mixtures (100:0, 70:30 and 30:70) during 3-month storage were studied. The increase in buffalo milk concentration resulted in increasing total cheese yield, dry matter (DM) and fat retention and fat in DM content. However, it caused reductions in moisture content, salt intake, hardness, chewiness, elasticity, sensory hardness and sensory cohesiveness of the samples. The percentage of water-soluble nitrogen to total nitrogen increased during storage resulting in decreased fracturability, hardness (textural and sensory), cohesiveness (textural and sensory), springiness, chewiness and elasticity. The panellists evaluated the white soft cheese made with buffalo milk as the most acceptable.