Impact of new ingredients obtained from brewer's spent yeast on bread characteristics.

The impact of bread fortification with ß-glucans and with proteins/proteolytic enzymes from brewers' spent yeast on physical characteristics was evaluated. ß-Glucans extraction from spent yeast cell wall was optimized and the extract was incorporated on bread to obtain 2.02 g ß-glucans/100 g flour, in order to comply with the European Food Safety Authority guidelines. Protein/proteolytic enzymes extract from spent yeast was added to bread at 60 U proteolytic activity/100 g flour. Both ß-glucans rich and proteins/proteolytic enzymes extracts favoured browning of bread crust. However, breads with proteins/proteolytic enzymes addition presented lower specific volume, whereas the incorporation of ß-glucans in bread lead to uniform pores that was also noticeble in terms of higher specific volume. Overall, the improvement of nutritional/health promoting properties is highlighted with ß-glucan rich extract, not only due to bread ß-glucan content but also for total dietary fibre content (39% increase). The improvement was less noticeable for proteins/proteolytic enzymes extract. Only a 6% increase in bread protein content was noted with the addition of this extract and higher protein content would most likely accentuate the negative impact on bread specific volume that in turn could impair consumer acceptance. Therefore, only ß-glucan rich extract is a promising bread ingredient.

Fortification of Wheat Bread with Agroindustry By-Products: Statistical Methods for Sensory Preference Evaluation and Correlation with Color and Crumb Structure.

The use of agroindustry by-products (BP) for fortification of wheat bread can be an alternative to waste disposal because BP are appealing sources of dietary fiber. Moreover, it may also contribute to indirect income generation. In this study, sensory, color, and crumb structure properties of breads fortified with fiber rich fraction recovered from four types of agroindustry BP were tested, namely orange (OE), pomegranate (PE), elderberry (EE), and spent yeast (YE). Statistical models for sensory preference evaluation and correlation with color and crumb structure were developed. External preference mapping indicated consumer preferences and enabled selection of the concentrations of BP fibre-rich fraction with best acceptance, namely 7.0% EE, 2.5% OE, 5.0% PE, and 2.5% YE. Data collected from image analysis complemented sensory profile information, whereas multivariate PLS regression provided information on the relationship between "crust color" and "crumb color" and instrumental data. Regression models developed for both sensory attributes presented good fitting (R2 Y > 0.700) and predictive ability (Q2 > 0.500), with low RMSE. Crust and crumb a* parameters had a positive influence on "crust color" and "crumb color" models, while crust L* and b* had a negative influence.

In vitro bioacessibility and transport across Caco-2 monolayers of haloacetic acids in drinking water.

Water disinfection plays a crucial role in water safety but it is also a matter of concern as the use of disinfectants promotes the formation of disinfection by-products (DBPs). Haloacetic acids (HAAs) are one of the major classes of DBPs since they are frequently found in treated water, are ubiquitous, pervasive and have high water solubility, so a great concern emerged about their formation, occurrence and toxicity. Exposure to HAAs is influenced by consumption patterns and diet of individuals thus their bioavailability is an important parameter to the overall toxicity. In the current study the bioacessibility of the most representative HAAs (chloroacetic acid - MCAA, bromoacetic acid - MBAA, dichloroacetic acid - DCAA, dibromoacetic acid - DBAA, and trichloroacetic acid - TCAA) after simulated in vitro digestion (SIVD) in tap water and transport across Caco-2 monolayers was evaluated. Compounds were monitored in 8 points throughout the digestion phases by an optimized LC-MS/MS methodology. MCAA and MBAA were not bioaccessible after SIVD whereas DCAA, DBAA and TCAA are highly bioaccessible (85 ± 4%, 97 ± 4% and 106 ± 7% respectively). Concerning transport assays, DCAA and DBAA were highly permeable throughout the Caco-2 monolayer (apparent permeability and calculated fraction absorbed of 13.62 × 10(-6) cm/s and 90% for DCAA; and 8.82 × 10(-6) cm/s and 84% for DBAA), whereas TCAA showed no relevant permeability. The present results may contribute to efficient risk analysis studies concerning HAAs oral exposure from tap water taking into account the different biological behaviour of these chemically similar substances.

Impact of cooking and handling conditions on furanic compounds in breaded fish products.

This study evaluates the influence of cooking and handling conditions on the quantity of furanic compounds (furan, 2-furfural, furfuryl alcohol, 2-pentylfuran, 5-hydroxymethylfurfural) in breaded fish products. Oven-baking and reheating in the microwave lead to low furanic compounds formation in comparison with deep-frying. The use of olive oil for deep-frying promoted higher levels of furanic compounds than sunflower oil. The amounts of these compounds diminished as the temperature and time of deep-frying decreased as well as after a delay after deep-frying. Thus, the generation of furanic compounds can be minimized by adjusting the cooking method and conditions, such as using an electric oven, deep-frying in sunflower oil at 160°C during 4min, or waiting 10min after cooking. However, these conditions that reduce furanic compounds levels also reduce the content of volatile compounds related to the aroma and flavour of fried foods. In this sense, new efforts should be done to reduce the formation of furanic compounds without being detrimental to the volatile profile.

Quantification of furanic compounds in coated deep-fried products simulating normal preparation and consumption: optimisation of HS-SPME analytical conditions by response surface methodology.

The validation of a method for the simultaneous quantification of furanic compounds in coated deep-fried samples processed and handled as usually consumed is presented. The deep-fried food was grinded using a device that simulates the mastication, and immediately analysed by headspace solid phase microextraction coupled to gas chromatography-mass spectrometry. Parameters affecting the efficiency of HS-SPME procedure were selected by response surface methodology, using a 2(3) full-factorial central composite design. Optimal conditions were achieved using 2g of sample, 3g of NaCl and 40min of absorption time at 37°C. Consistency between predicted and experimented values was observed and quality parameters of the method were established. As a result, furan, 2-furfural, furfuryl alcohol and 2-pentylfuran were, for the first time, simultaneously detected and quantified (5.59, 0.27, 10.48 and 1.77µgg(-1) sample, respectively) in coated deep-fried fish, contributing to a better understanding of the amounts of these compounds in food.

Effect of solvent to sample ratio on total lipid extracted and fatty acid composition in meat products within different fat content.

The effect of solvent to sample ratio on total extracted lipids and fatty acid (FA) composition in meat products with different fat contents was evaluated. Total lipids were extracted according to the Folch et al. (1957) method, using a 20:1 ratio of chloroform:methanol (2:1, v/v) to sample (A), and also testing the solvent:sample ratio of 10:1 (B). Higher amounts of total lipids and total FA from neutral lipids were obtained using the A ratio, which could be due to an insufficient chloroform:dry-weight sample proportion which could be insufficient for solubilizing the total amount of lipids. In the polar lipid fraction, the total amount of FA was higher using the B rather than the A ratio, which may be caused by the higher volume of added water when using A than B. When studying the FA composition of different lipid fractions, the volume of both the solvent and the water for total lipid extraction should be considered.

Effect of charcoal types and grilling conditions on formation of heterocyclic aromatic amines (HAs) and polycyclic aromatic hydrocarbons (PAHs) in grilled muscle foods.

Grilling muscle foods involves high temperatures that lead to production of cooking toxicants, such as heterocyclic aromatic amines (HAs) and polycyclic aromatic hydrocarbons (PAHs). To obtain realistic exposure levels of these two groups of mutagens analyses of the same samples using similar separation/detection techniques were performed. HAs and PAHs were quantified in well-done meat and fish samples grilled with wood and coconut shell charcoal at 200°C. Quantitative HAs and PAHs profiles were different for beef and salmon using the same type of charcoal. Higher levels of HAs and PAHs were found in salmon samples. No significant differences were observed for HAs and PAHs in beef samples grilled with both charcoal types, whereas salmon grilled with coconut shell charcoal presented significantly lower amounts of HAs and PAHs than salmon grilled with usual wood charcoal. Continuous barbecuing with the same charcoal shown that combustion of fat that dropped along the grilling period contributed to higher formation of HAs and PAHs. Special attention must be given to the intake of barbecued foods since high amounts of HAs and PAHs can be taken in a single meal.

A comparison of the extraction procedures and quantification methods for the chromatographic determination of polycyclic aromatic hydrocarbons in charcoal grilled meat and fish.

A method for analysis of 15 PAHs in charcoal-grilled meat/fish was established by high performance liquid chromatography and fluorescence detection. Gradient elution was performed with methanol/water/ethyl acetate. Maxima excitation and emission wavelengths were selected for each PAH. Retention times were very stable with coefficients of variation below 0.24% within analytical day and below 0.60% across analytical days. Two different methods of cleanup and pre-concentration steps were compared. Solvent extraction assisted by sonication carried out with n-hexane on 2g of lyophilized meat or 1g of lyophilized fish allowed to obtain high sensitivity, reproducibility and better extraction efficiency. Limits of quantification (LOQs, s/n=10) were lower than 0.01ng/g of meat wet weight and lower than 0.02ng/g of fish wet weight for all PAHs (except for Na, Fl and IP that were lower than 0.1ng/g). Two different quantification methods were compared. Standard addition method compensated PAHs losses due to incomplete extraction and it is recommended for analyses of grilled meat and fish samples that usually contain very low amounts of the eight high molecular weight PAHs (BaA, Ch, BbF, BkF, BaP, IP, BgP, DhA).

Groundwater from infiltration galleries used for small public water supply systems: contamination with pesticides and endocrine disruptors.

Infiltration galleries are among the oldest known means used for small public water fountains. Owing to its ancestral origin they are usually associated with high quality water. Thirty-one compounds, including pesticides and estrogens from different chemical families, were analysed in waters from infiltration galleries collected in Alto Douro Demarcated Wine region (North of Portugal). A total of twelve compounds were detected in the water samples. Nine of these compounds are described as presenting evidence or potential evidence of interfering with the hormone system of humans and wildlife. Although concentrations of the target analytes were relatively low, many of them below their limit of quantification, four compounds were above quantification limit and two of them even above the legal limit of 0.1 µg/L: dimethoate (30.38 ng/L), folpet (64.35 ng/L), terbuthylazine-desethyl (22.28 to 292.36 ng/L) and terbuthylazine (22.49 to 369.33 ng/L).

Quantification of endocrine disruptors and pesticides in water by gas chromatography-tandem mass spectrometry. Method validation using weighted linear regression schemes.

A multi-residue methodology based on a solid phase extraction followed by gas chromatography-tandem mass spectrometry was developed for trace analysis of 32 compounds in water matrices, including estrogens and several pesticides from different chemical families, some of them with endocrine disrupting properties. Matrix standard calibration solutions were prepared by adding known amounts of the analytes to a residue-free sample to compensate matrix-induced chromatographic response enhancement observed for certain pesticides. Validation was done mainly according to the International Conference on Harmonisation recommendations, as well as some European and American validation guidelines with specifications for pesticides analysis and/or GC-MS methodology. As the assumption of homoscedasticity was not met for analytical data, weighted least squares linear regression procedure was applied as a simple and effective way to counteract the greater influence of the greater concentrations on the fitted regression line, improving accuracy at the lower end of the calibration curve. The method was considered validated for 31 compounds after consistent evaluation of the key analytical parameters: specificity, linearity, limit of detection and quantification, range, precision, accuracy, extraction efficiency, stability and robustness.

Short communication: effect of kefir grains on proteolysis of major milk proteins.

The effect of kefir grains on the proteolysis of major milk proteins in milk kefir and in a culture of kefir grains in pasteurized cheese whey was followed by reverse phase-HPLC analysis. The reduction of kappa-, alpha-, and beta-caseins (CN), alpha-lactalbumin (alpha-LA), and beta-lactoglobulin (beta-LG) contents during 48 and 90 h of incubation of pasteurized milk (100mL) and respective cheese whey with kefir grains (6 and 12 g) at 20 degrees C was monitored. Significant proteolysis of alpha-LA and kappa-, alpha-, and beta-caseins was observed. The effect of kefir amount (6 and 12 g/100mL) was significant for alpha-LA and alpha- and beta-CN. alpha-Lactalbumin and beta-CN were more easily hydrolyzed than alpha-CN. No significant reduction was observed with respect to beta-LG concentration for 6 and 12 g of kefir in 100mL of milk over 48 h, indicating that no significant proteolysis was carried out. Similar results were observed when the experiment was conducted over 90 h. Regarding the cheese whey kefir samples, similar behavior was observed for the proteolysis of alpha-LA and beta-LG: alpha-LA was hydrolyzed between 60 and 90% after 12h (for 6 and 12 g of kefir) and no significant beta-LG proteolysis occurred. The proteolytic activity of lactic acid bacteria and yeasts in kefir community was evaluated. Kefir milk prepared under normal conditions contained peptides from proteolysis of alpha-LA and kappa-, alpha-, and beta-caseins. Hydrolysis is dependent on the kefir:milk ratio and incubation time. beta-Lactoglobulin is not hydrolyzed even when higher hydrolysis time is used. Kefir grains are not appropriate as adjunct cultures to increase beta-LG digestibility in whey-based or whey-containing foods.

Quantification of residual nitrite and nitrate in ham by reverse-phase high performance liquid chromatography/diode array detector.

Nitrite and nitrate are used as additives in ham industry to provide colour, taste and protect against clostridia. The classical colorimetric methods widely used to determine nitrite and nitrate are laborious, suffer from matrix interferences and involve the use of toxic cadmium. The use of chromatography is potentially attractive since it is more rapid, sensitive, selective and provides reliable and accurate results. A rapid and cost-effective RP-HPLC method with diode array detector was optimized and validated for quantification of nitrites and nitrates in ham. The chromatographic separation was achieved using a HyPurity C18, 5 microm chromatographic column and gradient elution with 0.01 M n-octylamine and 5mM tetrabutylammonium hydrogenosulphate to pH 6.5. The determinations were performed in the linear range of 0.0125-10.0mg/L for nitrite and 0.0300-12.5 g/L for nitrate. The detection limits were 0.019 and 0.050 mg/kg, respectively. The reliability of the method in terms of precision and accuracy was evaluated. Coefficients of variation lower than 2.89% and 5.47% were obtained for nitrite and nitrate, respectively (n=6). Recoveries of residual nitrite/nitrate ranged between 93.6% and 104.3%. Analysis of cooked and dried ham samples was performed, and the results obtained were in agreement with reference procedures.

Casein breakdown in terrincho ovine cheese: comparison with bovine cheese and with bovine/ovine cheeses.

Terrincho cheese is an uncooked, pressed cheese made from raw whole ovine milk from the "Churra da Terra Quente" breed. It requires a minimum ripening time of 30 d. A detailed evaluation of the effect of ripening time on the breakdown of the casein fractions, along with the formation of major breakdown products of casein hydrolysis, was monitored by HPLC to contribute to a more complete characterization of this product. In 30-d-old cheeses, only 20% of alpha(S1)-casein remained intact; the beta-casein fraction was more resistant to hydrolysis. The ripening time of Terrincho cheese can be predicted using 2 variables of normalized peak areas of alpha(S1)-casein and alpha(S1)-I peptide, and a constant; the estimation error is 2.5 d. The pH 4.3-insoluble fraction of Terrincho and cheeses manufactured with bovine milk and with ovine milk combined with 2 levels of bovine milk (10 and 20%) revealed different chromatographic and electrophoretic profiles, especially the alpha(S1)-casein fraction. Similar proteolysis progress was observed, particularly in the percentage of casein fraction degradation. However, using both analytical methods, the detection of 10% bovine milk at 30 d of ripening was no longer possible as result of alpha(S1)-casein hydrolysis. The discriminate analysis applied to HPLC data indicated that at 30 d of ripening, differences between the casein fractions of Terrincho cheese and mixture cheeses were mainly from beta1-casein content. The function thus obtained was able to correctly classify all the samples according to cheese type. Using the descriptive sensory profile, Terrincho cheese at 30 d of ripening could be distinguished from bovine and mixture cheeses owing to its higher fracturability and adhesiveness and lower elasticity and hardness, which correlated with its lower total casein content.

Chemical, physical, and sensorial characteristics of "Terrincho" ewe cheese: changes during ripening and intravarietal comparison.

The objectives of this study were to monitor the changes in chemical [moisture, acidity, pH, and water activity (a(w))] and physical (color and texture) parameters of "Terrincho" ewe cheese during 60 d of ripening, and to determine the correlations between the changes in instrumental texture and color parameters and the ripening time of the product. Intravarietal comparison of Terrincho ewe cheese from 5 different dairy plants was performed by evaluation of mechanical parameters from texture profile analysis (TPA) and color parameters in terms of CIELAB color space (L*, a*, and b*). In addition to mechanical and color tests, composition analyses and sensory tests were performed. The results were evaluated with statistical methods (single valued and multivariate analysis). During the first 20 d of ripening, an increase in hardness, fracturability, gumminess, chewiness, and yellowness occurred. Simultaneously, adhesiveness, resilience, L* (inside cheese, "i" and external "e"), and cohesiveness decreased. After 20 d of ripening hardness, fracturability, gumminess, and chewiness decreased and cohesiveness increased. The ripening time of Terrincho cheeses can be estimated with 6 variables: L* (external, e), L* (i), b* (inside cheese, i), hardness, a* (i), chewiness, and a constant. The estimation error was 4.2 d. Evaluation of composition, pH, texture profile analyses, color, and related sensory characteristics of Terrincho cheeses from 5 different dairy plants (with 30 d of ripening) revealed correlations between these parameters.

Quantification of short-chain free fatty acids in "Terrincho" ewe cheese: intravarietal comparison.

Headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry was employed for the quantification of volatile free fatty acids (FFA) in "Terrincho" ewe cheese. Solid-phase microextraction quantitative analysis was feasible under equilibrium situations as long as the conditions of agitation and the adsorption time were held constant. An excellent linear relationship between the amount of the adsorbed analyte and its initial concentration in the sample matrix was obtained when an adequate amount of sample was chosen. Thus, quantification was possible if biases due to competition or linear range excesses were controlled. Solid-phase microextraction sampling was carried out at 65 degrees C, and a fiber coated with an 85-micro/m polyacrylate film was chosen. After equilibration at 65 degrees C for 40 min, the fiber was exposed to the headspace above the sample for 20 min and then inserted into the gas chromatograph. The evolution of the volatile FFA during Terrincho ewe cheese ripening was analyzed for a 60-d period. An overall increase in FFA contents was verified up to 30 d of ripening. Between 30 and 45 d most FFA did not suffer significant changes. All FFA increased significantly by the 60-d ripening period. The excessive lipolysis observed at 60 d of ripening may result in the presence of off-flavors. Principal component analysis performed for intravarietal comparison of volatile FFA composition of 19 Terrincho cheeses, analyzed at 30 ripening days, enabled discrimination between cheeses produced at five different dairy plants.

Solid-phase microextraction of volatile compounds in "Terrincho" ewe cheese. Comparison of different fibers.

Solid-phase microextraction coupled to gas chromatography-mass spectrometry (GC-MS) was applied to study the volatile compounds in "Terrincho" ewe cheese. Six types of fibers were tested and the main extraction parameters were studied. Carboxen-polydimethylsiloxane fiber 75 microm (CAR-PDMS) achieved the most complete profile of ewe cheese volatile compounds. The optimised conditions used for characterization of "Terrincho" ewe cheese were: sample vial equilibration at 20 degrees C for 20 min, followed by CAR-PDMS fiber exposure to the headspace above the sample for 30 min and finally thermal desorption of the adsorbed substances, in the injector port for GC-MS analysis. This technique was a useful tool for the differentiation of 11 "Terrincho" ewe cheeses, all taken from the same cheesemaking season with 30 days of ripening but from three different farmhouses, according to their volatile fraction. Results obtained were statistically treated by categorical principal component analysis. Subsequently, 49.15% of the variation in data was due to the first dimension (k = 12.7) and the second dimension (k = 8.88) accounted for 34.2% of the total information. Volatile profiles among samples indicated cheese group separation according to farmhouse of production.

Quantification of non-protein nitrogen components of infant formulae and follow-up milks: comparison with cows' and human milk.

The composition of fourteen infant formulae and six follow-up milks with regard to their free amino acids (including taurine), free nucleotides, orotic acid, and free and total l-carnitine content was studied. The levels found were compared with the limits established in European legislation and with the composition of human and cows' milk samples. HPLC methodologies, optimized and validated for the matrices under study, were used, except for free and total l-carnitine contents that were quantified using a flow-injection manifold, also optimized and validated for the matrices under study. Global statistical treatment of the results by cluster analysis indicated similarities between the contents of the N compounds under study of infant formulae, follow-up milks and cows' milk and differences with regard to human milk composition. The principal component analysis showed that 60.2 % of the variation in data was due to the first principal component, and the second component represented 23.8 % of the total information. Nucleotide profiles, orotic acid, and free and total l-carnitine contents explain the main differences observed between human milk and the other milks studied (cows' milk, infant formulae and follow-up milks). Cows' milk is distinguished from infant formulae and follow-up milks mainly owing to the different uric acid contents and free amino acids profiles.

Solid-phase microextraction in combination with GC/MS for quantification of the major volatile free fatty acids in ewe cheese.

This work describes a method for quantification of the major free fatty acids of ewe cheese that contribute to its distinct and strongly marked flavor. A headspace SPME method in combination with GC/MS was used for the extraction, identification, and quantification of butanoic, hexanoic, octanoic and decanoic acids in ewe cheeses. The method used for sample preparation was simple. A fiber coated with 85-microm polyacrylate film was chosen to extract the free fatty acids. To perform a reliable quantification, several factors were taken into consideration for reliable quantification, namely, (i) the influence of addition of water, of an electrolyte or of a hygroscopic salt, on the release of free fatty acids from the matrix; (ii) the linear relationship between the amount of analyte adsorbed by the SPME polymer film and the initial concentration of the analyte in the cheese sample; and (iii) the competition for adsorption by fiber. Water removal with sodium sulfate promoted a more efficient extraction of volatile free fatty acids; biases due to competition or linear range excesses were controlled by choosing the appropriate amount of sample for each ewe cheese. The method of standard additions was used with success for the quantification of free fatty acids. Calibration curves that were constructed for the major short-chain free fatty acids (butanoic, hexanoic, octanoic, and decanoic acids) spiked into cheese followed linear relationships with highly significant (p < 0.001) correlation coefficients (r > 0.999). Coefficients of variation of <7.9% indicated that the technique was reproducible. A marked increase in concentration of short-chain free fatty acids was observed during cheese ripening, ranging from 0.35 to 9.33 mg/100 g for butanoic acid, 0.363 to 4.34 mg/100 g for hexanoic acid, 0.343 to 2.0 mg/100 g for octanoic acid, and 1.291 to 3.85 mg/100 g for decanoic acid. The limits of quantification were registered at levels of parts per million. The absolute quantification of butanoic acid was also carried out by using isotope dilution assays (IDA). The levels of acid obtained with this method were similar to those obtained by the standard additions method.