Combining different biomarkers to distinguish Chemlali virgin olive oils from different geographical areas of Tunisia.

BACKGROUND: Despite their high potential, Tunisian virgin olive oils (VOOs) are mainly exported in bulk or blended with VOOs of other origin, hindering their international market placement. To face this situation, their valorization is needed by highlighting their unique features and by developing tools to guarantee their geographical authenticity. Compositional features of Chemlali VOOs produced in three Tunisian regions were assessed to identify suitable authenticity markers. RESULTS: Quality indices ensured the quality of the VOOs studied. Volatile compounds, total phenols, fatty acid (FA) and chlorophylls are significantly influenced by the region of origin, which was justified by the differences found in soil and climatic conditions of the three geographical regions. To explore the capabilities of these markers for the geographical authentication of Tunisian Chemlali VOOs, classification models based on partial least squares-discriminant analysis (PLS-DA) were developed by grouping the minimum number of variables allowing the highest discrimination power, minimizing in this way the analytical procedure. The PLS-DA authentication model based on combining volatile compounds with FA or with total phenols achieved a correct classification of 95.7% of the VOOs according to their origin, as assessed by 10%-out cross-validation. Sidi Bouzid Chemlali VOOs achieved 100% of correct classification, while the misclassification between Sfax and Enfidha ones did not exceed 10%. CONCLUSIONS: These results allowed to establish the most promising and affordable combination of markers for the geographical authentication of Tunisian Chemlali VOOs from distinct production regions and provide the basis to further develop authentication models based on wider datasets. © 2023 Society of Chemical Industry.

Effect of Feeding Acid Oils on European Seabass Fillet Lipid Composition, Oxidative Stability, Color, and Sensory Acceptance.

Acid oils (AO) are fat by-products of edible oil refining with a high energetic value, being an interesting option for a more sustainable aquaculture nutrition. This study was conducted to evaluate the effects of the partial replacement of fish oil (FO) in diets by two AO instead of crude vegetable oils on the lipid composition, lipid oxidation and quality of fresh European seabass fillets, and after their commercial refrigerated storage for 6 days. Fish were fed with five different diets, the added fat being FO (100%) or a blend of FO (25%) and another fat (75%): crude soybean oil (SO), soybean-sunflower acid oil (SAO), crude olive pomace oil (OPO), or olive pomace acid oil (OPAO). Fresh and refrigerated fillets were assessed for fatty acid profile, tocopherol (T) and tocotrienol (T3) composition, lipid oxidative stability, 2-thiobarbituric acid (TBA) value, volatile compound content, color, and sensory acceptance. Refrigerated storage did not affect T + T3 total content but increased secondary oxidation products (TBA values and volatile compound contents) in fillets from all diets. The FO substitution decreased EPA and DHA and increased T and T3 in fish fillets, but the recommended human daily intake of EPA plus DHA could still be covered with 100 g of fish fillets. Both a higher oxidative stability and a lower TBA value were found in SO, SAO, OPO, or OPAO fillets, obtaining the greatest oxidative stability in OPO and OPAO fillets. Sensory acceptance was not affected by the diet or the refrigerated storage, while the differences found in color parameters would not be perceived by the human eye. According to the oxidative stability and acceptability of flesh, SAO and OPAO are adequate replacements of FO as energy source in European seabass diets, which implies that these by-products can be upcycled, improving the environmental and economical sustainability of aquaculture production.

Determination and Comparison of the Lipid Profile and Sodium Content of Gluten-Free and Gluten-Containing Breads from the Spanish Market.

The objective is to verify if gluten-free (GF) and gluten-containing (G) breads differ in their sodium content and lipid profile. Samples of GF (n = 20) and G (n = 14) sliced white sandwich bread of commercial brands most frequently consumed in Spain were collected. The fatty acid (FA) composition and the contents of sodium, fat, cholesterol and phytosterols were determined. Sodium, fat and cholesterol contents were significantly higher in GF bread. The FA composition also differed, while G breads declared in most instances the use of sunflower oil as fat ingredient and presented a higher polyunsaturated FA percentage; GF breads declared a wide variety of fats and oils as ingredients (coconut, palm, olive, sunflower, etc.) which was reflected in their FA profile. Cholesterol content was higher in GF bread because five samples declared the use of whole egg, while G samples did not include any egg product in their formulas. Phytosterol content was higher in G bread but its variability was greater in GF bread. In conclusion, nutritional quality of GF bread varied depending on the ingredients used and might be lower than that of G bread. However, these differences in composition could be reduced or eliminated through changes in the formulation of GF bread. Moreover, the comparison of the results obtained in our laboratory for fat and salt content with the declared contents on the labels showed a much higher deviation for GF samples and it can be concluded that the quality of the nutritional information declared was lower in GF samples.

Influence of cell-cell contact between L. thermotolerans and S. cerevisiae on yeast interactions and the exo-metabolome.

Sequential fermentation of grape must inoculated with L. thermotolerans and then S. cerevisiae 24 h later (typical wine-making practice) was conducted with or without cell-cell contact between the two yeast species. We monitored cell viability of the two species throughout fermentation by flow cytometry. The cell viability of S. cerevisiae decreased under both conditions, but the decrease was greater if there was cell-cell contact. An investigation of the nature of the interactions showed competition between the two species for nitrogen compounds, oxygen, and must sterols. Volatile-compound analysis showed differences between sequential and pure fermentation and that cell-cell contact modifies yeast metabolism, as the volatile-compound profile was significantly different from that of sequential fermentation without cell-cell contact. We further confirmed that cell-cell contact modifies yeast metabolism by analyzing the exo-metabolome of all fermentations by FT-ICR-MS analysis. These analyses show specific metabolite production and quantitative metabolite changes associated with each fermentation condition. This study shows that cell-cell contact not only affects cell viability, as already reported, but markedly affects yeast metabolism.

Toward a Harmonized and Standardized Protocol for the Determination of Total Hydroxytyrosol and Tyrosol Content in Virgin Olive Oil (VOO). The Pros of a Fit for the Purpose Ultra High Performance Liquid Chromatography (UHPLC) Procedure.

Τoward a harmonized and standardized procedure for the determination of total hydroxytyrosol and tyrosol content in virgin olive oil (VOO), the pros of a recently published in house validated ultra high performance liquid chromatography (UHPLC) protocol are discussed comparatively with those of other procedures that determine directly or indirectly the compounds hosted under the health claim on "olive oil polyphenols" (EC regulation 432/2012). Authentic VOOs were analyzed with five different liquid chromatographic separation protocols and 1H-NMR one in five different laboratories with expertise in VOO phenol analysis within three months. Data comparison indicated differences in absolute values. Method comparison using appropriate tools (Passing-Bablok regression and Bland Altman analyses) for all protocols vs. the UHPLC one indicated slight or statistically significant differences. The results were also discussed in terms of cost effectiveness, detection means, standard requirements and ways to calculate the total hydroxytyrosol and tyrosol content. Findings point out that the in-house validated fit for the purpose UHPLC protocol presents certain pros that should be exploited by the interested parties. These are the simplicity of sample preparation, fast elution time that increase the number of samples analyzed per day and integration of well-resolved peaks with the aid of only two commercially available external standards. Importance of correction factors in the calculations is stressed.

Effect of the Storage Conditions and Freezing Speed on the Color and Chlorophyll Profile of Premium Extra Virgin Olive Oils.

Premium extra virgin olive oils (PEVOO) are oils of exceptional quality and retail at high prices. The green color of recently extracted olive oils is lost during storage at room temperature, mainly because of the pheophytinization of chlorophylls. Since a green color is perceived as a mark of high-quality oils by consumers, it is especially important for PEVOO to maintain their initial green color. This study assessed the effect of applying low temperatures (refrigeration and freezing) and modified atmospheres on the color of four PEVOO for 24 months. Also, the effect of two freezing methods (slow freezing by placing the oil at -20 °C and fast freezing by immersing the oil in a bath of liquid nitrogen) was studied. Results showed that the green color was better preserved in oils frozen and stored at -20 °C whereas in oils frozen with liquid nitrogen the green color was lost much faster during frozen storage. An in-depth study of this unexpected phenomenon showed that this loss of green color was mainly due to a pheophytinization of chlorophylls. This phenomenon did not happen at the moment of freezing with liquid nitrogen, but over the first 100 days of storage at -20 °C. In addition, correlations between single chlorophyll and pheophytin contents and chromatic coordinates were established.

Revealing adulterated olive oils by triacylglycerol screening methods: Beyond the official method.

Official control methods to detect olive oil (OO) adulteration fail to provide satisfactory consumer protection. Thus, faster and more sensitive screening tools are needed to increase their effectiveness. Here, the official method for adulterant detection in OO was compared with three untargeted screening methods based on triacylglycerol analysis using high-throughput (FIA-HESI-HRMS; HT-GC-MS; HPLC-RID) and pattern recognition techniques (PLS-DA). They were assayed on a set of genuine and adulterated samples with a high natural variability (n = 143). The sensitivity of the official method was 1 for high linoleic (HL) blends at ≥2 % but only 0.39 for high oleic (HO) blends at ≥5 %, while specificity was 0.96. The sensitivity of the screening methods in external validation was 0.90-0.99 for the detection of HL and 0.82-0.88 for HO blends. Among them, HT-GC-MS offered the highest sensitivity (0.94) and specificity (0.76), proving to be the most suitable screening tool for OO authentication.

Subzero Temperature Storage to Preserve the Quality Attributes of Veiled Virgin Olive Oil.

Unfiltered olive oils (UO) have gained popularity in the global market, but they lose their quality characteristics faster than filtered oils (FO). In this work, refrigeration and freezing temperatures were explored to maintain UO quality features during storage. A full factorial design was applied to an UO and to the same oil after filtration to evaluate the effect of storage temperature (room temperature, 4 °C and -20 °C) and freezing speed (slow-freezing, in the freezer at -20 °C and fast-freezing, in a bath of liquid nitrogen). Official quality parameters, polar and nonpolar phenolic compounds, oxidative stability index, volatile compounds and descriptive sensory profile were measured periodically over 24 months of storage in the dark. Storage temperature influenced the quality of both UO and FO, but in different ways. At non-freezing temperature, UO experienced a severe decrease in its sensory quality compared to FO, mainly due to the hydrolysis of secoiridoids and degradation of the volatile fraction, but storage at -20 °C allowed to effectively preserve UO quality traits, thus resulting as a suitable strategy to increase the shelf-life of UO to satisfy the demand of consumers for this particular product. The results showed that slow-freezing was the most appropriate method for freezing.

Thiamine and Biotin: Relevance in the Production of Volatile and Non-Volatile Compounds during Saccharomyces cerevisiae Alcoholic Fermentation in Synthetic Grape Must.

Vitamins are major cofactors to numerous key metabolic pathways in enological yeasts, and both thiamine and biotin, notably, are believed to be essential to yeast fermentation and growth, respectively. In order to further assess and clarify their role in winemaking, and in the resulting wine, alcoholic fermentations of a commercial Saccharomyces cerevisiae active dried yeast were conducted in synthetic media containing various concentrations of both vitamins. Growth and fermentation kinetics were monitored and proved the essential character of biotin in yeast growth, and of thiamine in fermentation. The synthetic wine volatile compounds were quantified, and notable influences of both vitamins appeared, through a striking positive effect of thiamine on the production of higher alcohols, and of biotin on fatty acids. Beyond the evidence of this influence on fermentations and on the production of volatiles, this work proves, for the first time, the impact held by vitamins on wine yeasts' exometabolome, investigated through an untargeted metabolomic analysis. This highlighted chemical differences in the composition of synthetic wines through a notably marked influence of thiamine on 46 named S. cerevisiae metabolic pathways, and especially in amino acid-associated metabolic pathways. This provides, overall, the first evidence of the impact held by both vitamins on the wine.

Effect of Feeding Olive Pomace Acid Oil on Dark Chicken Meat Lipid Composition, Oxidative Stability, Color, and Sensory Acceptance.

This study evaluated the effect of using olive pomace acid oil (OPAO) instead of crude palm oil (PO) or refined olive pomace oil (ROPO) on lipid composition, lipid oxidation, and quality of chicken meat. Broiler chickens were fed diets with 6% of PO, ROPO, or OPAO, and deboned legs with skin were sampled. Fresh and refrigerated (commercial conditions; 7 days) chicken meat samples were assessed for fatty acid (FA) composition, tocopherol (T) and tocotrienol (T3) content, lipid oxidative stability, 2-thiobarbituric acid (TBA) values, volatile compounds, color, and sensory acceptance. Using ROPO and OPAO led to meat richer in monounsaturated FAs and OPAO to lower α-T levels compared to PO. Oxidative stability, TBA values, volatile compounds, and overall acceptance of meat were not affected by diet. Refrigeration increased TBA values and some volatile compounds' concentrations, but it did not decrease redness or consumers' overall acceptance. Therefore, the OPAO used was an adequate fat source for chicken diets at 6%, as it produced dark meat lower in saturated FAs than PO without affecting lipid oxidation or overall acceptance. According to this, upcycling OPAO as an energy source in chicken diets would be possible, which can contribute to the sustainability of the food chain.

Method for the analysis of volatile compounds in virgin olive oil by SPME-GC-MS or SPME-GC-FID.

During the course of the EU H2020 OLEUM project, a harmonized method was developed to quantify volatile markers of the aroma of virgin olive oil with the aim to support the work of sensory panel test to assess the quality grade. A peer validation of this method has been carried out, with good results in terms of analytical quality parameters. The method allows the quantification of volatile compounds by SPME-GC with two possible detectors, flame ionization detector and mass spectrometry, depending on the technical facilities of the labs applying this method. The method was optimized for the quantification of 18 volatile compounds that were selected as being markers responsible for positive attributes (e.g. fruity) and sensory defects (e.g. rancid and winey-vinegary). The quantification is carried out with calibration curves corrected by the internal standards. Additionally, a protocol is provided to prepare the calibration samples. This procedure enhances reproducibility between labs since one of the main sources of errors is the application of different procedures in calibration.

Yeast-yeast interactions revealed by aromatic profile analysis of Sauvignon Blanc wine fermented by single or co-culture of non-Saccharomyces and Saccharomyces yeasts.

There has been increasing interest in the use of selected non-Saccharomyces yeasts in co-culture with Saccharomyces cerevisiae. The main reason is that the multistarter fermentation process is thought to simulate indigenous fermentation, thus increasing wine aroma complexity while avoiding the risks linked to natural fermentation. However, multistarter fermentation is characterised by complex and largely unknown interactions between yeasts. Consequently the resulting wine quality is rather unpredictable. In order to better understand the interactions that take place between non-Saccharomyces and Saccharomyces yeasts during alcoholic fermentation, we analysed the volatile profiles of several mono-culture and co-cultures. Candida zemplinina, Torulaspora delbrueckii and Metschnikowia pulcherrima were used to conduct fermentations either in mono-culture or in co-culture with S. cerevisiae. Up to 48 volatile compounds belonging to different chemical families were quantified. For the first time, we show that C. zemplinina is a strong producer of terpenes and lactones. We demonstrate by means of multivariate analysis that different interactions exist between the co-cultures studied. We observed a synergistic effect on aromatic compound production when M. pulcherrima was in co-culture with S. cerevisiae. However a negative interaction was observed between C. zemplinina and S. cerevisiae, which resulted in a decrease in terpene and lactone content. These interactions are independent of biomass production. The aromatic profiles of T. delbrueckii and S. cerevisiae in mono-culture and in co-culture are very close, and are biomass-dependent, reflecting a neutral interaction. This study reveals that a whole family of compounds could be altered by such interactions. These results suggest that the entire metabolic pathway is affected by these interactions.

Using fluorescence excitation-emission matrices to predict bitterness and pungency of virgin olive oil: A feasibility study.

Unlike other food products, virgin olive oil must undergo an organoleptic assessment that is currently based on a trained human panel, which presents drawbacks that might affect the efficiency and robustness. Therefore, disposing of instrumental methods that could serve as screening tools to support sensory panels is of paramount importance. The present work aimed to explore excitation-emission fluorescence spectroscopy (EEFS) to predict bitterness and pungency, since both attributes are related with fluorophore compounds, such as polar phenols. Bitterness and pungency intensities of 250 samples were provided by an official sensory panel and used to build and compare partial least squares regressions (PLSR) with the excitation-emission matrix. Both PARAFAC scores and two-way unfolded data led to successful PLSR. The most relevant PARAFAC scores agreed with virgin olive oil phenolic spectra, evidencing that EEFS would be the fit-for-purpose screening tool to support the sensory panel.

Use of a Minimal Microbial Consortium to Determine the Origin of Kombucha Flavor.

Microbiological, chemical, and sensory analyses were coupled to understand the origins of kombucha organoleptic compounds and their implication in the flavor of the kombucha beverage. By isolating microorganisms from an original kombucha and comparing it to monocultures and cocultures of two yeasts (Brettanomyces bruxellensis and Hanseniaspora valbyensis) and an acetic acid bacterium (Acetobacter indonesiensis), interaction effects were investigated during the two phases of production. 32 volatile compounds identified and quantified by Headspace-Solid Phase-MicroExtraction-Gas Chromatography/Mass Spectrometry (HS-SPME-GC/MS) were classified according to their origin from tea or microorganisms. Many esters were associated to H. valbyensis, while alcohols were associated to both yeasts, acetic acid to A. indonesiensis, and saturated fatty acids to all microorganisms. Concentration of metabolites were dependent on microbial activity, yeast composition, and phase of production. Sensory analysis showed that tea type influenced the olfactive perception, although microbial composition remained the strongest factor. Association of B. bruxellensis and A. indonesiensis induced characteristic apple juice aroma.

Geographical authentication of virgin olive oil by GC-MS sesquiterpene hydrocarbon fingerprint: Verifying EU and single country label-declaration.

According to the last report from the European Union (EU) Food Fraud Network, olive oil tops the list of the most notified products. Current EU regulation states geographical origin as mandatory for virgin olive oils, even though an official analytical method is still lacking. Verifying the compliance of label-declared EU oils should be addressed with the highest priority level. Hence, the present work tackles this issue by developing a classification model (PLS-DA) based on the sesquiterpene hydrocarbon fingerprint of 400 samples obtained by HS-SPME-GC-MS to discriminate between EU and non-EU olive oils, obtaining an 89.6% of correct classification for the external validation (three iterations), with a sensitivity of 0.81 and a specificity of 0.95. Subsequently, multi-class discrimination models for EU and non-EU countries were developed and externally validated (with three different validation sets) with successful results (average of 92.2% of correct classification for EU and 96.0% for non-EU countries).

Stepwise strategy based on 1H-NMR fingerprinting in combination with chemometrics to determine the content of vegetable oils in olive oil mixtures.

1H NMR fingerprinting of edible oils and a set of multivariate classification and regression models organised in a decision tree is proposed as a stepwise strategy to assure the authenticity and traceability of olive oils and their declared blends with other vegetable oils (VOs). Oils of the 'virgin olive oil' and 'olive oil' categories and their mixtures with the most common VOs, i.e. sunflower, high oleic sunflower, hazelnut, avocado, soybean, corn, refined palm olein and desterolized high oleic sunflower oils, were studied. Partial least squares (PLS) discriminant analysis provided stable and robust binary classification models to identify the olive oil type and the VO in the blend. PLS regression afforded models with excellent precisions and acceptable accuracies to determine the percentage of VO in the mixture. The satisfactory performance of this approach, tested with blind samples, confirm its potential to support regulations and control bodies.

Oxidative Quality of Acid Oils and Fatty Acid Distillates Used in Animal Feeding.

Acid oils (AO) and fatty acid distillates (FAD) are byproducts from chemical and physical refining of edible oils and fats, respectively. Their high energy value makes their upcycling interesting as alternatives to conventional fats in animal feeding. The objective of this study is to characterize their oxidative quality and to provide recommendations about their evaluation for animal feeding purposes. The oxidation status (peroxide value (PV), p-Anisidine value (p-AnV), % polymeric compounds (POL)), the oxidative stability (induction time by the Rancimat at 120 °C (IT)), the fatty acid composition (FA), and tocopherol and tocotrienol content of 92 AO and FAD samples from the Spanish market were analyzed. Both AO and FAD showed low PV (0.8 and 1 meq O2/kg); however, p-AnV was higher in FAD (36.4 vs. 16.4 in AO) and POL was higher in AO (2.5% vs. not detected in FAD) as a consequence of the type of refining process. The botanical origin of AO and FAD influenced FA and tocol composition, and they influenced IT. A high variability was observed for most analyzed parameters, reinforcing the need for standardizing AO and FAD to obtain reliable feed ingredients and to include primary and secondary oxidative parameters within their quality control.

Methods to determine the quality of acid oils and fatty acid distillates used in animal feeding.

Acid oils and fatty acid distillates are by-products from the refining of edible oils and fats. They are used as feed ingredients, but their highly variable composition sometimes affects the productive parameters of the animals. Thus, their quality control and standardization are necessary. The official methods recommended for crude and refined fats and oils must be modified to give reliable results when applied to acid oils and fatty acid distillates. This article summarizes the drawbacks that were encountered during the setup of the analytical methods and how were they overcome by adapting the methods to these type of fat samples. Some methods such as the determinations of fatty acid composition, tocopherol and tocotrienol content, unsaponifiable matter, acidity and peroxide value had to be minimally adapted. However, others such as the determinations of moisture and volatile matter, insoluble impurities, lipid classes and p-anisidine value showed important drawbacks that required a more significant adaptation.•All the analytical methods have been successfully applied to acid oils and fatty acid distillates.•A detailed description of the sample preparation for analysis and applied analytical methods is provided as a compendium of methods in the supplementary material.•These methods will be extremely useful to improve the quality control of these heterogeneous feed ingredients.

Composition and Nutritional Value of Acid Oils and Fatty Acid Distillates Used in Animal Feeding.

Acid oils (AO) and fatty acid distillates (FAD) are oil refining by-products rich in free fatty acids. The objective of this study is their characterization and the identification of their sources of variability so that they can be standardized to improve their use as feed ingredients. Samples (n=92) were collected from the Spanish market and the MIU value (sum of moisture, insoluble impurities, and unsaponifiable matter), lipid classes, fatty acid composition, and tocol content were analyzed. Their composition was highly variable even between batches from the same producer. As FAD originated from a distillation step, they showed higher free fatty acid amounts (82.5 vs 57.0 g/100 g, median values), whereas AO maintained higher proportions of moisture, polymers, tri-, di-, and monoacylglycerols. Overall, the MIU value was higher in AO (2.60-18.50 g/100 g in AO vs 0.63-10.44 g/100 g in FAD), with most of the contents of insoluble impurities being higher than those in the guidelines. Tocol and fatty acid composition were influenced by the crude oil's botanical origin. The calculated dietary energy values were, in general, higher for AO and decreased when a MIU correction factor was applied. The analytical control and standardization of these by-products is of the outmost importance to revalorize them as feed ingredients.

Different Wines from Different Yeasts? "Saccharomyces cerevisiae Intraspecies Differentiation by Metabolomic Signature and Sensory Patterns in Wine".

Alcoholic fermentation is known to be a key stage in the winemaking process that directly impacts the composition and quality of the final product. Twelve wines were obtained from fermentations of Chardonnay must made with twelve different commercial wine yeast strains of Saccharomyces cerevisiae. In our study, FT-ICR-MS, GC-MS, and sensory analysis were combined with multivariate analysis. Ultra-high-resolution mass spectrometry (uHRMS) was able to highlight hundreds of metabolites specific to each strain from the same species, although they are characterized by the same technological performances. Furthermore, the significant involvement of nitrogen metabolism in this differentiation was considered. The modulation of primary metabolism was also noted at the volatilome and sensory levels. Sensory analysis allowed us to classify wines into three groups based on descriptors associated with white wine. Thirty-five of the volatile compounds analyzed, including esters, medium-chain fatty acids, superior alcohols, and terpenes discriminate and give details about differences between wines. Therefore, phenotypic differences within the same species revealed metabolic differences that resulted in the diversity of the volatile fraction that participates in the palette of the sensory pattern. This original combination of metabolomics with the volatilome and sensory approaches provides an integrative vision of the characteristics of a given strain. Metabolomics shine the new light on intraspecific discrimination in the Saccharomyces cerevisiae species.