Enzyme-assisted aqueous extraction of fish oil from Baltic herring (Clupea harengus membras) with special reference to emulsion-formation, extraction efficiency, and composition of crude oil.

Enzyme-assisted aqueous extraction (EAAE) is a green, and scalable method to produce oil and protein hydrolysates from fish. This study investigated the role of different parameters on emulsion formation, oil recovery, and the composition of crude oil during EAAE of Baltic herring (Clupea harengus membras). Fatty acid compositions, lipid classes, tocopherols, and oxidation status of the EAAE crude oils were studied. Compared to solvent-extracted oil, EAAE resulted in a lower content of phospholipids accompanied by a 57% decrease in docosahexaenoic acid. Changing fish to water ratio from 1:1 to 2:1 (w/w) with ethanol addition led to the greatest reduction (72%) of emulsion, which resulted in an increase in oil recovery by 11%. The addition of ethanol alone, or reduction of enzyme concentration from 0.4% to 0.1% also reduced emulsion-formation significantly. Overall, emulsion reduction resulted in higher content of triacylglycerols and n - 3 polyunsaturated fatty acids in the crude oil extracted.

Green technologies for production of oils rich in n-3 polyunsaturated fatty acids from aquatic sources.

Fish and algae are the major sources of n-3 polyunsaturated fatty acids (n-3 PUFAs). Globally, there is a rapid increase in demand for n-3 PUFA-rich oils. Conventional oil production processes use high temperature and chemicals, compromising the oil quality and the environment. Hence, alternative green technologies have been investigated for producing oils from aquatic sources. While most of the studies have focused on the oil extraction and enrichment of n-3 PUFAs, less effort has been directed toward green refining of oils from fish and algae. Enzymatic processing and ultrasound-assisted extraction with environment-friendly solvents are the most promising green technologies for extracting fish oil, whereas pressurized extractions are suitable for extracting microalgae oil. Lipase-catalysed ethanolysis of fish and algae oil is a promising green technology for enriching n-3 PUFAs. Green refining technologies such as phospholipase- and membrane-assisted degumming deserve investigation for application in fish and algal oils. In the current review, we critically examined the currently existing research on technologies applied at each of the steps involved in the production of oils rich in n-3 PUFAs from fish and algae species. Special attention was placed on assessment of green technologies in comparison with conventional processing methods.

Enzyme-Assisted Extraction of Fish Oil from Whole Fish and by-Products of Baltic Herring (Clupea harengus membras).

Baltic herring (Clupea harengus membras) is one of the most abundant commercially caught fish species from the Baltic Sea. Despite the high content of fat and omega-3 fatty acids, the consumption of Baltic herring has decreased dramatically over the last four decades, mostly due to the small sizes and difficulty in processing. At the same time there is an increasing global demand for fish and fish oil rich in omega-3 fatty acids. This study aimed to investigate enzyme-assisted oil extraction as an environmentally friendly process for valorizing the underutilized fish species and by-products to high quality fish oil for human consumption. Three different commercially available proteolytic enzymes (Alcalase®, Neutrase® and Protamex®) and two treatment times (35 and 70 min) were investigated in the extraction of fish oil from whole fish and by-products from filleting of Baltic herring. The oil quality and stability were studied with peroxide- and p-anisidine value analyses, fatty acid analysis with GC-FID, and volatile compounds with HS-SPME-GC-MS. Overall, longer extraction times led to better oil yields but also increased oxidation of the oil. For whole fish, the highest oil yields were from the 70-min extractions with Neutrase and Protamex. Protamex extraction with 35 min resulted in the best fatty acid composition with the highest content of eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3) but also increased oxidation compared to treatment with other enzymes. For by-products, the highest oil yield was obtained from the 70-min extraction with Protamex without significant differences in EPA and DHA contents among the oils extracted with different enzymes. Oxidation was lowest in the oil produced with 35-min treatment using Neutrase and Protamex. This study showed the potential of using proteolytic enzymes in the extraction of crude oil from Baltic herring and its by-products. However, further research is needed to optimize enzymatic processing of Baltic herring and its by-products to improve yield and quality of crude oil.

Impact of lactic acid fermentation on sensory and chemical quality of dairy analogues prepared from lupine (Lupinus angustifolius L.) seeds.

Lupine (Lupinus sp.) is a valuable source of plant proteins. There is little knowledge on the impact of food processing on composition and sensory properties of lupine products. In this research, we investigated the impact of fermentation with five starters of lactic acid bacteria on the sensory quality and flavor-active compounds in dairy analogues prepared from sweet lupine (Lupinus angustifolius L.). The sensory qualities of unfermented and fermented products were studied with generic descriptive analysis and affective tests. Acids and sugars were analyzed with GC-FID and volatiles with HS-SPME-GC-MS and GC-O. Fermentation increased sourness and 'vinegar' odor and reduced the 'beany' odor and flavor as well as the unpleasantness of flavor. Formation of volatiles during the fermentation was dependent on the starters. However, all fermentations increased the contents of lactic, acetic, and hexanoic acids, while reducing the contents of hexanal, described as 'grassy' in the unfermented lupine sample.

Effect of Saccharomyces cerevisiae and Schizosaccharomyces pombe strains on chemical composition and sensory quality of ciders made from Finnish apple cultivars.

Composition of volatile compounds and concentrations of sugars and organic acids were studied in apple ciders produced with Saccharomyces cerevisiae and Schizosaccharomyces pombe yeasts using eleven different Finnish apple cultivars. Moreover, sensory quality of selected ciders was studied using check-all-that-apply test with untrained panelists. Seventy-seven volatile compounds were detected in the samples using HS-SPME-GC-MS. In general, the ciders had higher concentrations of higher alcohols, aldehydes, and acetals whereas the juices contained higher contents of C6-alcohols. Simultaneously, fermentation using S. pombe resulted in lower concentrations of malic acid, ethyl pentanoate, ethyl hexanoate, and volatile acids and higher concentrations of residual sugars compared to S. cerevisiae. Ciders made using S. cerevisiae were characterized as 'alcoholic' and 'yeasty' while S. pombe ciders were more frequently described as 'sweet', 'honey-like', and less rated as sour. Besides the strong effect by the yeasts, apple cultivars had significant effects on the compositional and sensorial properties of apple ciders.

Influence of enzymatic treatment on the chemical composition of lingonberry (Vaccinium vitis-idaea) juice.

The effects of different enzymes, their dosages, and incubation times on juice yield and chemical composition of lingonberry juice were assessed. Nonvolatile composition including sugars, acids and phenolic compounds (anthocyanins, hydroxycinnamic acids, flavonols and flavan-3-ols) was analyzed using GC-FID, LC-DAD and LC-MS methods, whereas the volatile compound profile was examined using HS-SPME-GC-MS. Selection of the enzyme had a significant effect on the juice yield by increasing it from 70% without enzyme treatment up to 81%. Enzymes significantly increased the contents of most of the phenolic compounds in the juices but had little impact on the contents of sugars and acids. Heat treatment without enzyme addition also significantly affected the composition. Generally, increasing incubation time increased the contents of various phenolic compounds, especially flavan-3-ols and hydroxycinnamic acids, whereas the effects of dosage were mostly negligible. Enzyme treatment increased the contents of volatile compounds, regardless of dosage or incubation time.

Characterization and Quantification of Nonanthocyanin Phenolic Compounds in White and Blue Bilberry (Vaccinium myrtillus) Juices and Wines Using UHPLC-DAD-ESI-QTOF-MS and UHPLC-DAD.

The nonanthocyanin phenolic compounds in juice and wine produced from fruits of white bilberry, a nonpigmented mutant of Vaccinium myrtillus, and blue bilberry (pigmented variety) were analyzed using liquid chromatography with a diode array detector (LC-DAD) and LC-DAD-electrospray ionization-quadrapole/time of flight hybrid mass spectrometry (ESI-QTOF-MS). On the basis of elution order, UV-vis spectra, accurate mass data, and fragmentation pattern and standards, 42 compounds including 22 phenolic acids, 15 flavonols, and 5 flavan-3-ols, were identified in juices and wines prepared from the two bilberry varieties. The levels of most individual nonanthocyanin phenolic compounds in white bilberry products were significantly lower than those in pigmented ones. In bilberry juices, phenolic acids were the most predominant, accounting for approximately 80% of total phenolic content, with p-coumaroyl monotropeins and caffeic acid hexoside being the major phenolic acids. After fermentation, the total contents of phenolic acids, flavonols, and nonanthocyanin phenolic compounds significantly increased, while the content of total flavan-3-ols decreased significantly. p-Coumaroyl monotropeins still dominated in the wine products, while caffeic acid content showed dramatic elevation with the significant drop of caffeic acid hexoside.

Comparison of Volatile Composition between Alcoholic Bilberry Beverages Fermented with Non-Saccharomyces Yeasts and Dynamic Changes in Volatile Compounds during Fermentation.

The profile of volatile compounds was investigated using headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (HS-SPME-GC-MS) during bilberry juice fermentation with nine non-Saccharomyces yeasts, including Pachysolen tannophilus, Metschnikowia pulcherrima, Hanseniaspora uvarum, Torulaspora delbrueckii, Zygosaccharomyces bailii, Schizosaccharomyces pombe, Lachancea thermotolerans, Issatchenkia orientalis, and Saccharomycodes ludwigii. Dynamic changes in volatile compounds were determined simultaneously with the development of ethanol concentration during fermentation. H. uvarum or I. orientalis produced more ethyl acetate than other yeast strains throughout fermentation, while fermentation with M. pulcherrima resulted in high accumulation of higher alcohols. S. pombe was associated with high productions of pentane-2,3-dione, 3-hydroxybutan-2-one, 2-methylbutanal, and 3-methylbutanal. Among the 59 volatile compounds detected, generally, higher alcohols and monoterpenes accumulated constantly and reached the maximum concentration at the middle or later fermentation stage, whereas aldehydes, ketones, and acetals accumulated first followed by a significant drop. The production and accumulation dynamics of metabolites were highly dependent on the yeast species and the developing ethanol content.

Changes in the volatile profile, fatty acid composition and other markers of lipid oxidation of six different vegetable oils during short-term deep-frying.

Oil deterioration during deep-frying influences the quality of fried foods to a great extent. In this study, the frying performance of six vegetable oils, i.e., hemp, lupin, oat, rapeseed, soy, and sunflower, was evaluated following short-term (60 min) deep-frying of French fries at 180 °C. The frying oils were investigated for fatty acid profile, volatile compound composition, and parameters of oxidative stability, such as iodine, peroxide, and p-anisidine values. The examination showed that the content of Æ©PUFA in hemp oil decreased significantly (p < 0.05) after 60 min of deep-frying, although the degree of change was relatively small (close to 1.5%). Similarly, soy oil presented a fatty acid profile prone to oxidation, and generated the highest level of peroxides at the end of the thermal treatment (PV = 16.6 ±â€¯2.3 mEq O2 kg-1). As for the volatile compound composition of the oils, sunflower oil was extensively affected by the deep-frying treatment with a significant decrease (p > 0.05) in total terpenes, accompanied by a considerable rise in total aldehydes. Oppositely, the proportions of MUFA and PUFA of lupin and oat oils remained stable (p > 0.05) during the short-term deep-frying, indicating high stability of these oils. The research provided new data for evaluating the suitability of these oils for household food preparations.

Effects of processing and storage conditions on volatile composition and odor characteristics of blackcurrant (Ribes nigrum) juices.

Blackcurrant juices were prepared without enzyme (NEB-juice) and with conventional enzyme-aided berry pressing (EB-juice). Juices were pasteurized and stored at ambient temperature (in light and dark conditions) and at +4 °C for a 1-year period of time. Volatile composition and odor attributes were followed by HS-SPME-GC-MS and a sensory panel, respectively. Volatiles were decreased in the EB-juice, showing 100-fold lower contents of the main terpenoids, e.g., α-pinene, δ-3-carene, limonene, terpinolene, bornyl acetate and ß-caryophyllene; whereas esters, such as methyl butanoate and ethyl butanoate, showed 2- and 4-fold lower contents for the EB-juice. Pasteurization produced minor changes in both juices. Volatiles decreased during storage at room temperature while at +4 °C esters and eucalyptol were kept at 100%. The descriptive sensory analysis reported a significant decrease (p < 0.05) of the berry-like odor after 12-month storage at RT. Storage at +4 °C for 12 months did not affect the odor quality.

Bulk industrial fruit fibres. Characterization and prevalence of the original fruit metabolites.

Here we analysed the content of primary and secondary metabolites in nine types of industrially processed fibres derived from the juice industry. Specifically, we examined fibre from: apple, peach, and pear, as non-citrus fruits; the peel and flesh of orange and tangerine, and lemon flesh, as citrus fruits; and carrot, as vegetable. Regarding primary metabolites, the sugar content ranged from 21.6 mg/g in lemon to 290 mg/g in orange peel and lower mass organic acid content ranged from 25.0 mg/g in pear to 250 mg/g in lemon. The content of fatty acids were constant during fibre processing, ranging from 0.5 to 1.46%. Furthermore, the fatty acid profile was not affect for the processing. Concerning secondary metabolites, industrial processing did not decrease the sterols content, which ranged from 0.51 to 1.66 µg/g. Regarding carotenoids, of note was the presence of epoxycarotenoids, which may reflect the quality of the industrial process, thus giving added value to the by-product.

Profiles of Volatile Compounds in Blackcurrant ( Ribes nigrum) Cultivars with a Special Focus on the Influence of Growth Latitude and Weather Conditions.

The volatile profiles of three blackcurrant ( Ribes nigrum L.) cultivars grown in Finland and their responses to growth latitude and weather conditions were studied over an 8 year period by headspace solid-phase microextraction (HS-SPME) followed by gas-chromatographic-mass-spectrometric (GC-MS) analysis. Monoterpene hydrocarbons and oxygenated monoterpenes were the major classes of volatiles. The cultivar 'Melalahti' presented lower contents of volatiles compared with 'Ola' and 'Mortti', which showed very similar compositions. Higher contents of volatiles were found in berries cultivated at the higher latitude (66° 34' N) than in those from the southern location (60° 23' N). Among the meteorological variables, radiation and temperature during the last month before harvest were negatively linked with the volatile content. Storage time had a negative impact on the amount of blackcurrant volatiles.

Development of a SBSE-TD method coupled to GC-MS and chemometrics for the differentiation of variety and processing conditions in peach juices.

Peach juices of distinct varieties, namely yellow- and red-fleshed, and commercial and freshly blended were analyzed. The method used was based on Stir Bar Sorptive Extraction (SBSE) involving a polydimethylsiloxane-coated stir bar with thermal desorption (TD), followed by gas chromatography coupled to mass spectrometry (GC-MS) analysis. The resulting analytical data included 41 compounds belonging to several chemical classes, such as aldehydes, alcohols, lactones, terpenoids, fatty aldehydes, fatty acids and hydrocarbons. Furthermore, chemometric data treatment using unsupervised analysis (PCA) proved useful to classify peach juices on the basis of variety. Stepwise Linear Discriminant Analysis (SLDA) showed that a reduced number of variables (14 compounds), including lactones (6-pentyl-α-pyrone, γ-decalactone, γ-dodecalactone, and δ-dodecalactone), fatty acids (hexadecanoic acid), fatty aldehydes (tetracosanal and octacosanal), hydrocarbons (C23, C26, C27, C29, and C33), and alcohols (phytol and α-tocopherol), were necessary to classify the juice samples according to variety and processing conditions.

Dispersive liquid-liquid microextraction and injection-port derivatization for the determination of free lipophilic compounds in fruit juices by gas chromatography-mass spectrometry.

A method consisting of dispersive liquid-liquid microextraction (DLLME) followed by injection-port derivatization and gas chromatography-mass spectrometry (GC-MS) for the analysis of free lipophilic compounds in fruit juices is described. The method allows the analysis of several classes of lipophilic compounds, such as fatty acids, fatty alcohols, phytosterols and triterpenes. The chromatographic separation of the compounds was achieved in a chromatographic run of 25.5min. The best conditions for the dispersive liquid-liquid microextraction were 100µL of CHCl3 in 1mL of acetone. For the injection-port derivatization, the best conditions were at 280°C, 1min purge-off, and a 1:1 sample:derivatization reagent ratio (v/v) using N-methyl-N-(trimethylsilyl)trifluoroacetamide (MSTFA):pyridine (1:1) as reagent. Quality parameters were assessed for the target compounds, giving a limits of detection (LODs) ranging from 1.1 to 5.7ng/mL and limits of quantification (LOQs) from 3.4 to 18.7ng/mL for linoleic and stearic acid, respectively. Repeatability (%RSD, n=5) was below 11.51% in all cases. In addition, the method linearity presented an r2 ≥0.990 for all ranges applied. Finally, the method was used to test the lipophilic fraction of various samples of commercial fruit juice.

A rapid gas chromatographic injection-port derivatization method for the tandem mass spectrometric determination of patulin and 5-hydroxymethylfurfural in fruit juices.

A novel method consisting of injection-port derivatization coupled to gas chromatography-tandem mass spectrometry is described. The method allows the rapid assessment of 5-hydroxymethylfurfural (HMF) and patulin content in apple and pear derivatives. The chromatographic separation of the compounds was achieved in a short chromatographic run (12.2min) suitable for routine controls of these compounds in the fruit juice industry. The optimal conditions for the injection-port derivatization were at 270°C, 0.5min purge-off, and a 1:2 sample:derivatization reagent ratio (v/v). These conditions represent an important saving in terms of derivatization reagent consumption and sample preparation time. Quality parameters were assessed for the target compounds, giving LOD of 0.7 and 1.6µg/kg and LOQ of 2 and 5µg/kg for patulin and HMF, respectively. These values are below the maximum patulin concentration in food products intended for infants and young children. Repeatability (%RSD n=5) was below 12% for both compounds. In addition, the method linearity ranged between 25 and 1000µg/kg and between 5 and 192µg/kg for HMF and patulin, respectively. Finally, the method was applied to study HMF and patulin content in various fruit juice samples.

Injection-port derivatization coupled to GC-MS/MS for the analysis of glycosylated and non-glycosylated polyphenols in fruit samples.

Polyphenols, including glycosylated polyphenols, were analyzed via a procedure based on injection-port derivatization coupled to gas chromatography-tandem mass spectrometry (GC-MS/MS). The polyphenols in lyophilized fruit samples were extracted with an acidified MeOH mixture assisted by ultrasound. Samples were dried under vacuum, and carbonyl groups were protected with methoxylamine. Free hydroxyl groups were subsequently silylated in-port. Mass fragmentations of 17 polyphenol and glycosylated polyphenol standards were examined using Multiple Reaction Monitoring (MRM) as the acquisition mode. Furthermore, in-port derivatization was optimized in terms of optimal injection port temperature, derivatization time and sample: N-methyl-N-(trimethylsilyl)trifluoroacetamide (MSTFA) volume ratio. A C18 solid-phase-extraction clean-up method was used to reduce matrix effects and injection liner degradation. Using this clean-up method, recoveries for samples spiked at 1 and 10µg/g ranged from 52% to 98%, depending on the chemical compound. Finally, the method was applied to real fruit samples containing the target compounds. The complete chromatographic runtime was 15min, which is faster than reported for recent HPLC methods able to analyze similar compounds.

Comprehensive Two-dimensional Gas Chromatography Time-of-flight Mass Spectrometry to Assess the Presence of α,α-Trehalose and Other Disaccharides in Apple and Peach.

INTRODUCTION: Carbohydrates are important constituents in fruits. Among the carbohydrates, disaccharides have rarely been studied in apple and peach. Indeed, the abiotic stress biomarker and preservation agent α,α-trehalose is a disaccharide. OBJECTIVES: To establish a comprehensive method based on two-dimensional gas chromatography combined with time-of-flight MS detection (GC × GC-ToF/MS) to analyse the disaccharide composition of apple and peach. METHODS: The sample preparation was based on aqueous-methanolic extraction of the analytes, followed by oxime formation and trimethylsilylation of the disaccharides. First, three columns were tested with standards on the one-dimensional system. Next, to perform the sample analysis using GC × GC-MS (which offers significant advantages over conventional GC because it allows higher separation efficiencies), various column configurations were assessed on the two-dimensional system to obtain enhanced separation and low detection limits. The column sets tested included non-polar/semi-polar, semi-polar/polar and polar/non-polar. RESULTS: Using the method that proved to be more efficient, namely the method developed with the semi-polar/non-polar configuration, ten disaccharides were identified, based on analytical standards, retention index and mass spectra. These compounds were quantified in several varieties of apple and peach fruit using the developed GC × GC method and linear curve calibration, resulting in substantial differences among the fruits. However, cultivars within the fruits exhibited no significant differences. CONCLUSION: The proposed method allowed for the identification and quantification of several disaccharides in apple and peach, including the biomarker α,α-trehalose.

Ultra performance liquid chromatography analysis to study the changes in the carotenoid profile of commercial monovarietal fruit juices.

We have developed an analytical method that allows the simultaneous determination of epoxycarotenoids, hydroxycarotenoids and carotenes in monovarietal fresh homemade and industrially processed fruit products. Analyses were carried out using ultra performance liquid chromatography (UPLC). The extraction method was optimized using methanol as the first extraction solvent for lyophilized samples followed by a saponification step. Recoveries ranged between 75% and 104% depending on the compound. Repeatability was better than 10% for all compounds (%RSD, n=3). The chromatographic analysis takes less than 17min. In this short period, up to 27 carotenoids were identified in apple, peach and pear products. The developed method allowed us to differentiate juice from six varieties of apple by their carotenoid profile. Moreover, the methodology allows us to differentiate the carotenoid profiles from commercial juices and homemade fresh peach and pear juices, as well as to study the rearrangements of 5,6- to 5,8-epoxycarotenoids.