A novel UHPLC-ESI-MS/MS method and automatic calculation software for regiospecific analysis of triacylglycerols in natural fats and oils.

Regioisomeric analysis of triacylglycerols (TAGs) in natural oils and fats is a highly challenging task in analytical chemistry. Here we present a software (TAG Analyzer) for automatic calculation of regioisomeric composition of TAGs based on the mass spectral data from recently reported ultra-high performance liquid chromatography electrospray ionization tandem mass spectrometry (UHPLC-ESI-MS/MS) method for analyzing TAG regioisomers. The software enables fast and accurate processing of complex product ion spectra containing structurally informative diacylglycerol [M+NH4-RCO2H-NH3]+ and fatty acid ketene [RCO]+ fragment ions. Compared to manual processing, the developed software offers higher throughput with faster calculation as well as more accurate interpretation of chromatographically overlapping isobaric TAGs. The software determines results by constructing a synthetic spectrum to match the measured fragment ion spectrum, and by reporting the optimal concentrations of TAGs used to create the synthetic spectrum. This type of calculation is often extremely challenging for manual interpretation of the fragment ion spectra of isobaric TAGs with shared fragments, hence the need for automated data processing. The developed software was validated by analyzing a wide range of mixtures of regiopure TAG reference compounds of known composition and a commercial olive oil sample. Additionally, the method was also applied for regiospecific analysis of TAGs in human milk as an example of natural fats and oils with a highly complex TAG profile. The results indicate that the software is capable of resolving regioisomeric composition of natural TAGs even of the most complex composition. This novel calculation software combined with our existing UHPLC-ESI-MS/MS method form a highly efficient tool for regioisomeric analysis of TAGs in natural fats and oils.

Impact of cultivar, growth temperature and developmental stage on phenolic compounds and ascorbic acid in purple and yellow potato tubers.

Phenolic compounds and ascorbic acid were analyzed in one yellow and four purple-flesh potato cultivars grown at 13 °C and 18 °C and harvested at different stages of tuber development, using HPLC-DAD and UHPLC-MS. The expression of genes in the phenylpropanoid pathway was studied at transcription level using qPCR. Petunidin-3-p-coumaroylrutinoside-5-glucoside was the most abundant anthocyanin in 'Blue Congo', 'Blaue Schweden', and 'Synkeä Sakari', whereas malvidin-3-p-coumaroylrutinoside-5-glucoside dominated in 'Blaue Veltlin'. In mature tubers, the purple cultivar 'Synkeä Sakari' showed the highest content of anthocyanins (2.4 mg/g freeze-dried sample), and 'Blaue Veltlin' had the highest content of phenolic acids (5.5 mg/g). Cultivar was the main variable affecting the biosynthesis of the studied metabolites, whereas the temperatures studied did not show different impact. The content of the main phenolic acids and anthocyanins in the potato cultivars correlated positively with the expression levels of the genes involved in the phenylpropanoid pathway.

Anthocyanin-rich extract from purple potatoes decreases postprandial glycemic response and affects inflammation markers in healthy men.

Our recent clinical study suggested that polyphenol-rich purple potatoes lowered postprandial glycemia and insulinemia compared to yellow potatoes. Here, 17 healthy male volunteers consumed yellow potatoes with or without purple potato extract (PPE, extracted with water/ethanol/acetic acid) rich in acylated anthocyanins (152 mg) and other phenolics (140 mg) in a randomized cross-over trial. Ethanol-free PPE decreased the incremental area under the curve for glucose (p = 0.019) and insulin (p = 0.015) until 120 min after the meal, glucose at 20 min (p = 0.015) and 40 min (p = 0.004), and insulin at 20 min (p = 0.003), 40 min (p = 0.004) and 60 min (p = 0.005) after the meal. PPE affected some of the studied 90 inflammation markers after meal; for example insulin-like hormone FGF-19 levels were elevated at 240 min (p = 0.001). These results indicate that PPE alleviates postprandial glycemia and insulinemia, and affects postprandial inflammation.

Effects of Latitude and Weather Conditions on Proanthocyanidins in Blackcurrant (Ribes nigrum) of Finnish Commercial Cultivars.

Blackcurrants of three Finnish commercial cultivars 'Mortti', 'Ola', and 'Melalahti' cultivated in southern and northern Finland were compared on the basis of the content and composition of proanthocyanidins (PAs). Seventeen B-type PA oligomers (degree of polymerization 2-5 and 7) were detected by hydrophilic interaction liquid chromatography and electrospray ionization mass spectrometry. Total PAs, dimers, trimers, and tetramers were quantified. Among the three cultivars, 'Ola' had the highest contents of both total PAs and PA oligomers. 'Melalahti' was separated from both 'Mortti' and 'Ola' by PA profiles in the partial least-squares discriminant analysis model. All three cultivars revealed distinct responses to latitude and weather conditions. The content of total PAs showed a positive correlation to latitude in 'Ola' and 'Melalahti'. Among the meteorological variables, high temperature and radiation correlated negatively with total PAs, while only specific variables showed a correlation with PA oligomers.

Effects of a sea buckthorn oil spray emulsion on dry eye.

PURPOSE: To investigate the effects of a sea buckthorn oil and sodium hyaluronate-containing eyelid spray emulsion (SB spray) on dry eye. METHODS: A randomized controlled study was carried out. Adults (25-70 years) with Ocular Surface Disease Index (OSDI) ≥20 and moderate or severe dryness, burning or grittiness of the eyes were included. In study part one (n = 2), SB spray was used on both closed eyelids four times in one day. In part two (n = 10), SB spray was used on one randomized eyelid, and a commercial reference spray on the other for nine days. In part three (n = 40), eyes were randomized to one eye receiving SB spray and an untreated control for 1.5 months. Dry eye tests were carried out at baseline, during, and at the end of each study section. Symptoms were recorded in questionnaires and daily logs. RESULTS: In part one, the SB spray was well tolerated. In part two, OSDI decreased significantly (P = 0.022) in the SB spray eye compared to the reference spray, indicating a beneficial effect on symptoms. In part three, OSDI in the SB spray eye decreased significantly compared to the untreated control (P = 0.0007). The scores for dryness at the study end were lower in the SB spray eye compared to control (P = 0.0070). Symptom sums and frequencies of dryness (sum P = 0.0046, frequency P = 0.0016) and watering (sum P = 0.0003, frequency P = 0.013) in the daily logs were lower in the eye treated with SB spray. CONCLUSIONS: SB spray on closed eyelids relieved the symptoms of dry eye.

Historical Review on the Identification of Mesifurane, 2,5-Dimethyl-4-methoxy-3(2 H)-furanone, and Its Occurrence in Berries and Fruits.

Mesifurane, 2,5-dimethyl-4-methoxy-3(2 H)-furanone, is a natural compound used a worldwide as a flavoring for foods, beverages, and cosmetics. Global sales of mesifurane are around $100 million. Its significance as a flavor-impact compound in some Nordic berries was discovered in the early 1970s in Finland. Synthesized mesifurane was used as a key compound in aroma mixes exploited in a Finnish patent. Mesifurane is a significant flavorant in arctic brambles, mangoes, strawberries, and many other fruits and berries and is an enzymatic methylation product of 2,5-dimethyl-4-hydroxy-3(2 H)-furanone. Because of the obscurity of the information on the history of the commonly used trivial name, mesifurane, it is time to lift the veil and reveal the background of the present situation. The key player was a northern berry, arctic bramble ( Rubus arcticus), the Finnish name of which is mesimarja. Forty years ago, aroma research was limited by technical factors, but nowadays there is a surplus of information.

Red/Green Currant and Sea Buckthorn Berry Press Residues as Potential Sources of Antioxidants for Food Use.

The potential for using extracts of press residues from black, green, red, and white currants and from sea buckthorn berries as sources of antioxidants for foods use was investigated. Press residues were extracted with ethanol in four consecutive extractions, and total Folin-Ciocalteu (F-C) reactive material and authentic phenolic compounds were determined. Radical quenching capability and mechanisms were determined from total peroxyl radical-trapping antioxidant capacity (TRAP) and oxygen radical absorbance capacity (ORAC) assays and from diphenylpicrylhydrazyl (DPPH) kinetics, respectively; specific activities were normalized to F-C reactive concentrations. Levels of total F-C reactive materials in press residue extracts were higher than in many fruits and showed significant radical quenching activity. Black currant had the highest authentic phenol content and ORAC, TRAP, and DPPH reactivity. Sea buckthorn grown in northern Finland showed extremely high total specific DPPH reactivity. These results suggest that berry press residues offer attractive value-added products that can provide antioxidants for use in stabilizing and fortifying foods.

Improved analysis of anthocyanins and vitamin C in blue-purple potato cultivars.

Methods were optimized for extraction and quantification of anthocyanins (ACY) and vitamin C in potatoes. Acidified aqueous methanol (70%) was the optimal extraction solvent and freeze-drying significantly improved the extraction yield of ACY. The content of ACY varied widely in five potato cultivars from 0.42 to 3.18mg/g dry weight, with the latter being the highest value found in the Finnish cultivar 'Synkeä Sakari'. Compared with dithiothreitol (DTT), tris(2-carboxyethyl) phosphine hydrochloride (TCEP) was more efficient in reducing dehydroascorbic acid (DHA) to ascorbic acid (AA) and for quantifying the content of total ascorbic acid (TAA). For raw potatoes, quantification of TAA after treatment with TCEP was more reliable than a direct analysis of AA, whereas AA can be analyzed directly in steam-cooked samples. The TAA contents in the three potato cultivars were around 0.30-0.35mg/g dry weight. The loss of AA in steam cooking was 24%.

Role of Flavonols and Proanthocyanidins in the Sensory Quality of Sea Buckthorn (Hippophaë rhamnoides L.) Berries.

Sensory profile, flavonols, proanthocyanidins, sugars, and organic acids were investigated in purees of six sea buckthorn (Hippophaë rhamnoides) cultivars. The sensory profiles of the purees were dominated by intense sourness followed by astringency and bitterness due to the high content of malic acid. Malic acid and isorhamnetin glycosides, especially isorhamnetin-3-O-sophoroside-7-O-rhamnoside, had close association with the astringent attributes in the different purees, whereas some of the known astringent compounds such as proanthocyanidin dimers and trimers or quercetin glycosides, had less impact. Moreover, the ratios between contents of acids and phenolic compounds were more important predictors of bitterness than the individual variables alone. Astringency and bitterness are important sensory factors for the consumer acceptance of sea buckthorn products. The current study provides new knowledge on the correlations between sensory properties and composition and supports industrial utilization of the sea buckthorn berries.

Triacylglycerol regioisomers in human milk resolved with an algorithmic novel electrospray ionization tandem mass spectrometry method.

A highly sensitive mass spectrometric (MS) method was developed and validated to analyze ratios of regioisomeric triacylglycerols (TAGs) in fats and oils. UPLC resolution of lithiated TAGs followed by daughter scan MS/MS of positive ions revealed several indicative ions for quantitative analysis. Reference TAGs containing C14-C20 fatty acids (FAs) showed good linear response. Analysis of Finnish and Chinese pooled human milk samples revealed hundreds of regioisomeric TAGs. At least 64mol% of the TAGs were quantified with relative standard deviation <17%. When present in the same TAG molecule together with C18 FAs, palmitic acid was typically in the sn-2 position. When together with FAs 10:0, 12:0, 14:0, 20:1 and 20:2, the sn-2 preference of 16:0 was less clear. Oleic acid occupied typically the sn-1/sn-3 positions but when together with FAs 20:1, 20:2, 18:2, 14:1, 12:0 or 10:0 the positioning of 18:1 did not follow these rules.

Effects of latitude and weather conditions on proanthocyanidins in berries of Finnish wild and cultivated sea buckthorn (Hippophaë rhamnoides L. ssp. rhamnoides).

Sea buckthorn (Hippophaë rhamnoides ssp. rhamnoides) of varieties 'Terhi' and 'Tytti' and one of wild origin were cultivated in southern and northern Finland, harvested during 2007-2013. Proanthocyanidins (PAs) were analyzed with HILIC UPLC-ESI-MS. The southern and northern samples were separated in the partial least squares discriminant analysis model (four factors, R(2) 0.75, Q(2) 0.70). The total PAs were more abundant in berries from the north (610-970mg/100gDW) than in those from the south (340-450mg/100gDW) (p<0.05). In northern Finland, the length of the growth season as well as the temperature sum and radiation sum of the growth season until harvest were negatively correlated with the total PAs in all the samples but positively with PA oligomers in 'Tytti' and 'Terhi'. In southern Finland no respective correlations were seen. 'Terhi' and 'Tytti' had different trends in the content of total PA and oligomers in overripe stages.

Phenolic compounds extracted by acidic aqueous ethanol from berries and leaves of different berry plants.

Phenolic compounds of berries and leaves of thirteen various plant species were extracted with aqueous ethanol and analyzed with UPLC-DAD-ESI-MS, HPLC-DAD, and NMR. The total content of phenolics was consistently higher in leaves than in berries (25-7856 vs. 28-711mg/100g fresh weight). Sea buckthorn leaves were richest in phenolic compounds (7856mg/100g f.w.) with ellagitannins as the dominant compound class. Sea buckthorn berries contained mostly isorhamnetin glycosides, whereas quercetin glycosides were typically abundant in most samples investigated. Anthocyanins formed the dominating group of phenolics in most dark-colored berries but phenolic acid derivatives were equally abundant in saskatoon and chokeberry berries. Caffeoylquinic acids constituted 80% of the total phenolic content (1664mg/100g f.w.) in bilberry leaves. B-type procyanidins and caffeoylquinic acids were the major phenolic compounds in hawthorn and rowanberry, respectively. Use of leaves of some species with prunasin, tyramine and ß-p-arbutin, may be limited in food applications.

CO2 Plant Extracts Reduce Cholesterol Oxidation in Fish Patties during Cooking and Storage.

Cholesterol oxidation products (COPs) in foods may pose risks for human health. Suitable antioxidants can reduce the formation of COPs in industrial products. Consumer awareness of food additives has brought a need for more natural alternatives. This is the first study on the effects of supercritical CO2 extracts of rosemary, oregano, and an antimicrobial blend of seven herbs, tested at two levels (1 and 3 g/kg fish), against cholesterol oxidation in patties made of a widely consumed fish species, Atlantic salmon (Salmo salar), during baking and storage. Cholesterol oxidation was reduced by the extracts as indicated by lowered levels of 7α-hydroxycholesterol, 7ß-hydroxycholesterol, and 7-ketocholesterol, which were quantified by GC-MS. The total amount of COPs was smaller in all of the cooked samples containing the plant extracts (<1 µg/g extracted fat) than in the cooked control (14 µg/g). Furthermore, the plant extracts exhibited protective effects also during cold storage for up to 14 days.

Stability of Hydroxycinnamic Acid Derivatives, Flavonol Glycosides, and Anthocyanins in Black Currant Juice.

The stability of phenolic compounds was followed in black currant juice at ambient temperatures (in light and in dark conditions) and at +4 °C for a year. Analyses were based on high-performance liquid chromatography-diode-array detection-electrospray ionization-mass spectrometry (or tandem mass spectrometry) and high-performance liquid chromatography-diode-array detection-electrospray ionization-quadrupole time-of-flight mass spectrometry methods supported by nuclear magnetic resonance after selective high-performance liquid chromatography isolation. Altogether, 43 metabolites were identified, of which 2-(Z)-p-coumaroyloxymethylene-4-ß-d-glucopyranosyloxy-2-(Z)-butenenitrile, 2-(E)-caffeoyloxymethylene-4-ß-d-glucopyranosyloxy-2-(Z)-butenenitrile, 1-O-(Z)-p-coumaroyl-ß-d-glucopyranose, (Z)-p-coumaric acid 4-O-ß-d-glucopyranoside, and (Z)-p-coumaric acid were novel findings in black currant juice. Hydroxycinnamic acid derivatives degraded 20-40% at room temperature during one year of storage, releasing free hydroxycinnamic acids. O-Glucosides of hydroxycinnamic acid compounds were the most stable, followed by O-acylquinic acids, acyloxymethyleneglucosyloxybutenenitriles, and O-acylglucoses. Light induced the isomerization of (E)-coumaric acid compounds into corresponding Z-isomers. Flavonol glycosides stayed fairly stable. Flavonol aglycones were derived mainly from malonylglucosides. Over 90% of anthocyanins were lost at room temperature in a year, practically independent of light. Storage at low temperatures, preferably excluding light, is necessary to retain the original composition of phenolic compounds.

Sea Buckthorn (Hippophaë rhamnoides ssp. rhamnoides) Berries in Nordic Environment: Compositional Response to Latitude and Weather Conditions.

Flavonol glycosides (FGs) in sea buckthorn (Hippophaë rhamnoides ssp. rhamnoides) berries of varieties 'Tytti' and 'Terhi', cultivated in northern Finland (68°02' N) for six years and southern Finland (60°23' N) for seven years, were investigated and compared by HPLC-DAD-ESI-MS/MS. The average total content of 23 identified glycosides of isorhamnetin and quercetin was 103 ± 23 and 110 ± 21 mg/100 g fresh berries in 'Terhi' and 'Tytti', respectively. The total contents of FGs, flavonol diglycosides, and triglycosides in both varieties were higher in the north than in the south, whereas total flavonol monoglycoside content behaved vice versa (p < 0.05). Among the 89 weather variables studied, the sum of the daily mean temperatures that are 5 °C or higher from the start of growth season until the day of harvest was the most important variable which associated negatively with the accumulation of FGs in berries. Such influence was much stronger in berries from the north than from the south.

Proanthocyanidins in Sea Buckthorn (Hippophaë rhamnoides L.) Berries of Different Origins with Special Reference to the Influence of Genetic Background and Growth Location.

Wild sea buckthorn berries from Finland (Hippophaë rhamnoides ssp. rhamnoides) and China (ssp. sinensis) as well as berries of two varieties of ssp. rhamnoides cultivated in Finland and five of ssp. mongolica cultivated in Canada were compared on the basis of the content and composition of proanthocyanidins (PAs). Among all of the samples, only B-type PAs were found. The contents of dimeric, trimeric, tetrameric, and total PAs were in the range of 1.4-8.9, 1.3-9.5, 1.0-7.1, and 390-1940 mg/100 g of dry weight, respectively. The three subspecies were separated by three validated factors (R(2), 0.724; Q(2), 0.677) in the partial least squares discriminant analysis model. Significant differences in total PAs were found between the ssp. rhamnoides and mongolica samples (p < 0.05). In ssp. rhamnoides, samples grown in northern Finland were characterized by a high amount of total PAs, typically 2-3 times higher than that in the level found in southern Finland. In ssp. sinensis, altitude did not have a systematic effect on the PA composition, suggesting the significance of the interaction between genetic background and growth location.

Flavonol Glycosides in Currant Leaves and Variation with Growth Season, Growth Location, and Leaf Position.

Flavonol glycosides (FG) were analyzed in the leaves of six currant cultivars (Ribes spp.) with HPLC-DAD, HPLC-MS/MS, and NMR. The average amounts of the 12 major, identified FG constituted 86-93% (9.6-14.1 mg/g DW) of the total of 27 FG found. Quercetin and kaempferol were the major aglycones with trace amounts of myricetin. Quercetin-3-O-(2,6-α-dirhamnopyranosyl-ß-glucopyranoside), quercetin-3-O-(2-ß-xylopyranosyl-6-α-rhamnopyranosyl-ß-glucopyranoside), and kaempferol-3-O-(3,6-α-dirhamnopyranosyl-ß-glucopyranoside) were identified for the first time in currant leaves and existed in a white currant cultivar 'White Dutch' only. Kaempferol-3-O-ß-(6'-malonyl)glucopyranoside was also a new compound existing in abundance in five cultivars but not in the white one. The results show the primary importance of the genetic background of the cultivars. The content of malonylated FG of special importance in cardiovascular health decreased regularly during summer. Time of collection and leaf position were more prominent factors affecting the composition than were the year of harvest or the growth latitude. Randomly collected leaves differed in their FG profiles from those collected from the middle position of new branches.

Effects of CO2 plant extracts on triacylglycerol oxidation in Atlantic salmon during cooking and storage.

Increasing concern of consumers on the safety of synthetic food additives has created high interest in natural preservatives in food industry. Plant extracts produced by supercritical CO2 technology from rosemary (R), oregano (O) and an antimicrobial blend (AB) consisting of seven different plants were studied for their effects on lipid oxidation in Atlantic salmon (Salmo salar). Fish pieces were marinated with rapeseed oil containing 0, 1, 2 or 4 g of plant extracts/kg of fish. After cooking the pieces were stored in refrigerator for 26 days. Peroxide values (PVs) were determined and oxidised triacylglycerols (TAGs) measured by UHPLC-ESI/MS at 0, 7, 14 and 26 days of storage. During the first two weeks of storage, AB delayed oxidation by at least one week compared to control samples as shown by PVs (<10 meq. O2) and by the oxidised TAGs. Oregano and rosemary showed also some antioxidative potential.

Comparison of the postprandial effects of purple-fleshed and yellow-fleshed potatoes in healthy males with chemical characterization of the potato meals.

The aim of the current study was to characterize the anthocyanin content and composition of a purple potato landrace cultivar (Solanum tuberosum 'Synkeä Sakari') and to compare the postprandial effects of purple-fleshed potatoes, yellow-fleshed potatoes and bilberries in potato starch on postprandial glycemia and insulinemia in healthy males. The purple potato meal caused smaller insulinemia than the yellow potato meal (iAUC 120 min 1347 and 2226, respectively, p = 0.012 and iAUC 240 min 1448 and 2403, p = 0.007) or the bilberry meal (iAUC 120 min 1920, p = 0.027). The purple potato meal caused a smaller plasma glucose at 40 min postprandially compared with the yellow potato meal (p = 0.044). The results of this study suggest that anthocyanin-containing purple-fleshed potatoes influence the postprandial insulinemia positively. Since potatoes are the world's largest non-grain commodity, replacing yellow-fleshed potatoes with purple-fleshed potatoes as staple food could have large potential in maintaining public health.

Effects of sea buckthorn oil intake on vaginal atrophy in postmenopausal women: a randomized, double-blind, placebo-controlled study.

BACKGROUND: Vaginal atrophy, the thinning and drying of vaginal mucosa, is associated with menopause. The standard estrogen treatment is not suitable for all women. OBJECTIVE: To investigate the effects of oral sea buckthorn (SB) oil supplementation on vaginal atrophy. METHOD: A total of 116 postmenopausal women experiencing symptoms of vaginal dryness, itching or burning were randomized to this placebo-controlled, double-blind study. Ninety-eight participants completed the intervention of three months, during which they consumed 3g of SB or placebo oil daily. At the beginning and end, factors of vaginal health were scored by a gynecologist, vaginal pH and moisture were measured and vaginal health index was calculated. Symptoms of atrophy and menopause were evaluated at study visits and by daily logbooks. Serum samples were collected for the analysis of circulating lipids, liver enzymes and C-reactive protein. RESULTS: Compared to placebo, there was a significantly better rate of improvement in the integrity of vaginal epithelium in the SB group when both compliant and noncompliant participants were included (odds ratio (OR)=3.1, 95% CI 1.11-8.95). A beneficial trend was observed when only the compliant participants were included (OR=2.9; 95% CI 0.99-8.35). There was a tendency (P=0.08) toward better improvement of vaginal health index from baseline to the end in the SB group [(0.8 (SD 2.8)] compared to placebo [-0.1 (SD 2.0)]. CONCLUSIONS: SB oil showed beneficial effects on vaginal health, indicating it is a potential alternative for mucosal integrity for those women not able to use estrogen treatment for vaginal atrophy.