Classification of olive oils according to geographical origin by using H NMR fingerprinting combined with multivariate analysis

Authentic extravirgin olive oils from 7 different regions (Italy – 3 regions, Greece – 4 regions) have been investigated by ¹H Nuclear Magnetic Resonance (NMR) fingerprinting in combination with multivariate statistical analysis. In order to cover the dominating lipid signals as well as signals from compounds of low abundance in the oil, both a simple one pulse experiment and an experiment with multiple saturation of the lipid signals was applied to each sample. Thus, the dynamic range of concentrations covered by the two experiments was of the order of 100,000 allowing for a more comprehensive NMR assessment of the samples. Monte-Carlo embedded cross-validation was used to demonstrate that a combination of principal component analysis, canonical analysis, and classification via nearest class mean can be used to predict the origin of olive oil samples from ¹H NMR data. Given the rather limited number of samples tested, correct prediction probabilities of 78% were achieved with region specific correct predictions between 53% and 100%.     

Classification of Brazilian honeys by physical and chemical analytical methods and low field nuclear magnetic resonance (LF H NMR)

This study evaluated the possibility of differentiating the botanical origin of honeys using Low Field Nuclear Magnetic Resonance (LF ¹H NMR). Eighty samples of honey from 8 different botanical sources (eucalyptus, “assa-lipto”, oranges, Barbados cherry, cashew tree, “assa-peixe”, “cipó-uva” and polyfloral) were analyzed. A close correlation (p < 0.01) was established between the LF ¹H NMR analysis and physical and chemical measurements, including water content, water activity, pH and color. Bi-exponential fitting of the transverse relaxation (T₂) data revealed two water populations in all samples, T₂₁ and T₂₂, corresponding to relaxation times of 0.6–1.8 ms and 2.3–5.4 ms respectively. The observed differences in the relaxation times suggest that these were influenced by the differences in botanical origins. Good linear correlations were observed between the T₂ and T₂₁ parameters and the physical and chemical data. This study demonstrated that LF ¹H NMR can be a viable technique for use in classifying honeys by their botanical origin.     

Biochemical monitoring of black raspberry (Rubus coreanus Miquel) fruits according to maturation stage by H NMR using multiple solvent systems

Nuclear magnetic resonance (NMR) techniques coupled with multivariate data analysis were used to conduct monitoring of biochemical changes of black raspberry fruits at different stages of maturation and under various extraction and NMR dissolution solvent conditions: extraction with 50% methanol and D₂O as an NMR dissolution solvent, extraction with 50% methanol and 50% methanol-d₄ as an NMR dissolution solvent, and extraction with 100% ethyl acetate and 100% methanol-d₄ as an NMR dissolution solvent. Partial least-squares discriminant analysis reliably distinguished black raspberry fruits according to the maturation stage, whereby the relative levels of various compounds such as amino acids, organic acids, sugars and phenolic compounds were compared using analysis of variance. Sucrose and most of the amino acids, and organic acids decreased, whereas fructose, glucose, and cyanidins increased in relative concentration according to maturation of black raspberry fruits. The total number and kinds of assigned compounds of the three solvent systems were also compared. This research demonstrates that the metabolic profile of black raspberry fruits changes during maturation, and provides objective criteria for determining the stage of black raspberry fruit maturation via a ¹H NMR-based metabolomics technique using multiple solvent systems.     

Compositional study of different soybean (Glycine max L.) varieties by H NMR spectroscopy,chromatographic and spectrometric techniques

Compositional study of different soybean varieties was performed by use of ¹H nuclear magnetic resonance (NMR) spectroscopy, chromatographic and spectrometric technique. Compositions of amino acids, fatty acids, sugars, elements, and isoflavones in two glyphosate-tolerant soybean varieties and three Chinese conventional soybean varieties were studied by ¹H NMR spectroscopy, gas chromatography-mass spectrometry, high performance liquid chromatography and inductively coupled plasma-atomic emission spectrometry, respectively. Principal component analysis suggested that glyphosate-tolerant and conventional soybeans had different compositional profile characteristics of amino acids and fatty acids. Then, the contents of some typical soybean components involving main nutrients and antinutrients were compared. As key nutrients of glyphosate-tolerant soybeans, the content of crude protein increased at 8.9–40%, while the contents of α-tocopherol and γ-tocopherol decreased at 12–64%. As antinutrients of glyphosate-tolerant soybeans, the content of tannin decreased at 32–51%, while the content of raffinose increased at 63–197%, compared with conventional soybeans. Systematical study of compositional profile characteristics provided an effective method for discriminating different soybean varieties and useful reference values for soybean consumption.     

Characterisation of the lipidic components of margarines by H Nuclear Magnetic Resonance

In this work, ¹H Nuclear Magnetic Resonance (¹H NMR) has been used to study the lipidic fraction of margarines of different compositions, determining simultaneously both major components, which is to say triglycerides, and other minor ones, such as 1- and 2-monoglycerides, 1,2-diglycerides, vegetable stanols and sterols, and sorbic acid. The results show a wide variety of acyl group compositions, with polyunsaturated groups ranging from 22% to 50%, monounsaturated from 23% to 50%, and saturated from 21% to 55%. 1,2-Diglycerides and vegetable stanols and/or sterols have been detected in all the studied margarines, in concentrations varying between 2.73 and 26.06 mmol/kg, and between 3.40 and 240.01 mmol/kg, respectively. A good agreement has been found between the results obtained by ¹H NMR and some composition data of these margarines, showing the usefulness of this technique to analyse the lipidic composition of margarine in a quick and easy way.     

Study of the suitability of HRMAS NMR for metabolic profiling of tomatoes: Application to tissue differentiation and fruit ripening

The feasibility of HRMAS NMR as an efficient technique for metabolic studies of tomato fruit and tomato tissues is described. A good-taste tomato variety from Almería (Spain) (commercial name Zayno) was chosen for this study. Fruits of three different ripening stages (green, turning and red) were selected, based on their external appearance. NMR data of whole fruits were obtained through ¹H HRMAS NMR spectra of tomato purée. Flesh, peel and seeds from mature red fruits were separately analysed. ¹H HRMAS NMR spectra showed resolutions similar to that of solution ¹H NMR with the advantages of minimal sample manipulation and possibility of analysing polar and non-polar compounds simultaneously. Seeds could be clearly distinguished by visual comparison of the spectra. Tryacylglycerols, in which linoleic (47%) and oleic (28%) acyl chains preponderate, were the major components of seeds. By contrast, signals of fructose, glucose, citric acid, and the amino acids GABA, glutamine and glutamate dominated the spectra of flesh, peel and tomato purée. Principal components analysis, performed on the covariance matrix, revealed a clear separation between peel and flesh. Apart form the presence of the cutin biopolymer, peel showed a specific accumulation of sugar and sugar moieties related to pectin degradation, as well as glycosylated metabolites. Chemometric analysis also allowed the metabolic trajectory from green to red mature stages to be visualised. However, sample heterogeneity in the analysis of the whole fruit proved to be an important issue. The source of such heterogeneity was assigned to the presence of tissues of different hardness in the small amount of sample used. The presence of two clusters of samples at the turning stage indicated that the levels of certain metabolites could diverge significantly in fruits with almost identical appearance.     

Comparative NMR relaxometry of gels of amylomaltase-modified starch and gelatin

With the aim of generating gelatin-like starch gel functionality, starches extracted from normal potato, high amylose potato, maize, waxy maize, wheat and pea and oxidized potato starch were modified with amylomaltase (AM) (4-α-glucanotransferase; E.C. 2.4.1.25) from Thermus thermophilus. Gel characteristics after storage for 1 and 10 days at 20 °C of 12.0% gels were assessed by monitoring proton relaxation for the resulting 51 enzyme-modified starches and two gelatins using low-field ¹H nuclear magnetic resonance (LF NMR) relaxometry. Discrete and distributed exponential analysis of the Carr–Purcell–Meiboom–Gill (CPMG) LF NMR relaxation data revealed that the pastes and gels contained one water component and that the spin–spin relaxation time constants (T₂) and distributions differed with respect to starch type and enzyme modification. Typically, AM modification resulted in starches with decreased T₂ relaxation time and a more narrow T₂ distribution indicating a more homogeneous water population. In contrast, treatment with a branching enzyme (BE) (EC 2.4.1.18) combined with AM increased T₂ relaxation time and a broadened T₂ distribution. As evaluated by the principal component analysis (PCA), long chains of amylopectin generated hard gels and decreased T₂ relaxation time at both day 1 and day 10. Especially at day 10, T₂ relaxation time could be predicted from the amylopectin chain length (CL) distribution. Reconstructed amylopectin CL distribution required to emulate gelatin LF NMR data suggest the importance of combined fractions of long (DP 60–80) and short (DP 10–25) amylopectin chains.     

Water properties and salt uptake in Atlantic salmon fillets as affected by ante-mortem stress,rigor mortis,and brine salting: A low-field H NMR and H/Na MRI study

A low-field (LF) ¹H NMR T₂ relaxation and ²³Na/¹H MRI study was performed on Atlantic salmon to study the effect of ante-mortem handling stress and rigor mortis on muscle water properties and subsequent post-rigor salting. Compared to rested fish, exhausted fish exhibited a more rapid and stronger development of rigor mortis. This resulted in significant differences in post-rigor water-holding capacity and salt uptake. By LF NMR T₂ relaxation analysis significant differences in water distribution according to (1) ante-mortem handling, (2) fillet location, and (3) brine salting were detected. Furthermore, ²³Na MRI revealed differences between the two treatments in fillet salt distribution, where the salt penetration in exhausted fillets was more pronounced. By combining ¹H and ²³Na MR images, the salt diffusion and distribution seemed to be highly affected by the distribution of fat.     

A NMR metabolomics study of the ripening process of the Fiore Sardo cheese produced with autochthonous adjunct cultures

Fiore Sardo (FS) is a traditional Italian raw ewe’s milk cheese carrying a Protected Designation of Origin (PDO). This study investigated the kinetics of FS cheese ripening by physicochemical parameters, microbial counting, and NMR metabolomics using aqueous extracts. Four Fiore Sardo cheeses, manufactured from milk with deliberately added autochthonous lactic acid bacteria (LAB) or commercial starters were studied during a period of 90 days of ripening. Major differences in the metabolic profiles were observed amongst the samples as a function of the adjunct culture utilised. ¹H NMR metabolomics in combination with multivariate data analysis was able to classify cheese samples on the basis of their maturation age and the type of added cultures. These findings lay the metabolic basis for the authentication of Fiore Sardo cheese produced in compliance with PDO specifications which allow the use of only native LAB cultures.     

Structural characterisation of a heteropolysaccharide by NMR spectra

An undocumented water-soluble polysaccharide, LZ-C-1, was isolated from the fruiting bodies of the fungus, Ganoderma lucidum. The polysaccharide had a molecular weight of 7 × 10³ Da, and was mainly composed of l-Fuc, d-Glc and d-Gal. LZ-C-1 had a sugar content of ∼96.4% as measured using the phenol–sulphuric acid method. As a precondition to understand the bioactivity, structural features of LZ-C-1 were investigated by a combination of total hydrolysis, methylation analysis, FT-IR and NMR studies. The results indicated that LZ-C-1 had a backbone of 1,6-disubstituted-α-galactopyranosyl, 1,2,6-trisubstituted-α-galactopyranosyl, 1,3-disubstituted-β-glucopyranosyl and 1,4,6-trisubstituted-β-glucopyranosyl residues. The branches were mainly composed of 1-substituted-β-glucopyranosyl and 1-substituted-α-fucopyranosyl residues.     

利用模式识别技术鉴别中国葡萄酒的产地和品种

通过测定样品的物理化学指标来辨别中国葡萄洒的品种和产地.本实验借助葡萄洒分析仪和紫外可见分光光度计,共测了20项物理化学指标.33个样品来自3个不同的产地,34个样品来自3个不同品种.方差分析破用来鉴别具有显著差异的变量.主成分分析和逐步判别分析被用来对数据进行进一步的分类分析.最终,通过典则判别分析分别建立了有效的产地鉴别和品种鉴别的判别模型,并且交义检验被用来对该模型的预测能力进行估计,正确率分别为93.94%和100%.     

Water molecular dynamics during bread staling by Nuclear Magnetic Resonance

Bread staling is a complex phenomenon that originates from multiple physico-chemical events (amylopectin retrogradation, water loss and redistribution) that are not yet completely elucidated. Molecular properties of white bread loaves were characterized by multiple proton Nuclear Magnetic Resonance (NMR) techniques (proton FID, T₂ and T₁ relaxation time) over 14 days of storage. Changes at a molecular level (faster decay of proton FIDs and shifting of proton T₂ relaxation times distributions towards shorter times), indicating a proton mobility reduction of the bread matrix, were observed during storage. Multiple ¹H T₂ populations were observed and tentatively associated to water-gluten and water-starch domains. Proton T₁ of bread was for the first time measured at variable frequencies (Fast Field Cycling NMR) and found to be strongly dependent upon frequency and to decrease in bread during storage, especially at frequencies ≤ 0.2 MHz. An additional proton T₁ population, relaxing at 2 ms, was detected at 0.52 MHz only at early storage times and tentatively attributed to a water-gluten domain that lost mobility during storage.     

白酒模拟体系氢键缔合时效性核磁共振研究

采用氢谱核磁共振测定了40%和60%乙醇含量白酒模拟体系在190d时间范围内的氢键缔合情况。结果表明,随着储藏时间的延长,模拟体系的羟基质子峰不断向低场移动,说明体系内氢键缔合逐渐增强;不同乙醇含量的白酒模拟体系氢键缔合强度不同,低乙醇含量溶液的氢键缔合整体强于高乙醇含量溶液;而且二者出现缔合最强峰的时间也不相同,高乙醇含量溶液比低乙醇含量溶液较晚出现最强缔合。     

Evaluation of antioxidant activity and total phenols index in persimmon vinegars produced by different processes

The total phenols index (TPI) and antioxidant activity of persimmon vinegars produced by different processes were evaluated. A novel extraction method was designed and optimised for this purpose with respect to the type and concentration of solvent and ultrasonication time. The best extraction conditions found were the use of 80% ethanol and 25 min of ultrasonication. Antioxidant capacity was determined by the oxygen-radical absorbance capacity of fluorescein (ORAC-FL) and 2,2′-diphenyl-1-pycrylhydrazyl (DPPH) free-radical assays. The antioxidant activities were the same in the fruit and the vinegar, except in the ORAC assay, which showed a significant decrease during the acetification process. The results showed that using the wild yeast strain native to the persimmon produced vinegars with higher antioxidant activity than that of an inoculated alcoholic fermentation. Finally, a comparison between our vinegars and other commercial examples was made. The TPI and antioxidant activity values of persimmon vinegars were always higher than those obtained from white and red-wine vinegars. The antioxidant activity and total phenols of the final product indicate that persimmon vinegar is a competitive product in the market.     

Oxidation of corn oil at room temperature: Primary and secondary oxidation products and determination of their concentration in the oil liquid matrix from H nuclear magnetic resonance data

The oil liquid matrix of several corn oil samples that have been stored at room temperature in closed receptacles for different periods of time is studied by means of ¹H nuclear magnetic resonance (¹H NMR), in order to further knowledge about this type of edible oil oxidation. As expected, the degradation of linoleic acyl groups predominates. In samples at early oxidation stages the presence of hydroperoxides and of (Z,E) conjugated-dienic systems is demonstrated, the concentration of the first group being higher than that of the second. In addition to these compounds, the presence of hydroxy derivatives supporting (Z,E) conjugated-dienic systems, as well as of hydroperoxy derivatives supporting (E,E) conjugated-dienic systems, in samples at intermediate and advanced oxidation stages, is also shown. Corn oil samples at advanced stages of oxidation also contain aldehydes, among which there are alkanals, (E)-2-alkenals, (E,E)-2,4-alkadienals, 4-hydroxy-(E)-2-alkenals, 4-hydroperoxy-(E)-2-alkenals and 4,5-epoxy-(E)-2-alkenals. The concentrations of the different kinds of intermediate compounds above mentioned as well as of the different kinds of aldehydes present in the oil liquid matrix were determined. These latter compounds can be either free or joined to truncated structures of triglycerides. In addition, a principal component analysis between storage conditions and oxidation level of the samples was carried out.     

Effect of different mixers on physicochemical properties and water status of extruded and laminated fresh pasta

Physicochemical properties and water status of fresh pasta (both extruded and laminated) produced with recently designed mixers (Premix^® and Bakmix^®) that induce a uniform hydration of the solids and allow the formation of a dough in 1–2 s were evaluated.     

P NMR spectroscopic investigations of caseins treated with microbial transglutaminase

Caseins - the main constituents of bovine milk proteins - self-assemble into large supramolecular aggregates, so-called casein micelles. The enhancement of the stability of casein micelles is advantageous with respect to technological milk processing. A promising approach to accomplish this goal is the cross-linking of caseins using microbial transglutaminase (mTG). The present paper describes the combined use of liquid- and solid-state ³¹P NMR spectroscopy as well as dynamic light scattering in order to characterize the influence of an mTG treatment upon the structure of micelles in ultrahigh temperature (UHT)-treated skim milk at a molecular level. Liquid-state ³¹P NMR spectroscopy was applied to characterize milk, milk serum and casein dispersions. A narrow SerP signal in the liquid-state ³¹P NMR spectra of UHT-treated milk is shown to be due to casein molecules in the milk serum whereas the casein molecules bound in the micelles give rise to broad signals. Most of the caseins contribute to a 3 kHz broad background signal which can be visualized in the spectrum of suspensions of re-dispersed micellar material derived from UHT-treated milk. Treatment with mTG results in a cross-linking of caseins, which could be followed by liquid-state ³¹P NMR spectroscopy. Especially, the cross-linking of β-casein was demonstrated by quantitative liquid-state ³¹P NMR experiments. Furthermore, the stability of cross-linked micellar aggregates against EDTA could be investigated by liquid-state ³¹P HR NMR in combination with dynamic light scattering (DLS). Solid-state ³¹P NMR was used to show that the motional state of the κ-caseins located at the outer surface of the micelles derived from UHT-treated milk is not significantly changed by the applied mTG treatment.     

Enzymatic synthesis of primeverosides using transfer reaction by Trichoderma longibrachiatum xylanase

Enzymatic synthesis of two phenyl xylopyranosyl glucopyranosides, through transfer reaction by Trichoderma longibrachiatum endoxylanase, was achieved in the presence of n-hexane used as solvent, phenyl glucoside (10 mM) as acceptor and xylan (2 g/l) as donor. Kinetic study showed that only one compound, identified by ¹H and ¹³C NMR and heteronuclear 2D (¹H–¹³C) chemical shift correlation as phenyl primeveroside (phenyl 6-O-β-xylopyranosyl-1-β-d-glucopyranoside), was synthesized when the reaction time was beyond 1 h. Benzyl and hexyl primeverosides were obtained under the same conditions. When several phenyl glucoside concentrations, from 5 to 50 mM, were used with 2 g/l of xylan, a phenyl primeveroside isomer, identified as phenyl 4-O-β-xylopyranosyl-β-d-glucopyranoside, accumulated in the medium whereas the production of phenyl primeveroside decreased. Only phenyl primeveroside was produced when several xylan concentrations from 2 to 10 g/l were used with 10 mM of phenyl glucoside and its concentration in the reaction mixture increased with the increase of xylan concentration.     

Effect of Different Drying Treatments and Solvent Ratios on Phytochemical Constituents of Ipomoea aquatica and Correlation with α-Glucosidase Inhibitory Activity

Ipomoea aquatica is an aquatic plant that is widely consumed in Southeast Asia as a vegetable. In this study, the influence of various ethanol ratios (0, 20, 50, 80, and 100%) as an extraction solvent and different drying methods including air drying, sun drying, and oven drying on phytochemical constituents of I. aquatica was investigated using a proton nuclear magnetic resonance-based metabolomics approach. The effect on α-glucosidase inhibitory activity and total phenolic content was also examined. Clear discrimination was observed between different ethanol ratios and different drying processes by principal component analysis. The highest α-glucosidase inhibitory activity was observed for absolute ethanol extract from the oven drying method with IC₅₀ value of 204.0 ± 59.0 µg/mL and total phenolic content value of 22.0 ± 0.7 µg gallic acid equivalent/mg extract. Correlation between the α-glucosidase inhibitory activity and the metabolite were analyzed using a partial least square analysis. The metabolites that are responsible for the activity were quercetin derivatives, chlorogenic acid derivatives, sucrose, and fructose. This study highlights the basis for future investigations of I. aquatica as a source of food that has the potential for nutraceutical enhancement and as ingredient in medicinal preparation.