Rapid Identification and Relative Quantification of the Phospholipid Composition in Commercial Lecithins by <Superscript>31</Superscript>P NMR

³¹P NMR analysis of samples prepared in a sodium cholate detergent system was used as a method for the identification and quantification of enzymatic hydrolysis products of lecithin. To precisely characterize all of the hydrolysis products from commercial lecithin, a series of enzymatic reactions of each phospholipid with phospholipase PLA1 were conducted and monitored by ³¹P NMR at different times. Twenty-six phosphorus-containing hydrolysis products from six classes of phospholipids (PC, PI, PS, PE, PG, PA) were found and determined by ³¹P NMR measurement. The impact of pH on the chemical shift values for these hydrolysis products was observed and reported. To the best of our knowledge, this is the first report of ³¹P-NMR chemical shift values for entire lyso-phospholipids hydrolyzed from 6 classes of phospholipids. Rapid and routine analysis of phospholipid composition in commercial lecithins by ³¹P NMR was achieved without the need of phospholipid standards.     

Structure and Physical Properties of Organogels Developed by Sitosterol and Lecithin with Sunflower Oil

High linoleic acid sunflower oil (HLSO) with various sitosterol (Sit) to lecithin (Lec) mass ratios (i.e., 0:100–100:0) were used to develop organogels at two storage temperatures (T_s: 5 and 25 °C). The results showed that, at 25 °C, the hardness value of organogels obtained from HLSO with both Sit and Lec was higher than that of organogels developed from HLSO with only Sit or Lec. Microscopy revealed that the shapes of the crystals in the organogels varied significantly with the composition of the structurant and the T_s. At both T_s used, the Sit:Lec (80:20) system had a lower degree of supersaturation compared with the (100:0) system. X‐ray diffraction (XRD) revealed that Sit:Lec mass ratio of 70:30, 80:20 and 100:0 had similar short spacings, and the presence of Lec might be adverse to the formation of Sit crystal in oil. Small‐angel X‐ray scattering (SAXS) showed that the layer thickness of Sit/Lec/HLSO organogel was larger than that of Sit/HLSO organogel. It was found that the presence of Lec induced the change of self‐assembly structure of Sit in HLSO and caused the changes of physical properties of organogels obtained.     

Simple and Efficient Profiling of Phospholipids in Phospholipase D-modified Soy Lecithin by HPLC with Charged Aerosol Detection

Dietary phosphatidylinositol (PI) can be synthesized via phospholipase D (PLD)-catalyzed transphosphatidylation of phosphatidylcholine (PC), abundant in soy lecithin, with myo-inositol. However, a generated mixture of phospholipid (PL) classes poses a challenge for analysis. Our current work on Streptomyces PLD engineering requires a robust analytical method for profiling of PI and related PLs derived from the transphosphatidylation reactions. Therefore, we optimized an HPLC-based method with charged aerosol detector (CAD) for PL quantification. PLs were separated on a normal phase silica column by a gradient elution system using two solvents containing chloroform/methanol/1 M formic acid–triethylamine buffer in different ratios. Retention times of the PL standards and LC–MS under identical conditions were used to identity PL classes. PL standards gave linear response in 100- and 10-fold (lyso-PI) concentration range. The method provided a simple, sensitive, repeatable, and precise analysis of PI, PC, phosphatidylethanolamine, phosphatidic acid, and lyso forms of PC and PI. Compared to the similar existing method, introduction of CAD provided a three- to fivefold decrease at the lower end and a two- to fivefold increase at the upper end of the dynamic range. High precision, high sensitivity, and low limits of detection and quantification further underline the benefits of CAD in PL analysis.     

Fourier Transform Near Infrared Spectroscopy as a Quality Control Tool for the Analysis of Lecithin and By-Products During Soybean Oil Processing

Fourier transform near infrared (FT-NIR) spectroscopy was used to analyze multiple measurement parameters in lecithin production samples and soybean oil refining by-products. For lecithin, partial least squares (PLS) calibration models were developed for acetone insolubles, acid value and moisture and leave-one-out cross validation of the calibration models yielded root mean square error of cross validation (RMSECV) values of 0.37%, 0.59 (mg KOH/g) and 0.050%, respectively. An independent test set consisting of 40% of the lecithin production samples were predicted from the PLS calibration models and a root mean square error of prediction (RMSEP) of 0.41%, 0.53 (mg KOH/g) and 0.056% were obtained for acetone insolubles, acid value and moisture, respectively. Comparison of FT-NIR predictions and corresponding reference method values of 10 lecithin samples using a two-tailed t test showed no significant difference at the p = 0.05 level. A set of 51 samples of soybean oil refining by-products, including acidulated soapstock, fatty acids and black oil, were used for developing PLS calibration models for measuring acid value, moisture and iodine value and leave-one-out cross validations for each model gave values for RMSECV of 6.59 (mg KOH/g), 0.046% and 0.42 (mg I₂/g), respectively. Overall, the results of this study demonstrate the suitability of FT-NIR spectroscopy for the routine analysis of lecithin production samples and soybean oil refining by-products for quality control purposes.     

紫苏葶异构体的NMR定量分析

本文利用核磁共振波谱仪,通过对紫苏葶溶液不同溶剂、不同浓度和不同测试温度的选择,使顺式结构和反式结构的紫苏葶羟基峰在核磁共振一维氢谱中呈现出不同化学位移值,从而对其顺式结构和反式结构进行了定性、定量分析.     

Soy fatty acid oxidation with sodium hypochlorite monitored by nuclear magnetic resonance spectroscopy

The oxidation process of soy fatty acids by sodium hypochlorite with ruthenium trichloride catalyst was examined at different temperatures and active chlorine:fatty acid molar ratios. ¹H and ¹³C distortionless enhancement by polarized transfer nuclear magnetic resonance (NMR) spectroscopy techniques were used to monitor oxidation of the double bonds in unsaturated lipids by measuring the peak integration ratio of double-bond peaks:methylene-methyl peaks. This NMR monitoring technique proved to be an excellent means to quantify double-bond reactions. Gas chromatography-mass spectrometry was used to identify mono- and diacid products, separated by hexane/methylene chloride extraction, as well as other oxidation products. While the presence of ruthenium catalyst increased the initial rate of oxidation, it also catalyzed the decomposition of hypochlorite, decreasing the available reactive chlorine, resulting in a delay in complete oxidation. A 9:1 molar ratio of active chlorine to fatty acids completely oxidized all double bonds of soy fatty acids. However, the yield of low-molecular-weight monoacid oxidation products was only 17%, indicating the probable formation of hydroxy fatty acids.     

Quantification of Fatty Acids and their Regioisomeric Distribution in Triacylglycerols from Porcine and Bovine Sources Using 13C NMR Spectroscopy

The uptake of specific fatty acids in humans is dependent on their position on the glycerol backbone. There is a great interest in methods that can access this information fast and accurately. By way of high-resolution NMR, we have analyzed TAG extracted from pig and beef tissues and obtained quantitative data for the composition and regioisomeric distribution of all major unsaturated fatty acids usually found in these source materials, using a combination of manual integration and deconvolution of ¹³C NMR spectra. In addition, we have developed a method for determining composition and regioisomeric distribution of the two main saturated fatty acids found in pork (16:0, 18:0). The results are discussed in relation to species-specific genetic characteristics of fatty acid and TAG biosynthesis. The developed method could support decisions related to breeding for desired fatty acid profiles, and stimulate further methodology developments using high field NMR.     

Dietary Crude Lecithin Increases Systemic Availability of Dietary Docosahexaenoic Acid with Combined Intake in Rats

Crude lecithin, a mixture of mainly phospholipids, potentially helps to increase the systemic availability of dietary omega‐3 polyunsaturated fatty acids (n‐3 PUFA), such as docosahexaenoic acid (DHA). Nevertheless, no clear data exist on the effects of prolonged combined dietary supplementation of DHA and lecithin on RBC and plasma PUFA levels. In the current experiments, levels of DHA and choline, two dietary ingredients that enhance neuronal membrane formation and function, were determined in plasma and red blood cells (RBC) from rats after dietary supplementation of DHA‐containing oils with and without concomitant dietary supplementation of crude lecithin for 2–3 weeks. The aim was to provide experimental evidence for the hypothesized additive effects of dietary lecithin (not containing any DHA) on top of dietary DHA on PUFA levels in plasma and RBC. Dietary supplementation of DHA‐containing oils, either as vegetable algae oil or as fish oil, increased DHA, eicosapentaenoic acid (EPA), and total n‐3 PUFA, and decreased total omega‐6 PUFA levels in plasma and RBC, while dietary lecithin supplementation alone did not affect these levels. However, combined dietary supplementation of DHA and lecithin increased the changes induced by DHA supplementation alone. Animals receiving a lecithin‐containing diet also had a higher plasma free choline concentration as compared to controls. In conclusion, dietary DHA‐containing oils and crude lecithin have synergistic effects on increasing plasma and RBC n‐3 PUFA levels, including DHA and EPA. By increasing the systemic availability of dietary DHA, dietary lecithin may increase the efficacy of DHA supplementation when their intake is combined.     

Variations in Fatty Acids,Phospholipids and Sterols During the Seed Development of a High Oleic Sunflower Variety

The changes in the content and composition of total fatty acids, phospholipids and sterol esters, and their fatty acids, and of free sterols and tocopherols in developing seeds of a selection of high oleic acid sunflower varieties grown in Bulgaria were examined over a period of 15th to 90th day after flowering by means of various chromatographic methods. Under the climatic and geographical conditions typical for the South-East Balkans phospholipid, sterol-, sterol ester- and tocopherol- species are formed practically completely in the first 15 days after flowering. Until the 90th day, only quantitative changes were detected to give a product with 65% oil content, 1% phospholipids, 0.3% total sterols and 0.09% tocopherols. Oleic acid is the main component in all acyl derivatives, reaching 85% of the total fatty acids while palmitic and stearic acid content is about 4% each. The product is a good quality HOSO with beneficial content of FA and good prospects as a salad and cooking oil.     

Determining the Polymorphic Purity of ε‐CL20 Contaminated by Other Polymorphs through the Use of FTIR Spectroscopy with PLS Regression

CL20 (HNIW=hexanitrohexaazaisowurtzitane), is an energetic molecule that presents four polymorphs (called alpha (α‐), beta (β‐), gamma (γ‐), and epsilon (ε‐)). The ε form is the one used in propellant formulations (the rocket combustive agent). On the one hand, a method developed to measure the purity of this form should be the most reliable one. On the other hand, the amount of impurities – alpha (α‐), beta (β‐), gamma (γ‐), and of the monoacetylatedpentanitrated compound (PN) – should be taken into account in order to assess the influence of the impurities on this propellant. The aim of this work is to develop a quantitative analytical method to measure the polymorphic composition of CL20 using FTIR‐MIR spectroscopy and PLS regression.     

Development of Quantitative Analytical Methods to Measure the Polymorphic Composition of CL20 (Hexanitrohexaazaisowurtzitane – HNIW) with FTIR‐NIR or FTIR‐MIR Spectroscopy using PLS Regression

CL20 (HNIW=Hexanitrohexaazaisowurtzitane), is an energetic molecule that presents four polymorphs (called alpha (α), beta (β), gamma (γ), and epsilon (ε)). Industry is focused to synthesize the ε‐form. For them, it is very important to get an analytical method to quantify the concentration of the ε‐form, which has the best energetic performance. The aim of this work is to develop a quantitative analytical method to measure the polymorphic composition of CL20 using FTIR (mid IR and near IR, MIR/NIR) spectroscopy with PLS regression.     

两种罗汉果多糖的IR及13C NMR分析

利用FT IR和13C NMR技术对罗汉果中的两种多糖SGPS1(Siraitia grosvenorii Polysaccharides,SGPS)和SGPS2进行分析。从SGPS1和SGPS2的红外吸收(4000-500 cm-1)可知,它们具有典型的多糖吸收特征,SGPS1中的单糖残基以吡喃环和呋喃环的形式存在,SGPS2中的单糖残基以α-吡喃环的形式存在。同时运用13C NMR证实了这两种多糖的单糖残基环状构型及糖苷键构型。     

Quantitative Analysis of Dairy Phospholipids by 31P NMR

³¹P NMR analysis of samples prepared in a sodium cholate detergent system was assessed as a method for the quantitative measurement of dairy phospholipids. Major phospholipid (PL) classes measured included: phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), phosphatidylinositol (PI), sphingomyelin (SM) and dihydrosphingomyelin (DHSM). The ³¹P NMR method was validated by comparison with a quantitative two-dimensional thin-layer chromatography (2D-TLC) technique. The 2D-TLC system was more sensitive, able to detect some minor compounds not observed by ³¹P NMR. However, ³¹P NMR is more suited to routine analysis, with sample analysis taking 36 min. The method was also more versatile and sample analysis was possible on high phospholipid containing materials without prior lipid extraction (e.g. buttermilk protein concentrate, beta serum liquid).     

Bimodal Detection of Proteins by 129Xe NMR and Fluorescence Spectroscopy

A full understanding of biological phenomena involves sensitive and noninvasive detection. Herein, we report the optimization of a probe for intracellular proteins that combines the advantages of fluorescence and hyperpolarized ¹²⁹Xe NMR spectroscopy detection. The fluorescence detection part is composed of six residues containing a tetracysteine tag (−CCXXCC−) genetically incorporated into the protein of interest and of a small organic molecule, CrAsH. CrAsH becomes fluorescent if it binds to the tetracysteine tag. The part of the biosensor that enables detection by means of ¹²⁹Xe NMR spectroscopy, which is linked to the CrAsH moiety by a spacer, is based on a cryptophane core that is fully suited to reversibly host xenon. Three different peptides, containing the tetracysteine tag and four organic biosensors of different stereochemistry, are benchmarked to propose the best couple that is fully suited for the in vitro detection of proteins.     

Structural Analysis of Hepta-, Nona-, and Undeca-Cyclic Aromatic Hydrocarbons by NMR Spectroscopy

Large condensed polycyclic aromatic hydrocarbons (LCPAHs) generally have distorted structures due to steric repulsion between neighboring hydrogen atoms. The distortion may influence their physicochemical properties such as electric conductivity, photoconductivity, and photochromism. Accordingly we have synthesized LCPAHs and analyzed their structure. In the present study, we synthesized an undecacyclic aromatic compound, dibenzo[a,rst]-dinaphtho[2,1,8-klm:1′,2′-o]pentaphene (DDPP) and determined the structure by use of NMR spectroscopy. To assign the ¹H- and ¹³C-NMR chemical shifts completely, we used the HMQC-TOCSY (Heteronuclear Multiple Quantum Coherence-Total Correlation SpectroscopY) method in which the original proton-carbon correlation is relayed to neighboring protons in the same spin-system. Furthermore, we compared the chemical shifts (δs) of the heptacyclic aromatic hydrocarbon, peropyrene, the nonacyclic, violanthrene B (VEB), and the undecacyclic, DDPP, and investigated correlations between their local structure and δs. For the protons in the bay region, the averaged δs decrease as molecular size increases: 9.23 ppm (hepta-) > 9.07 ppm (nona-) > 8.77 ppm (undeca-). Similar change in the δs is also found for the protons in the cove region: 9.37 ppm (nona-) > 8.81 ppm (undeca-). Thus, it is noted that the averaged δs for the protons in the cove region become downfield than those for the protons in the bay region. The findings are explained in terms of the anisotropic effect of the local magnetic field of benzene rings.     

Study of Ozonated Sunflower Oil Using 1H NMR and Microbiological Analysis

Prior studies have proven that ozonated vegetable oils present a high germicidal power. Ozonation of sunflower oil at different applied ozone dosage was carried out and peroxide and aldehydes indices along with antimicrobial activity were determined. The reaction products were identified using Proton Nuclear Magnetic Resonance Spectroscopy (¹H NMR). The principal signals intensity values were used for following the reaction course between ozone and sunflower oil. The reaction was following up to peroxide index values of 1202 mmol-equi/kg. The intensities of olefinic proton signals decreased with the gradual increase in ozone concentration but without disappearing completely. The Criegee ozonides obtained at 107.1 mg/g of ozone doses were approximately 3.9-fold higher than that at beginning of the reaction. The aldehyde protons were observed as a weak intensity signal in all the spectra. The signals belonging to olefinic protons from hydroperoxides appeared weak and increased with the increase in ozone doses. Signals from other oxygenated groups were assigned. The highest action spectrum of antimicrobial activity was obtained with the higher peroxide index. It was concluded that at higher applied ozone doses, the higher the antimicrobial activity potential of ozonized sunflower oil     

Fatty Acid,Lipid Class,and Phospholipid Molecular Species Composition of the Soft Coral Xenia sp. (Nha Trang Bay,the South China Sea,Vietnam)

The soft corals of the genus Xenia are common for Indo‐Pacific reef ecosystems. Lipid class, fatty acid (FA), phospho‐ and phosphonolipid molecular species compositions were identified for the first time in the soft coral Xenia sp. from Vietnam. Total lipids consisted predominantly of waxes, monoalkyl diacylglycerols, triacylglycerols, sterols, and polar lipids (21.4, 7.7, 14.2, 10.5, and 36.7 %, respectively). Sesquiterpene alcohol, valerenenol, was found. Acids 16:0, 18:3n‐6, 20:4n‐6, and 20:5n‐3 dominated in total FA. The markers of zooxanthellae (18:4n‐3 and 18:5n‐3) and octocorals (24:5n‐6 and 24:6n‐3) were detected. Acids 18:5n‐3, 20:4n‐6, 22:4n‐6, and 24:5n‐6 concentrated in FA of polar lipids, whereas 14:0, 16:0, 16:1n‐7, 18:2n‐6, and 18:3n‐6 were the major FA of neutral lipids. ChoGpl, EtnGpl, SerGpl, CAEP, PtdIns, and lyso ChoGpl constituted 39.5, 20.8, 20.5, 9.7, 4.3, and 5.3 %, respectively, of the sum of phospho‐ and phosphonolipids. Thirty‐two molecular species of phospholipids and ceramide aminoethylphosphonate (CAEP) were determined by high resolution tandem mass spectrometry. Lyso 18:0e PakCho (4.1 %), 18:0e/20:4 PakCho (20.5 %), 18:1e/20:4 PlsEtn (18.0 %), 18:0e/24:5 PakSer (14.0 %), and 16:0 CAEP (9.6 %) were the major molecular species. EtnGpl and PtdIns mainly consisted of alkenyl acyl and diacyl forms, respectively. Alkyl acyl forms predominated in ChoGpl and SerGpl. Acid 24:5n‐6 was a principal FA in SerGpl, whereas 20:4n‐6 was more abundant in ChoGpl and EtnGpl. PtdIns contained various C_20–24 PUFA. In the context of chemotaxonomy of corals, Xenia sp. has the lipid composition typical for soft corals and the FA profile similar to that of alcyonarians with the high level of 18:3n‐6.     

Direct esterification of fatty acids with phenylalkanols by using dicyclohexylcarbodiimide

A mild one‐pot esterification method of fatty acids with alcohols at room temperature is described. The reaction of undec‐10‐enoic acid (I) with 1‐phenylethanol in the presence of N,N'‐di‐cyclohexylcarbodiimide and 4‐(N,N‐dimethylamino)pyridine gave 1'‐phenylethyl undec‐10‐enoate in quantitative yield. Similar reactions were also carried out with (Z)‐octadec‐9‐enoic acid, (Z)‐12‐hydroxyoctadec‐9‐enoic and (Z)‐9‐hydroxyoctadec‐12‐enoic acids. In addition, compound (I) was allowed to react with diphenyl methanol to form its corresponding ester. The structure elucidation of the prepared esters is based on the elemental analysis and spectral data (IR, ¹H‐ and ¹³C‐NMR, MS).     

协同区间PLS——紫外光谱法同时测定肉桂酸和苯乙酮

结合紫外光谱和化学计量学算法,建立了同时测定混合物中肉桂酸和苯乙酮的方法。常规全谱偏最小二乘(PLS)由于存在与组分无关或呈非线性关系的波长点,使模型预测性能降低。本文采用协同区间偏最小二乘(synergy interval PLS,siPLS)算法选择特征波段,并优化波段组合。与全谱PLS模型比较,结果显示,基于siPLS的方法可以明显简化模型,提高模型的预测精度。研究为肉桂酸和苯乙酮的同时分析提供参考,也为siPLS算法在光谱计量学中应用提供借鉴。     

Self-organization Properties of a GPCR-Binding Peptide with a Fluorinated Tail Studied by Fluorine NMR Spectroscopy

Conjugation of the bioactive apelin-17 peptide with a fluorocarbon chain results in self-organization of the peptide into micelles. Fluorine NMR spectroscopy studies show that the fluoropeptide‘s micelles are monodisperse, while proton NMR indicates that the peptide moiety remains largely disordered despite micellization. A very fast exchange rate is measured between the free and micellar states of the peptide which enables the number of molecules present in the micelle to be estimated as 200, in agreement with values found by dynamic light scattering measurements.