乳酸菌组合发酵菌种配方及其增殖培养基的优化

为了将甘肃牧区优良乳酸菌株用于直投式酸奶发酵剂的生产,以从甘肃牧区分离所得的乳酸乳球菌乳脂亚种(HM 598685)、嗜酸乳杆菌(HM 598684)、干酪乳杆菌(HM 598683),按照1:0:0、0:1:0、0:0:1、1:1:0、1:2:0、2:1:0、1:0:1、1:0:2、2:0:1、1:1:1、1:2:1、2:1:1、1:1:2的比例进行混合发酵,通过所制酸奶的感官性状和生化特性确定组合发酵的菌种配方,并对混合发酵的增殖培养基进行优化。结果发现,当3株菌比例为1:1:1时,其混合发酵的凝乳时间最短,为8.5h;所得酸奶的活菌数最大,高达7.90×10~8CFU/mL;而其感官评价中滋味气味及组织形态均显著优于其他组合。根据美蓝实验,当基础培养基中使用胰蛋白酶水解乳蛋白时,培养所得乳酸菌的活性最高,且活菌数可达2.93×10~8CFU/mL。通过L_9(3~4)正交设计,确定最佳增殖因子为酵母粉1%、果糖0.5%、吐温0.2%、乙酸钠0.5%,此时培养所得混合菌活菌数达2.11×10~9CFU/mL;各增殖因子对活菌数影响的主次顺序为:酵母粉>乙酸钠>果糖>吐温。     

氯霉素免疫抗原及包被抗原的制备

采用混合酸酐法合成30:1～10:1 三个不同起始物质的量比的氯霉素免疫抗原(HAP-KLH),采用活泼酯法(EDC-NHS)合成40:1～1:1 七个不同起始物质的量比的包被抗原(HAP-OVA),紫外测定HAP-KLH 偶联比25:1～8:1,HAP-OVA 偶联比23:1～1:3。间接竞争ELISA 实验优化包被抗原,在23:1～3:1 偶联比范围内,氯霉素(CAP) IC50值为36～15ng/mL,随着偶联比的减小而降低,当偶联比低于3:1 则略为上升。确定了包被抗原HAP-OVA 最佳偶联比为3:1。本研究表明包被抗原的偶联比对ELISA 测定IC50 值有重要影响,应合成不同偶联比的包被抗原,选出最佳偶联比。     

The association of lipophilic phospholipids with native bovine casein micelles in skim milk: Effect of lactation stage and casein micelle size

A known biological role of casein micelles is to transport calcium from mother to young and provide amino acids for growth and development. Previous reports demonstrated that modified casein micelles can be used to transport and deliver hydrophobic probes. In this study, the distribution of lipid-soluble phospholipids, including sphingomyelins (SM) and phosphatidylcholines (PC), was quantified in whole raw milk, skim raw milk, and casein micelles of various sizes during early, mid, and late lactation stages. Low-pressure size exclusion chromatography was used to separate casein micelles by size, followed by hydrophobic extraction and liquid chromatography–mass spectrometry for the quantification of PC and SM. Results showed that the SM d18:1/23:0, d18:1/22:0, d18:1/16:0, d16:1/22:0, d16:1/23:0, and d18:1/24:0 and the PC 16:0/18:1, 18:0/18:2, and 16:0/16:0 were dominating candidates appearing in maximum concentration in whole raw milk obtained from late lactation, with 21 to 50% of total SM and 16 to 35% of total PC appearing in skim milk. Of the total SM and PC found in skim milk, 35 to 46% of SM and 22 to 29% of PC were associated with the casein micelle fraction. The highest concentrations of SM d18:1/22:0 (341 ± 17 µg/g of casein protein) and PC 16:0/18:1 (180 ± 20 µg/g of casein protein) were found to be associated with the largest casein micelles (diameter = 149 nm) isolated in milk from late lactation, followed by a decrease in concentration as the casein micelle size decreased.     

Plant-Derived Supplementary Carbohydrates,Polysaccharides and Oligosaccharides in Management of Diabetes Mellitus: A Comprehensive Review

ABSTRACT

Background and aims: Diabetes mellitus is a chronic heterogenic disease characterized by the deregulation of the metabolism of insulin and glucose. The aim of this review has been to evaluate the efficacy of medical plant-based carbohydrates, excluding monosaccharides, to manage glycemic response in clinical trials.

Methods: The range of literature presented was compiled by searching electronic databases, including Scopus, PubMed and Cochrane library, from their inception through to June 2018. Only clinical-based studies were considered for this review. Dietary carbohydrates were investigated, especially those containing fiber possessing beneficial effects in the management of hyperglycemia. The most common oligosaccharides, including xylooligosaccharide, isomaltooligosaccharide, fructooligosaccharide, galactooligosaccharide, and sucrose, were able to manage glycemic and insulin metabolism in clinical trials.

Results: In light of 77 selected papers, several plant-based oligosaccharides and polysaccharides have been shown to significantly improve the management of glucose and insulin metabolism in healthy and diabetic patients.

Conclusions: Oligosaccharides and polysaccharides derived from plants possess promising hypoglycemic potential, similar to and even more effective than current synthetic drugs, with no deleterious side effects.

Abbreviations: AMPK: AMP-activated protein kinase; AX: Arabinoxylan; CAT: Catalase; CFDA: Chinese food and drug administration; DM: Diabetes mellitus; FBS, Fasting blood glucose; FDA: United States food and drug administration; FOS: Fructooligosaccharide; G6Pase: Glucose-6- phosphatase; GCK: glucokinase; GD: Gestational diabetes; GI: Glycemic index; GL: Glycemic load; Glut4: Glucose transporter 4; GOS: Glucooligosaccharide; GPx: glutathione peroxidase; GRAS: generally regarded as safe HCF: high-carbohydrate, high fiber diet; HNF1α: Hepatic nuclear factor 1 alpha, HNF4α: Hepatic nuclear factor 4α; HNF1β: Hepatic nuclear factor 1β; II: Insulinemic index; IOS: Isomaltooligosaccharides; KGM: Konjac glucomannan; LBP: L. barbarum polysaccharide; MODY: Maturity onset diabetes of the young; PECK: phosphoenolpyruvate carboxykinase; RDS: Rapidly digestible starch; RS: Resistant starch; SBOS: Soybean oligosaccharides; SDS: Slowly digestible starch; SNP: Single nucleotide polymorphism; SOD: Superoxide dismutase; T1DM: Type 1 diabetes; T2DM: Type 2 diabetes mellitus; TBARS: Thiobarbituric acid reactive substances; XOS: Xylooligo-saccharide.     

Composition and thermal analysis of ternary mixtures of avocado fat:palm stearin:cocoa butter (Avo:PS:CB)

Avocado fat is a semi-solid substance with potential functional lipid characteristics. A study was carried out to evaluate the effect of addition of palm stearin and cocoa butter on the solidification behavior of avocado fat to formulate a mixture to become similar to lard. A total of three mixtures were prepared: avocado fat:palm stearin:cocoa butter (88:7:5), avocado fat:palm stearin:cocoa butter (86:7:7), avocado fat:palm stearin:cocoa butter (84:7:9; w/w), and identified by the mass ratio of avocado fat to palm stearin and cocoa butter. The fat mixtures were compared with lard in terms of the fatty acid and triacylglycerol compositions using gas chromatography and high-performance liquid chromatography, thermal properties using differential scanning calorimetry and solid fat content using p-nuclear magnetic resonance. Although there were considerable differences between lard and the fat mixtures with regard to fatty acid and triacylglycerol compositions, some similarities were seen with regard to thermal properties and solid fat content profile. Of all the fat mixtures, avocado fat:palm stearin:cocoa butter (84:7:9) displayed closer similarity to lard with respect to thermal transitions at –3.59°C and its solid fat content profile showed the least difference to that of lard throughout the temperature range measured.     

Enzymatic interesterification of anhydrous butterfat with flaxseed oil and palm stearin to produce low-trans spreadable fat

Lipase-catalysed interesterification was performed to produce low-trans spreadable fat (LTSF) with anhydrous butterfat (ABF), flaxseed oil (FSO), and palm stearin (PS) using biocatalysts (Lipozyme RM IM and Novozym SP435). The reaction was carried out for 24 h at 60 °C in a shaking water bath (180 rpm). Seven substrate blends for the production of LTSF were prepared: 12:6:2, 10:6:4, 9:6:6, 8:6:6, 6:6:8, 6:6:9, and 4:6:10 (ABF/PS/FSO, by weight). After 24-h interesterification, decreased saturated fatty acid (SFA) contents in LTSFs were observed with ranges from 67% to 41%. LTSFs exhibited low atherogenic index (AI) with ranges from 2.8 to 0.8, compared with 3.3–4.7 AI of butter fat. Equivalent carbon number (ECN) 42–46 in the newly produced triacylglycerol (TAG) increased according to rearrangement of FAs on TAG backbone. The α-linolenic acid contents (mol%) of each LTSF were 5.6% (12:6:2), 10.7% (10:6:4), 15.1% (9:6:6), 15.8% (8:6:6), 20.8% (6:6:8), 22.4% (6:6:9) and 26% (4:6:10). The trans fatty acids in the LTSFs was not more than 2%. Melting point ranges of LTSF were from 37 (9:6:6) to 32 °C (4:6:10).     

不同产地大花红景天多糖的单糖组成分析

通过气相色谱-质谱联用技术对不同产地大花红景天水提和碱提粗多糖的单糖组成及其比例进行分析。结果表明:西藏3个产地大花红景天水提粗多糖ST-Ⅰ、ST-Ⅱ、ST-Ⅲ均由鼠李糖、阿拉伯糖、甘露糖、葡萄糖和半乳糖构成,摩尔比分别为:1∶2.86∶0.43∶2.12∶0.63、1∶2.81∶0.28∶1.72∶0.37、1∶3.37∶0.58∶2.36∶0.66;碱提粗多糖JT-Ⅰ、JT-Ⅱ由甘露糖、葡萄糖和半乳糖构成,摩尔比分别为:1∶0.93∶2.90、1∶2.01∶2.92,JT-Ⅲ与其他两地相比,单糖组成有较大差异,由鼠李糖、岩藻糖、阿拉伯糖、甘露糖、葡萄糖和半乳糖,其摩尔比为5.60∶1.02∶13.20∶1∶9.27∶0.92。     

Physicochemical and volatiles characterization of trans-free solid fats produced by lipase-catalyzed interesterification

ABSTRACT:  Trans -free solid fats were synthesized from fully hydrogenated soybean oil (FHSBO), olive oil (OO), and palm stearin (PS) at different substrate weight ratios (10:20:70, 10:40:50 and 10:50:40) via lipase-catalyzed interesterification. The interesterified products contained mostly TAG (98.8% to 99.0%), and small amounts of MAG and DAG as by-products. The major fatty acids were oleic acid, palmitic acid, and stearic acid in the interesterified products, and the melting points ranged from 39 to 45 °C. The amount of α-tocopherol was reduced by 75% to 92%. Volatile analysis by solid-phase microextraction indicated that OO and PS had distinct volatile profiles, in which 18 volatiles were retained in interesterified products. Furthermore, some volatiles disappeared or formed during processing. Electronic nose showed that the odors of substrates (OO and PS) were different from each other, and the odors of interesterified products were distinguishable from that of OO or PS. Among the interesterified products, the odor of blend FHSBO:OO:PS of 10:40:50 or 10:50:40 was different from that of blend FHSBO:OO:PS (10:20:70). However, no odor difference was observed between products blend FHSBO:OO:PS 10:40:50 and 10:50:40.     

Cocoa butter equivalents prepared by blending fractionated palm stearin and shea stearin

The aim of this study was to assess the possibility of commercial use as a cocoa butter equivalent (CBE). The CBE was prepared by blending fractionated palm stearin and shea stearin in a weight ratio of 40:60 and contained 81.9% total symmetric monounsaturated triacyl-glycerols. The CBE was blended with cocoa butter in weight ratios (CBE:cocoa butter) of 5:95, 10:90, 20:80, 30:70, 40:60, 50:50, 60:40, 70:30, 80:20, and 90:10. The blends were evaluated for their fatty acid and triacylglycerol compositions, thermal melting/crystallization behaviors, and solid fat content. The 5:95, 10:90, 20:80, and 30:70 blends showed similar melting/crystallization temperature ranges and enthalpies to those of cocoa butter. Furthermore, they showed similar changes in solid fat content to those of cocoa butter as a function of temperature. These results indicate that the CBE can be blended with cocoa butter at 30% for the manufacture of chocolate products without significantly altering their physical properties.     

O:8 serotype Yersinia enterocolitica strains in China

Serotypes O:3, O:8 and O:9 Yersinia enterocolitica strains carrying virulence determinants are common pathogens causing human infections. In many years of surveillance in China for Y. enterocolitica, no pathogenic O:8 strains have been found where the isolated O:8 serotypes lacked the major virulence genes and in contrast to O:3 and O:9 strains, none of the O:8 isolates were from humans. These O:8 isolates lack ail, ystA, yadA and virF genes but possess the ystB gene and all belong to Biotype 1A. These O:8 strains did not kill mice and could protect immunized mice against challenge with a pathogenic O:8 strain. Compared to the Chinese pathogenic O:3 and O:9 strains which have similar pulsed-field gel electrophoresis patterns, the 39 Chinese O:8 animal and food isolates were different from the pathogenic O:8 reference strains. This suggests the O:8 strains lacking virulence determinants may not disseminate rapidly in humans and are maintained in animal reservoirs; and therefore exhibit higher variance and divergence from the virulent type.     

Chitosan–tripolyphosphate submicron particles as the carrier of entrapped rutin

Chitosan (CH)–tripolyphosphate (TPP) submicron particles were formed as carriers of entrapped rutin, and the release properties characterized using simulated gastric juices and fluids of the small intestine. Particle size, charge and entrapment efficiencies were investigated as a function of the CH:TPP molar ratio (2.0:1.0–5.0:1.0). Size was found to decrease from ∼814 nm for the 2.0:1:0 mass ratio to ∼528 nm for the ratios between 2.5:1.0 and 4.0:1.0, and then again to ∼322 nm for the 5:0:1.0 mass ratio, whereas all particles carried a positive surface charge, increasing from +21 to +59 mV as the ratio increased from 2.0:1.0 to 5.0:1.0. The percent entrapment was found to rise from 3.68% to 57.6% as the ratios increased from 2.0:1:0 to 4.0:1:0, before reaching a plateau. Submicron particles (4.0:1.0 mass ratio only) were found to retain rutin in simulated gastric fluids, whereas in conditions which simulated fluids from the small intestine, only 20% of the entrapped rutin was released and 80% remained absorbed to the CH:TPP carriers. Such particles have applications for the delivery of phenolics in food and natural health products.     

Effects of long chain fatty acids on lipid and glucose metabolism in monolayer cultures of bovine hepatocytes

The objectives were to determine the long-term (48 h) effects of specific long chain fatty acids on hepatic lipid and glucose metabolism in monolayer cultures of bovine hepatocytes. From 16 to 64 h after plating, hepatocytes from three 7- to 10-d-old calves were exposed to one of the following treatments: 1 mM palmitic acid (1 mM C16:0), 2 mM palmitic acid (2 mM C16:0), or 1 mM palmitic acid plus 1 mM of either stearic (C18:0), oleic (C18:1), linoleic (C18:2), linolenic (C18:3), eicosapentaenoic (C20:5), or docosahexaenoic (C22:6) acid, or 0.5 mM each of eicosapentaenoic and docosahexaenoic acid (C20:5 + C22:6). The two treatments containing 2 mM of saturated fatty acids, 2 mM C16:0 and 1 mM C16:0 plus 1 mM C18:0, increased beta-hydroxybutyrate concentrations in the medium and [1-(14)C]palmitic acid oxidation to acid-soluble products compared with all other treatments. The treatment containing C22:6 increased total cellular triglyceride content and incorporation of [1-(14)C]palmitic acid into cellular triglycerides. The treatments containing C22:6 or C20:5 + C22:6 increased [1-(14)C]palmitic acid metabolism to phospholipids and cholesterol. The presence of C22:6 in the medium decreased metabolism of [2-(14)C]propionic acid either to glucose in the medium or to cellular glycogen. Overall, fatty acids differed in their effects on lipid and glucose metabolism in monolayer cultures of bovine hepatocytes with C22:6 eliciting the most profound changes.     

Characterization of edible coatings consisting of pea starch, whey protein isolate, and Carnauba wax and their effects on oil rancidity and sensory properties of walnuts and pine nuts

Edible coatings made of whey protein isolate (WPI), pea starch (PS), and their combinations with carnauba wax (CW) were prepared and characterized. WPI combined with CW formed stable emulsion while PS with CW formed unstable emulsion and both formulations produced non-homogeneous films. Addition of PS to WPI: CW combination at the ratio of 1:1:1, respectively, resulted in stable emulsion and homogenous films. The emulsion PS: WPI: CW (1:1:2) was stable and formed a continuous film but had less homogenous droplets size distribution when compared to 1:1:1 film. Combined films had a reduced tensile strength and elongation compared to single component films. WPI : CW (1:1) films had higher elastic modulus than the WPI films, but the modulus reduced by the addition of PS. All the coating formulations were effective in preventing oxidative and hydrolytic rancidity of walnuts and pine nuts stored at 25 °C throughout the storage (12 d) but were less effective at 50 °C. Increasing the concentration of CW from 1:1:1 to 1:1:2 in PS: WPI: CW formulation did not contribute in further prevention of oil rancidity at 25 °C. Using of PS: WPI: CW (1:1:1) coating on both nuts significantly (P < 0.05) improved their smoothness and taste but the PS: WPI: CW (1:1:2) coatings imparted unacceptable yellowish color on walnuts. PRACTICAL APPLICATION: Edible coating of walnuts and pine nuts by whey protein isolate, pea starch, and carnauba wax reduced the oxidative and hydrolytic rancidity of the nuts and improved sensory characteristics.     

Preparation of Recombined Milk Using Modified Butterfats Containing α‐Linolenic Acid

Abstract: Modified butterfats (MBFs) were produced by lipase‐catalyzed interesterification with 2 substrate blends (6:6:8 and 4:6:10, by weight) of anhydrous butterfat (ABF), palm stearin, and flaxseed oil in a stirred‐batch type reactor after short path distillation. The 6:6:8 and 4:6:10 MBF contained 21.7% and 26.5%α‐linolenic acid, respectively. Total saturated fatty acids of the MBFs ranged from 41.4% to 47.4%. The cholesterol contents of the 6:6:8 and 4:6:10 MBFs were 21.0 and 12.1 mg/100 g, respectively. In addition, the melting points of the 6:6:8 and 4:6:10 MBFs were 32 °C and 31 °C, respectively. After preparation of recombined milks (oil‐in‐water emulsions) with MBFs, the stability of emulsions prepared with the MBFs (6:6:8 and 4:6:10) was compared to those with ABF during 10‐d storage at 30 °C. Skim milk powder (containing 1% protein) was added to prepare emulsions as an emulsifier. Microstructures of emulsions freshly prepared with the ABF and the MBFs consisted of uniform fat globules with no flocculation during 10‐d storage. With respect to fat globule size distribution, the volume‐surface mean droplet diameter (d₃₂) of the 6:6:8 and 4:6:10 MBF emulsions ranged between 0.33 and 0.34 μm, which was similar to the distribution in ABF emulsion. Practical Application: Milk, an expensive dairy food, has been widely used in various milk‐derived food products. Modified butterfats (MBFs) contain α‐linolenic acid as an essential fatty acid. Emulsion stability of recombined milks (oil‐in‐water emulsions) with MBFs was similar to that in anhydrous butterfat emulsion during 10‐d storage. They may be a promising alternative for reconstituted milks to use in processed milk‐based products.     

Development, Utilization and Protein Quality of Potato:Soy:Egg Flakes

Various ratios of potato:full-fat soy:whole egg (P:S:E) were combined and drum dried to yield flakes which were rehydrated and formed into baked or fried patties. Methods were developed to utilize either fresh or dehydrated potatoes for flake production. Sensory evaluations showed 60:34:6 P:S:E fried patties to be more generally accepted than baked patties. Freezing of the uncooked dough prior to frying or baking was shown to be a good storage method. P:S:E flakes (60:34:6) were also substituted for 12% of patent wheat flour in bread. P:S:E supplementation improved the protein efficiency ratio (PER) of patent flour wheat bread. PER's for 60:34:6 P:S:E flakes and baked patties were equivalent to casein.     

Effects of fat source and copper on unsaturation of blood and milk triacylglycerol fatty acids in Holstein and Jersey cows

Fatty acid composition of plasma triacylglycerides and milk fat was analyzed from Holstein and Jersey cows with control or depleted copper status and fed roasted whole soybeans or tallow. Conjugated linoleic acid in plasma was higher in Jersey cows. Dietary fat source influenced the proportions of all fatty acids in plasma and in milk, except for conjugated linoleic acid in milk. Feeding soybeans increased plasma C14:1, C18:0, C18:2, and conjugated linoleic acid, and decreased C14:0, C16:0, C16:1, and cis- and trans-C18:1 compared with feeding tallow. Low copper diets decreased C18:0 and increased cis- and trans-C18:1, and conjugated linoleic acid in plasma. A fat source x copper status interaction occurred for cis-C18:1 in plasma. Proportions of C4:0 to C14:0 were higher, and cis16:1, cis- and trans-C18:1, and conjugated linoleic acid were lower in milk fat of Jersey compared with Holstein cows. Generally, the effects of copper depletion were less apparent in milk than in plasma. Copper depletion increased C4:0, trans-C18:1, and conjugated linoleic acid, and decreased C16:1 in milk. Feeding whole soybeans increased C4:0 to C14:0, C18:0, C18:2, and C18:3, and decreased C14:1, C16:0, C16:1, and cis- and trans-C18:1 in milk. Breed x fat interactions occurred for C4:0, C14:1, C16:1, and conjugated linoleic acid in milk. Copper status x fat source interaction occurred for trans-C18:1. The breed x copper status interaction was apparent in milk fat for C16:1 and C18:0 and conjugated linoleic acid in milk. Both C18:0 and trans-C18:1 were desaturated by mammary tissue; however, whereas desaturation of C18:0 was linear, desaturation of trans-C18:1 reached a plateau that could have been caused by presence of the trans-10 isomer, which is not desaturated and was not separated from trans-11 C18:1 in our analysis. Comparison of the plasma triacylglycerol fatty acid profile with the milk fat profile was useful to interpret separate events of biohydrogenation in the rumen and desaturation by the mammary gland.     

Frozen Dessert Attribute Changes with Increased Amounts of Unsaturated Fatty Acids

Vanilla flavor frozen desserts were made to be similar in composition except for the fat. Canola and soybean oils were combined with milk fat to provide fatty acid ratios of 1:1:1 and 3:2:1 (saturated: monounsaturated:polyunsaturated), whereas the milk fat control had a ratio of 24:8.6: 1. Sensory, physical and chemical tests indicated minimal differences between the control ice cream and the dessert with the 3:2:1 ratio. However, product with the 1:1:1 ratio was described as oily and oxidized in flavor. It melted faster, was softer in the penetration test and coarser in texture than the ice cream control.     

混合办公废纸高效脱墨剂的研发

针对混合办公废纸油墨的特点,采用复配的方法研制开发的高效脱墨剂SWI,脱墨效果优于同类进口产品,而成本则比进口脱墨剂低73.3%.复配配方为:油酸:硬脂酸:十八醇:oP-1 0:Tween-60:十二烷基苯磺酸=15:5:2:4:3:2.配合常规脱墨药品,该脱墨剂相对于绝干废纸的最佳用量为0.3%左右.     

A Review of Criteria and Methods for Evaluating the Probiotic Potential of Microorganisms

Probiotics are live organisms that, when consumed in sufficient quantities, confer health benefits to the host. There is global interest in probiotics research; many are isolated from diverse niches and identified in numerous ways. The challenge is selecting the precise range of tests and methods to use. Some tests are argued to be outdated, biased and/or irrelevant. Screening guidelines exist, but at times are not strictly followed. In many countries, firm regulations regarding probiotics declaration and use are nonexistent. Consequently, some organisms are presented as probiotics without strong scientific evidence. This review discusses common screening approaches, their relevance, strengths and weaknesses.

Abbreviations: ADP: Adenosine Diphosphate; ATCC: American Type Culture Collection; ATP: Adenosine Triphosphate; BSH: Bile Salt Hydrolase; Caco-2: Cancer coli-2; cFDA-SE: Carboxyfluorescein diacetate succinimidyl ester; CLSI: Clinical and Laboratory Standards Institute; DNA: Deoxyribonucleic acid; EPN: Epinephrine; FAO: Food and Agriculture Organization; FbPA: Fibronectin binding protein A; FDA: Food and Drug Authority; GIT: Gastrointestinal tract; GRAS: Generally Recognized as Safe; HEPES : 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; HT-29: Human colon adenocarcinoma cells; HT-29 MTX: Methotrexate treated human colon adenocarcinoma cells; ICMR-DBT: Indian Council of Medical Research and Department of Biotechnology; IBS: Irritable Bowel Syndrome; ICR: Inherited Cataract Rat; LAB: Lactic Acid Bacteria; MFI: Mean Fluorescence Intensity; MIC: Minimum Inhibitory Concentration; mRNA: Messenger Ribonucleic Acid; MRS: de Man Rogosa Sharpe; NCCB: Netherlands Culture Collection of Bacteria; OD: Optical Density; PBS: Phosphate Buffered Saline; PGE: Prostaglandin; RBC: Red Blood Cells; SHIME: Simulator of the Human Intestinal Microbial Ecosystem; SDS-PAGE: Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis; TLC: Thin-Layer Chromatography; TRAP-6: Thrombin Receptor Activator Peptide-6; WHO: World Health Organization     

The Compound Characteristics Comparison (CCC) approach: a tool for improving confidence in natural compound identification

ABSTRACT

Compound identification is the main hurdle in LC-HRMS-based metabolomics, given the high number of ‘unknown’ metabolites. In recent years, numerous in silico fragmentation simulators have been developed to simplify and improve mass spectral interpretation and compound annotation. Nevertheless, expert mass spectrometry users and chemists are still needed to select the correct entry from the numerous candidates proposed by automatic tools, especially in the plant kingdom due to the huge structural diversity of natural compounds occurring in plants. In this work, we propose the use of a supervised machine learning approach to predict molecular substructures from isotopic patterns, training the model on a large database of grape metabolites. This approach, called ‘Compounds Characteristics Comparison’ (CCC) emulates the experience of a plant chemist who ‘gains experience’ from a (proof-of-principle) dataset of grape compounds. The results show that the CCC approach is able to predict with good accuracy most of the sub-structures proposed. In addition, after querying MS/MS spectra in Metfrag 2.2 and applying CCC predictions as scoring terms with real data, the CCC approach helped to give a better ranking to the correct candidates, improving users’ confidence in candidate selection. Our results demonstrated that the proposed approach can complement current identification strategies based on fragmentation simulators and formula calculators, assisting compound identification. The CCC algorithm is freely available as R package (https://github.com/lucanard/CCC) which includes a seamless integration with Metfrag. The CCC package also permits uploading additional training data, which can be used to extend the proposed approach to other systems biological matrices.

List of abbreviations: Acidic: acidic moiety; aliph: aliphatic chain; AUC: area under the ROC curve; bs: best glycosidic structure; CCC: Compounds’ Characteristics Comparison; Cees: Carbons estimation errors; CO: Carbon to Oxygen ratio; Het: Heterocyclic moiety; IMD: Isotopic Mass Defect (and Pattern); LC-HRMS: Liquid Chromatography – High Resolution Mass Spectrometry; md: mass defect; MM: Monoisotopic Mass; MS: Mass Spectrometry; MSE: Mean Squared Error; nC: number of Carbons; NN: Nitrogen; pC: percentage of Carbon mass on the total mass; Pho: Phosphate; PLSr: Partial Least Square regression; ppm: parts per million; QSRR: Quantitative structure-retention relationship; RMD: Relative Mass Defect; ROC: Receiver Operating Characteristics; rRMD: residual Relative Mass Defect; RT: retention time; Sul: Sulphur; UPLC-ESI-Q-TOF-MS: Ultra Performance Liquid Chromatography – ElectroSpray Ionization –Quadropole – Time of Flight – Mass Spectrometry; VAT: Vitis arizonica Texas