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Short-chain fatty acids (SCFA) and lactate in ruminal fluid are products resulting from the microbial fermentation of substrates and can be used to reflect the composition and activity of the ruminal microbiome. Determination of SCFA and D-/L-lactate in ruminal fluid currently requires two separate protocols, which is time-consuming and costly. In this study, we have optimised and validated a simple and unified 3-nitrophenylhydrazine (3-NPH) derivatisation protocol and a 20 min chiral-LC-MS method for the simultaneous quantification of all SCFA and D- and L-lactate in ruminal fluid. This method, which requires no sample pretreatment or purification shows adequate sensitivity (limit of detection (LOD): 0.01 µg/mL), satisfactory accuracy (recovery: 88–103%), and excellent reproducibility (relative standard deviation (RSD) for repeated analyses < 3% for most analytes). The application of this method to a cohort of 24 animals allowed us to reveal a large inter-cow variation in ruminal SCFA and lactate level, the concentration range for each species, the widespread correlation between different SCFA, and the strong correlation between D- and L-lactate. [ABSTRACT FROM AUTHOR]

Simple Summary: Batocera horsfieldi (Hope) is an important wood-boring pest in China. This pest primarily infests tree trunks by feeding on the woody tissue, creating a network of interconnected tunnels within the trunk. These tunnels become blocked with insect feces and wood debris, causing damage, decay, and even the death of the host plant's tissues. In this study, single-molecule real-time sequencing (SMRT) and Illumina RNA-seq technologies were employed to conduct full-length transcriptome sequencing of male and female adults of B. horsfieldi. A total of 20,356,793 subreads (38.26 G, clean reads) were generated, including 432,091 circular consensus sequences and 395,851 full-length non-chimera reads. Clustering and redundancy removal of the full-length non-chimera reads resulted in 39,912 consensus reads. Additionally, functional annotation was performed on a total of 84,650 transcripts in seven different databases. This study provides an important foundation for future exploration of gene regulation in the interaction between B. horsfieldi and host plants using RNA interference (RNAi), and it offers a scientific basis for the prevention and control of B. horsfieldi. Batocera horsfieldi (Hope) (Coleoptera: Cerambycidae) is an important forest pest in China that mainly infests timber and economic forests. This pest primarily causes plant tissue to necrotize, rot, and eventually die by feeding on the woody parts of tree trunks. To gain a deeper understanding of the genetic mechanism of B. horsfieldi, this study employed single-molecule real-time sequencing (SMRT) and Illumina RNA-seq technologies to conduct full-length transcriptome sequencing of the insect. Total RNA extracted from male and female adults was mixed and subjected to SMRT sequencing, generating a complete transcriptome. Transcriptome analysis, prediction of long non-coding RNA (lncRNA), coding sequences (CDs), analysis of simple sequence repeats (SSR), prediction of transcription factors, and functional annotation of transcripts were performed in this study. The collective 20,356,793 subreads (38.26 G, clean reads) were generated, including 432,091 circular consensus sequences and 395,851 full-length non-chimera reads. The full-length non-chimera reads (FLNC) were clustered and redundancies were removed, resulting in 39,912 consensus reads. SSR and ANGEL software v3.0 were used for predicting SSR and CDs. In addition, four tools were used for annotating 6058 lncRNAs, identifying 636 transcription factors. Furthermore, a total of 84,650 transcripts were functionally annotated in seven different databases. This is the first time that the full-length transcriptome of B. horsfieldi has been obtained using SMRT sequencing. This provides an important foundation for investigating the gene regulation underlying the interaction between B. horsfieldi and its host plants through gene editing in the future and provides a scientific basis for the prevention and control of B. horsfieldi. [ABSTRACT FROM AUTHOR]

Milk is a rich source of lipids, with the major components being triglycerides (TAG) and phospholipids (mainly phosphatidylcholine (PC), sphingomyelin (SM), phosphatidylethanolamine (PE), phosphatidylserine (PS) and phosphatidylinositol (PI)). Liquid chromatography-mass spectrometry (LC-MS) is the predominant technique for lipid identification and quantification across all biological samples. While fatty acid (FA) composition of the major lipid classes of milk can be readily determined using tandem MS, elucidating the regio-distribution and double bond position of the FA remains difficult. Various workflows have been reported on the quantification of lipid species in biological samples in the past 20 years, but no standard or consensus methods are currently available for the quantification of milk phospholipids. This study will examine the influence of several common factors in lipid analysis workflow (including lipid extraction protocols, LC stationary phases, mobile phase buffers, gradient elution programmes, mass analyser resolution and isotope correction) on the quantification outcome of bovine milk phospholipids. The pros and cons of the current LC-MS methods as well as the critical problems to be solved will also be discussed. [ABSTRACT FROM AUTHOR]

An accurate dry matter intake (DMI) measurement method is of great importance for determining the performance of dairy cows. While n-alkane technique is commonly used to estimate the DMI of ruminants, the protocols used to measure alkanes are complex, time consuming and low throughput. Herein we present a simplified and miniaturised protocol that combines a smaller sample matrix (60 mg of faeces or herbage), a shorter (3 h) saponification step and a direct purification of the crude extract by filtration through silica powder or mixing with silica powder prior to GC-FID analysis. In addition to a much higher throughput compared with the existing methods, by skipping the commercial SPE columns and reducing the organic solvent volume, our miniaturised protocol should significantly reduce the sample processing cost and is thus suitable for alkane determination for large-scale experiments. [ABSTRACT FROM AUTHOR]

Short-chain fatty acids (SCFA, C2-C5) in milk and serum are derived from rumen bacterial fermentation and, thus, have the potential to be used as biomarkers for the health status of dairy cows. Currently, there is no comprehensive and validated method that can be used to analyse all SCFAs in both bovine serum and milk. This paper reports an optimised protocol, combining 3-nitrophenylhydrazine (3-NPH) derivatisation and liquid chromatography-mass spectrometry (LC-MS) analysis for quantification of SCFA and β-hydroxybutyric acid (BHBA) in both bovine milk and bovine serum. This method is sensitive (limit of detection (LOD) ≤ 0.1 µmol/L of bovine milk and serum), accurate (recovery 84–115% for most analytes) and reproducible (relative standard deviation (RSD) for repeated analyses below 7% for most measurements) with a short sample preparation step. The application of this method to samples collected from a small cohort of animals allowed us to reveal a large variation in SCFA concentration between serum and milk and across different animals as well as the strong correlation of some SCFAs between milk and serum samples. [ABSTRACT FROM AUTHOR]

Regioisomers (or positional isomers) of triacylglycerols (TAGs) of milk are known to show differential outcome in relation to human absorption. Quantitation of TAG regioisomers remains a big challenge due to the lack of facile chromatographic separation technique. The feasibility of using fragment ion intensity ratio to determine the ratio of co-eluting AAB/ABA-type regioisomer pairs was confirmed in this study. The ability of C30 stationary phase in resolving interfering TAG isomers was demonstrated for the first time. This allowed us to reveal the complexity of using fragment ion intensity to quantify 1,2-olein-3-palmitin (OOP), 1,3-olein-2-palmitin (OPO), 1,2-olein-3-stearin (OOS), and 1,3-olein-2-stearin (OSO) regioisomers in milk samples. A novel algorithm was proposed to consider the contribution of OPO/OOP and OSO/OOS double bond (DB)-isomers and to eliminate the interference of isobaric ions from other isomers, an aspect overlooked in previous studies. This liquid chromatography-mass spectrometry method that requires no pre-fractioning and a moderate chromatographic separation time of 36 min is simple and, thus, suitable for screening a large number of samples for genetic analysis of this trait. Preliminary results using a small cohort of animals showed that OPO/OOP ratio differs significantly between Jersey and Holstein cows, and a large variation was also observed across individual Holstein cows. [ABSTRACT FROM AUTHOR]

Volatile phenols (VPs) derived from smoke-exposed grapes are known to confer a smoky flavor to wine. Current methods for determination of VPs in grape berries either involve complex sample purification/derivatization steps or employ two analytical platforms for free and bound VP fractions. We report here a simple gas chromatography-tandem mass spectrometry (GC-MS/MS) method for quantification of both free and bound VPs in grapes, based on optimized (1) GC-MS/MS parameters, (2) an analyte extraction procedure, and (3) phenol glycoside hydrolysis conditions. Requiring neither sample cleanup nor a derivatization step, this method is sensitive (LOD ≤ 1 ng/g berries) and reproducible (RSD < 12% for repeated analyses) and is expected to significantly reduce the sample turnover time for smoke taint detection in vineyards. [ABSTRACT FROM AUTHOR]

Ensuring the large signal stability of the DC microgrid is the premise of the safe operation of the DC microgrid, but the research on the large-signal stability of microgrids with multiple droop control micro-sources is still scarce. In this paper, a DC microgrid system model with multiple droop control micro-sources was established by appropriate simplification. Addressing the problem that most stability research methods cannot be quantitatively analyzed, the mixed potential function method was used to analyze the large signal stability of the system. However, the criterion obtained by the conventional mixed potential function method is complicated and contains multiple time-varying parameters, which is not convenient for analysis. Therefore, the simple form of the criterion was obtained through simplification and the analysis proved the rationality of the simplification. On this basis, a nonlinear droop control method was proposed to improve the anti-interference ability of the system. Finally, the accuracy of the large signal stability criterion and the effectiveness of nonlinear droop control on the system's large signal stability were verified by simulation. [ABSTRACT FROM AUTHOR]

Milk contains 3-6% of fat, of which the dominant component is triacylglycerol (TAG). Over 100 TAG groups can be readily detected in any non-enriched milk sample by LC-MS; most TAG groups contain several isomers (TAG molecules with different fatty acid composition), which cannot be fully resolved chromatographically by any single stationary phase. TAG profile of mature milk from 19 cows was surveyed in this study for eight consecutive months using RP-LC-Orbitrap MS. It was found that TAG profile of milk was not constant throughout the milking season and the seasonal pattern varied with TAG groups. The overall unsaturation level of TAG was stable from October 2013 to January 2014, decreased in February/March 2014 and then increased from April and peaked in May 2014. In addition to the seasonal fluctuation in TAG profile, the proportion of different isomeric species within a TAG group also changed substantially across seasons. However, the proportion of different positional isomers within a given TAG group does not seem to vary during the milking season. To our knowledge, this is the first report on the seasonal change of milk lipid at the TAG group and isomer level., Competing Interests: The authors declare no conflict of interest.