Monitoring Changes in Biochemical and Metabolite Profiles in Garlic Cloves during Storage.

Storage is important for the garlic cloves industry because it is critical to enabling a year-round supply. This study aimed to investigate the changes in biochemical and metabolic profiles in garlic cloves in terms of different temperatures and cultivars during storage using nontargeted and targeted metabolomics. The results showed that the storage temperatures and times were important factors affecting the composition and metabolite content of garlic cloves. In detail, the metabolic profiling of garlic cloves changed significantly at 22 °C, which was mainly related to sprouting. Furthermore, γ-glutamyl peptide was converted into the corresponding flavor precursors or free amino acids, leading to the fluctuation in the amount of nutrients in garlic cloves. In contrast, the quality of garlic cloves remained stable for 290 days at 0 °C though metabolism still occurred, which indicated that the slight chemical changes did not impact the quality significantly and low temperature could prolong their dormancy.

The quality evaluation of 30 Asparagus officinalis L. varieties.

Asparagus, a vital economic contributor, is a well-liked vegetable grown around the globe, and some secondary metabolites in its spear are beneficial to human health. Asparagus spears possess a significant quantity of nutrients and phytochemicals; however, the difference in these chemical compositions among various varieties has not been sufficiently studied. This work aimed to detect the chemical compositions of 30 varieties of asparagus and to assess them by principal component analysis (PCA). The results showed that the contents of these chemical compositions varied in varieties. Selenium (Se, 1.12-2.9 µg/100 g dry-weight [DW]) was abundant in asparagus, with an average dry matter content of 8.25%. Free amino acids (5.60-9.98 g/100 g DW) and polyphenols (6.34-8.67 mg/g DW) were both present in high amounts, along with flavonoids (4.218-8.22 mg/g DW) and protodioscin (0.44-1.96 mg/g DW). Correlation analysis, PCA, and hierarchical cluster analysis were used to conduct a comprehensive evaluation of asparagus. Atlas, Appolo, Jinggang 111, Jingke 2, and WS-1 were the top five varieties with comprehensive scores. This study provided valuable data for the breeding, quality improvement, processing, and utilization of asparagus varieties in the future.

Silencing gram-negative bacteria binding protein 1 decreases the immunity of Tribolium castaneum against bacteria.

Gram-negative bacteria binding proteins (GNBPs) have the ability to recognize molecular patterns associated with microbial pathogens (PAMPs), leading to the activation of immune responses downstream. In the genome of Tribolium castaneum, three GNBP genes have been identified; however, their immunological roles remain unexplored. In our study, a GNBP1, designated as TcGNBP1, were identified from the cDNA library of T. castaneum. The coding sequence of TcGNBP1 consisted of 1137 bps and resulted in the synthesis of a protein comprising 378 amino acids. This protein encompasses a signal peptide, a low-complexity region, and a glycoside hydrolase 16 domain. TcGNBP1 was strongly expressed in early adult stages, and mainly distributed in hemolymph and gut. Upon being challenged with Escherichia coli or Staphylococcus aureus, the transcript levels of TcGNBP1 were significantly changed at different time points. Through molecular docking and ELISA analysis, it was observed that TcGNBP1 has the ability to interact with lipopolysaccharides, peptidoglycan, and ß-1, 3-glucan. Based on these findings, it was further discovered that recombinant TcGNBP1 can directly bind to five different bacteria in a Ca2+-dependent manner. After knockdown of TcGNBP1 with RNA interference, expression of antimicrobial peptide genes and prophenoloxidase (proPO) activity were suppressed, the susceptibility of T. castaneum to E. coli or S. aureus infection was enhanced, leading to low survival rate. These results suggest a regulatory mechanism of TcGNBP1 in innate immunity of T. castaneum and provide a potential molecular target for dsRNA-based insect pest management.

Effects of Spinach Extract and Licorice Extract on Growth Performance, Antioxidant Capacity, and Gut Microbiota in Weaned Piglets.

Anemia and weaning stress are important factors affecting piglet growth performance. Spinach extract and licorice extract have been used to improve anemia and antioxidant capacity, respectively. However, whether they have synergistic effects has not been reported. To evaluate the effects of mixed spinach extract and licorice extract on growth performance, serum biochemistry, antioxidant capacity, and gut microbiota in weaned piglets, a total of 160 weaned piglets were randomly allotted to four treatments with four replications of 10 piglets each. The four treatments were as follows: control (CON) group (basal diet), spinach extract (SE) group (basal diet + 1.5 kg/t spinach extract), licorice extract (LE) group (basal diet + 400 g/t licorice extract), and spinach extract and licorice extract (MIX) group (basal diet + 1.5 kg/t spinach extract + 400 g/t licorice extract). The results showed that, compared with the CON group, diets supplemented with spinach extract and licorice extract significantly increased the average daily gain (p < 0.05), while considerably reducing the feed-to-gain ratio (p < 0.05). Moreover, the MIX group exhibited a significant up-regulation of serum total protein, globulin, albumin, glucose, and triglyceride levels in comparison to the CON group (p < 0.05). Meanwhile, both the anemia and antioxidant capacity of piglets were effectively improved. Notably, the MIX group achieved even better results than the individual supplementation in terms of enhancing growth performance, which could potentially be attributed to the increased abundance of the Rikenellaceae_RC9_gut_group. These results demonstrated that the supplementation of diets with spinach extract and licorice extract improves the absorption of nutrients from the diet and antioxidant capacity in weaned piglets.

Development of an analytic method for organosulfur compounds in Welsh onion and its use for nutritional quality analysis of five typical varieties in China.

A reliable, simple, and sensitive method capable of quantifying six organosulfur compounds (OSCs) was established. The samples were extracted by water containing 3 % formic acid with a simple vortex, ultrasound, and centrifugation step, and the solutions were analyzed by ultra-high-performance liquid chromatography separation system coupled with a triple-quadrupole mass spectrometry (UHPLC - MS/MS). Then the method was applied for the analysis of six OSCs in five varieties of two types Welsh onions in China, and the moisture content, reducing sugar, total polyphenols, and 21 free amino acids were also analyzed to study the characters of these Welsh onions intensively. Multivariate statistical analysis was used to investigate the differences in OSCs and free amino acids profiles among the samples. This study showed that enzymatic inhibition method combined with UHPLC - MS/MS is an effective technique to analyze OSCs in Welsh onion, and could be valuable for the routine quantitation of OSCs in other foods.

Functional Analysis of a CTL-X-Type Lectin CTL16 in Development and Innate Immunity of Tribolium castaneum.

C-type lectins (CTLs) are a class of proteins containing carbohydrate recognition domains (CRDs), which are characteristic modules that recognize various glycoconjugates and function primarily in immunity. CTLs have been reported to affect growth and development and positively regulate innate immunity in Tribolium castaneum. However, the regulatory mechanisms of TcCTL16 proteins are still unclear. Here, spatiotemporal analyses displayed that TcCTL16 was highly expressed in late pupae and early adults. TcCTL16 RNA interference in early larvae shortened their body length and narrowed their body width, leading to the death of 98% of the larvae in the pupal stage. Further analysis found that the expression level of muscle-regulation-related genes, including cut, vestigial, erect wing, apterous, and spalt major, and muscle-composition-related genes, including Myosin heavy chain and Myosin light chain, were obviously down-regulated after TcCTL16 silencing in T. castaneum. In addition, the transcription of TcCTL16 was mainly distributed in the hemolymph. TcCTL16 was significantly upregulated after challenges with lipopolysaccharides, peptidoglycans, Escherichia coli, and Staphylococcus aureus. Recombinant CRDs of TcCTL16 bind directly to the tested bacteria (except Bacillus subtilis); they also induce extensive bacterial agglutination in the presence of Ca2+. On the contrary, after TcCTL16 silencing in the late larval stage, T. castaneum were able to develop normally. Moreover, the transcript levels of seven antimicrobial peptide genes (attacin2, defensins1, defensins2, coleoptericin1, coleoptericin2, cecropins2, and cecropins3) and one transcription factor gene (relish) were significantly increased under E. coli challenge and led to an increased survival rate of T. castaneum when infected with S. aureus or E. coli, suggesting that TcCTL16 deficiency could be compensated for by increasing AMP expression via the IMD pathways in T. castaneum. In conclusion, this study found that TcCTL16 could be involved in developmental regulation in early larvae and compensate for the loss of CTL function by regulating the expression of AMPs in late larvae, thus laying a solid foundation for further studies on T. castaneum CTLs.

2D Phthalocyanine-Assembled Porous Nanostructure-Based Electrochemical Platform for High-Efficiency Detection of Ascorbic Acid.

In this work, a novel two-dimensional (2D) porous nanostructure is constructed upon air/water interfacial assembly of 12-crown-ether-4-incorporated double-decker phthalocyanine (Pc2). The combination of the good electroconductivity of phthalocyanine and the great surface area of the porous structure endows the assembled film with excellent chemical sensing property for ascorbic acid (AA). The low limit of detection can be 0.15 µM with a large linear concentration range and strong anti-interfering ability, which can be comparable to the best results of tetrapyrrole-based electrochemical sensors for AA. Furthermore, the obtained 2D porous assembled film sensor can be applied in real-time monitoring of AA in commercial drinks, indicating its application potential in accurate detection of AA in real samples.

Distinct Quality Changes of Asparagus during Growth by Widely Targeted Metabolomics Analysis.

Asparagus is a popular vegetable and traditional medicine consumed worldwide due to its health benefits. The quality of asparagus, mainly attributed to small components like flavonoids and steroid, is quite differential as a result of different environments and maturities. However, the accumulation pattern and regulatory mechanism of metabolites in asparagus remain unclear so far. Herein, widely targeted metabolomics analysis was employed to study the quality and chemical composition variances of four asparagus, including three green asparagus of different maturities and one white asparagus. A total of 1045 metabolites were annotated in asparagus in which flavonoids and phenolic acids accounted for 37.51% of the total. Green asparagus was found to be rich in flavonoids, while white asparagus contained more steroids. Additionally, 461 biomarkers were screened between matured green and white asparagus, which is much more than that filtered among three green asparagus at different growth stages. These results indicated that sunlight has a stronger effect on the metabolism of asparagus compared to the general development of asparagus. Linoleic acid metabolism and alpha-linolenic acid metabolism were active during green asparagus growth, while flavone and flavonol biosynthesis and flavonoid biosynthesis resulted as two of the most important pathways when asparagus was exposed to sunlight.

Regulatory mechanisms of Beta-carotene and BCMO1 in adipose tissues: A gene enrichment-based bioinformatics analysis.

Beta-carotene (ß-carotene, BC) is one of the carotenoids most commonly consumed by humans. BCMO1 is expressed in various human tissues and is considered to be a key enzyme that converts BC into vitamin A. Studies indicated that BC-derived carotenoid signaling molecules affected the physiological functions of fat cells. In order to investigate the role and possible molecular mechanism of BC in mouse adipocytes, we conducted 4-group and 2-group difference analysis based on the data of GSE27271 chip in the Gene Expression Omnibus database. Genes differentially expressed in the inguinal white adipose tissue of mice were screened out and combined with the STRING database to construct protein-protein interaction (PPI) networks. Among them, Alb (albumin), Mug1 (murinoglobulin-1) and Uox (urate oxidase) genes were at relatively key positions and may affect the action of BC. Besides, Ppara (peroxisome proliferator-activated receptor alpha), Acly (ATP-citrate lyase) and Fabp5 (fatty acid-binding protein 5) genes constituted functional partners with many genes in the PPI network, and these genes may be Bcmo1 targeting molecules. Gene Ontology (GO) function and signaling pathways enrichment analysis were performed on the genes with protein interaction relationship in the PPI network. Fatty acid binding, cholesterol metabolic process, and regulation of fatty acid metabolic process were significantly enriched, and PPAR signaling pathway showed the most significant, indicating that BC and Bcmo1 might synergistically affect body metabolic functions such as fat metabolism. In general, BC and Bcmo1 may play a role in fat metabolism in mice, thereby affecting other functions or diseases.

Determination of Three Typical Metabolites of Pyrethroid Pesticides in Tea Using a Modified QuEChERS Sample Preparation by Ultra-High Performance Liquid Chromatography Tandem Mass Spectrometry.

Pyrethroid pesticides are widely used on tea plants, and their residues of high frequency and concentration have received great attention. Until recently, the residues of typical metabolites of pyrethroid pesticides in tea were unknown. Herein, a modified "quick, easy, cheap, effective, rugged and safe" (QuEChERS) method for the determination of three typical metabolites of pyrethroid pesticides in tea, using ultra performance liquid chromatography tandem mass spectrometry, was developed. The mixture of florisil, octadecylsilane, and graphite carbon black was employed as modified QuEChERS adsorbents. A Kinetex C18 column achieved good separation and chromatographic peaks of all analytes. The calibration curves of 3-phenoxybenzoic acid (3-PBA) and 4-fluoro-3-phenoxybenzoic acid (4-F-3-PBA) were linear in the range of 0.1-50 ng mL-1 (determination coefficient R2 higher than 0.999), and that of cis-3-(2-chloro-3,3,3-trifluoroprop-1-en-1-yl)-2,2-dimethylcyclopropanecarboxylic acid (TFA) was in the range of 1-100 ng mL-1 (R2 higher than 0.998). The method was validated and recoveries ranged from 83.0% to 117.3%. Intra- and inter-day precisions were lower than or equal to 13.2%. The limits of quantification of 3-PBA, 4-F-3-PBA, and TFA were 5, 2, and 10 µg kg-1, respectively. A total of 22 tea samples were monitored using this method, and 3-PBA and TFA were found in two green tea samples.

Metabolic Changes in Different Tissues of Garlic Plant during Growth.

The accumulation, distribution, and transportation of nutrients in different tissues of garlic during growth are unclear. Thereby, five tissues (leaf, pseudostem, bulb wrapper, clove skin, and clove) collected at 7 weeks were subjected to metabolomics analysis. A total of 84 biomarkers were identified during garlic plant growth. Most organosulfur compounds, amino acids, and dipeptides were upregulated in the clove, while a reversed trend was observed in other tissues. In addition, nucleotides and alkaloids increased because of senescence in the last 2 weeks except for the clove. The results also indicated that the garlic plant at an early stage is an ideal vegetable that is rich in nutrients. When the leaves began to wither, most nutrients were transported from other tissues to cloves, and the content of 7 total flavor precursors and 20 total amino acids in the clove increased by 113% and 65% after week 5, respectively. Therefore, delayed harvest may improve the nutritional quality of garlic bulbs.

Distinct Quality Changes of Garlic Bulb during Growth by Metabolomics Analysis.

Garlic-specialized metabolites contribute to both spicy flavor and healthy function of garlic. Their accumulation pattern and regulatory mechanism vary greatly at different environments and maturities. Herein, metabolomics models were built to evaluate and predict the quality and chemical composition variances of four garlic varieties in two regions at six growth stages. A total of 91 metabolites were identified, and their accumulation pattern during growth in three varieties of garlic in Shandong was similar but obviously distinct from that planted in Heilongjiang. Active metabolism for organosulfur compounds and amino acids was observed, and most metabolites with the "γ-glutamyl-" group were the storage compounds of nitrogen and sulfur in garlic because they increased remarkably during growth. The levels of functional components in garlic varied among different stages, and reliable prediction models for these compounds were provided, which may give a new idea for the estimation of garlic quality and confirmation of the best harvest time.

Profiling of organosulfur compounds and amino acids in garlic from different regions of China.

The quality of garlic is mainly attributed to organosulfur compounds that are the secondary metabolites of certain amino acids. Herein, a sensitive and rapid method for the simultaneous determination of 7 organosulfur compounds and 21 amino acids using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) was developed. Then the levels of these 28 compounds and allicin in garlic among 242 samples from 6 provinces in China were profiled with multivariate data analysis. The results showed that garlic was rich in organosulfur compounds and could be a good supplement for amino acids. Contents of 29 compounds varied greatly among the provinces. The multivariate data analysis demonstrated two major groups for all samples, indicating that the location has a significant influence on the metabolism of garlic. Besides, the compositional differences of garlic from 6 provinces and correlations for growing conditions and contents of components were also analyzed.

[Rapid simultaneous determination of 10 nutrients in ginger by ultra-high performance liquid chromatography-tandem mass spectrometry].

An analytical method by ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) was developed for the simultaneous determination of the two types of nutrients, viz. gingerol-related compounds and curcuminoids in ginger, including 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, 8-shogaol, 10-shogaol, tetrahydrocurcumin, curcumin, demethoxycurcumin and bisdemethoxycurcumin. The 10 target compounds were separated on a ZORBAX RRHD Eclipse Plus C18 column (100 mm×2.1 mm, 1.8 µm) using gradient elution with 0.1% (v/v) formic acid aqueous solution and methanol containing 0.1% (v/v) formic acid as the mobile phases. Qualitative confirmation and quantitative analysis of the target compounds were performed using an electrospray ionization (ESI) source in the positive ion and multiple reaction monitoring (MRM) modes. The linear correlation coefficients (r) of the 10 nutrients were ≥ 0.9995. The limits of quantification were 0.10-7.71 µg/L. The average spiked recoveries of the samples at the three levels were 82.8%-115.3%, the relative standard deviations (RSDs) were 0.58%-11.49%. The results showed that all the 10 nutrients in ginger were detected, and the content of 6-gingerol was the highest at 373.35-702.48 mg/kg. This method is convenient, rapid, accurate and reliable, and is suitable for the analysis of gingerol-related and curcuminoids in ginger. It provides a technical means for the quality identification and control of ginger.

Development and validation of an ultra performance liquid chromatography Q-Exactive Orbitrap mass spectrometry for the determination of fipronil and its metabolites in tea and chrysanthemum.

A fast, sensitive and reliable method for the determination of fipronil and its metabolites in tea and chrysanthemum was developed using a modified QuEChERS technique and an ultra performance liquid chromatography Q-Exactive Orbitrap mass spectrometry. The mixture of adsorbents containing primary secondary amine (PSA), octadecylsilane (C18) and carbon nanotubes (CNTs), was used as QuEChERS adsorbents. The use of mass resolution at 70000 full width at half maximum (FWHM) and narrow mass windows at 5 ppm achieved high selectivity and repeatability. Satisfactory linearity with correlative coefficient (R2) higher than 0.996 was achieved for all compounds. Recoveries at three levels (2, 10 and 50 µg kg-1) ranged from 86% to 112%, while the intra- and inter-day accuracies were less than 15%. Limits of quantification for fipronil and its metabolites were 2 µg kg-1, which fulfils the requirement of maximum residue limits formulated by European Union and Japan.

Residue pattern of polycyclic aromatic hydrocarbons during green tea manufacturing and their transfer rates during tea brewing.

Residues of polycyclic aromatic hydrocarbons (PAHs) in green tea and tea infusion were determined using gas chromatography-tandem mass spectrometry to study their dissipation pattern during green tea processing and infusion. Concentration and evaporation of PAHs during tea processing were the key factors affecting PAH residue content in product intermediates and in green tea. PAH residues in tea leaves increased by 2.4-3.1 times during the manufacture of green tea using the electric heating model. After correction to dry weight, PAH residue concentrations decreased by 33.5-48.4% during green tea processing because of PAH evaporation. Moreover, spreading and drying reduced PAH concentrations. The transfer rates of PAH residues from green tea to infusion varied from 4.6% to 7.2%, and PAH leaching was higher in the first infusion than in the second infusion. These results are useful for assessing exposure to PAHs from green tea and in formulating controls for the maximum residue level of PAHs in green tea.

Occurrence and Residue Pattern of Phthalate Esters in Fresh Tea Leaves and during Tea Manufacturing and Brewing.

The residues of 16 phthalate esters (PAEs) in fresh tea leaves and made tea were determined via gas chromatography-tandem mass spectrometry to study their distribution and degradation characteristics during tea planting and processing. Five PAEs were detected in all fresh tea leaves, and higher concentrations were detected in mature leaves. The distribution of PAEs in fresh tea leaves ranged from 69.7 to 2244.0 µg/kg. The degradative percentages of ∑5PAEs during green tea manufacturing ranged from 61 to 63% and were significantly influenced by the drying process. The transfer rates of PAEs-D4 ranged from 5.2 to 100.6%. PAEs with a high water solubility showed the highest transfer coefficient in the range of 91.8-100.6%, whereas PAEs with a high log Kow showed a low leaching efficiency below 11.9%. These results benefit the risk evaluation and establishment of a maximum residue limit for PAEs in tea.

[Effects of Cu2+ on growth hormone secretion of pig pituitary cells in culture].

AIM: To study the effects of Cu2+ on growth hormone (GH) secretion of pig pituitary cells in vivo. METHODS: Gland pituitary cells of 32-35 days old pigs were incubated for 48 h in presence of 10% fetal calf serum in DMEM. Cells were treated for 36 h with various concentrations of Cu2+ (0 mg/L, 0.025 mg/L, 0.1 mg/L, 0.4 mg/L, 1.6 mg/L) in serum-free DMEM. Culture medium was collected in 12 h, 24 h, 36 h thereafter for GH measurement and the radioimmunoassay kits were provided by Linco. RESULTS: The GH concentration was higher in treatments of 0.025 mg/L, 0.1 mg/L, 0.4 mg/L than that of 0 mg/L when the cells were treated with Cu2+ for 24 h, and it was significantly different (P < 0.05) between treatment 0.1 mg/L and treatment 0 mg/L. CONCLUSION: It was concluded that the presence of Cu2+ stimulated pig pituitary cells to secret GH in vivo.