Sphingolipids, including sphingosine and sphinganine, are one of the major classes of lipids. They serve as constituents of cell membranes and lipid rafts and aid in the performance of cell-cell communication and adhesion. Abnormal levels of sphingolipids in the aqueous humor can indicate impaired sphingolipid metabolism and associated ocular pathologies. Sphingolipids can be extracted from the aqueous humor by the methyl-tert-butyl ether (MTBE) lipid extraction method and subsequently analyzed by liquid chromatography-mass spectrometry (LC-MS). This chapter describes a modified protocol for an MTBE lipid extraction from the aqueous humor, followed by analysis with ultrahigh-performance liquid chromatography-mass spectrometry (UHPLC-MS).

The integration of complementary analytical platforms is nowadays the most common strategy for comprehensive metabolomics analysis of complex biological systems. In this chapter, we describe methods and tips for the application of a mass spectrometry multi-platform in Alzheimer\'s disease research, based on the combination of direct mass spectrometry and orthogonal hyphenated approaches, namely, reversed-phase ultrahigh-performance liquid chromatography and gas chromatography. These procedures have been optimized for the analysis of multiple biological samples from human patients and transgenic animal models, including blood serum, various brain regions (e.g., hippocampus, cortex, cerebellum, striatum, olfactory bulbs), and other peripheral organs (e.g., liver, kidney, spleen, thymus).

The chemical and biologically active characterization of jujube samples (fruits, cores, and leaves) were carried out by the integrated nontargeted metabolomics and bioassay. Firstly, collision cross-section values of active compounds in jujubes were determined by ultrahigh-performance liquid chromatography coupled with ion mobility quadrupole time-of-flight mass spectrometry. Then, a multidimensional statistical analysis that contained principal component analysis, partial least squares-discriminant analysis and hierarchical clustering analysis was employed to effectively cluster different tissues and types of jujubes, making identification more scientific. Furthermore, angiotensin-converting enzyme (ACE) and 2, 2-diphenyl-1-picrylhydrazyl (DPPH) were used to evaluate the quality of jujubes from a double activity dimension. The analytical results obtained by using ACE and DPPH to evaluate the quality of jujube were different from multivariate statistics, providing a reference for the application of jujube. Therefore, integrating chemical and biological perspectives to evaluate the quality of jujube provided a more comprehensive evaluation and effective reference for clinical needs.

Despite the considerable steps taken in the last decade in the context of antineoplastic drug (AD) handling procedures, their mutagenic effect still poses a threat to healthcare personnel actively involved in compounding and administration units. Biological monitoring procedures usually require large volumes of sample and extraction solvents, or do not provide adequate sensitivity. It is here proposed a fast and automated method to evaluate the urinary levels of cyclophosphamide and iphosphamide, composed of a miniaturized solid phase extraction (µSPE) followed by ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) analysis. The extraction procedure, developed through design of experiments (DoE) on the ePrep One Workstation, required a total time of 9.5 min per sample, with recoveries of 77-79% and a solvent consumption lower than 1.5 mL per 1 mL of urine sample. Thanks to the UHPLC-MS/MS method, the limits of quantification (LOQ) obtained were lower than 10 pg/mL. The analytical procedure was successfully applied to 23 urine samples from compounding wards of four Italian hospitals, which resulted in contaminations between 27 and 182 pg/mL.

Huangqi Guizhi Wuwu decoction (HGWWD) is a widely used traditional Chinese medicine (TCM) preparation for the treatment of ischemic stroke and diabetes peripheral neuropathy. However, the material basis for the efficacy of HGWWD remains unclear. In this study, a rapid, sensitive and selective ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) method was developed to separate and identify the absorbed components and metabolites of HGWWD in rat plasma after oral administration for the first time. By comparing the retention time, high-resolution mass spectrometry primary and secondary mass spectrometry data of blank plasma and drug-containing plasma, a total of 42 constituents, including 24 prototype compounds and 18 metabolites, were identified or tentatively characterized. The results indicated that monoterpenes, flavonoids, organic acids, amino acids, gingerols and alkaloids were main prototype compounds in rat plasma, and flavonoid-related metabolites, organic acid-related metabolites and gingerol-related metabolites were major metabolites. It is concluded the developed UHPLC-Q-TOF-MS method with high sensitivity and resolution is suitable for identifying and characterizing the absorbed components and metabolites of HGWWD, and the results will provide important data for further study on the relationship between the chemical constituents and pharmacological activities of HGWWD.

This study presents an innovative, efficient, environmentally friendly and rapid mechanical inclusion extraction (MIE) method for active ingredients in functional food. 2-hydroxypropyl-beta-cyclodextrin was used as the inclusion reagent, and water was used as the extraction solvent in MIE. The experimental parameters affecting the extraction efficiency of the target compounds were systematically investigated using single-factor experiments and surface response methodology optimization. The method showed satisfactory linearity (coefficient of determination > 0.991), precision (0.02 % to 4.89 %), limit of detection (1.1-11.3 ng/mL), and recoveries of 80.4-108.7 % and 86.3-112.3 % at spiked concentration levels of 1 and 5 μg/mL, respectively. Consequently, the MIE method provided a novel green alternative and extended its applications for the simultaneous extraction of hydrophobic and hydrophilic compounds from functional food.

Various forms of mycotoxins commonly exist in food and pose a significant risk to human health. Here a comprehensive suspect and nontarget screening strategy for both parent and modified mycotoxins was developed using ultrahigh-performance liquid chromatography-high resolution mass spectrometry (UHPLCHRMS). We constructed an in-house MS/MS database containing 82 mycotoxins in 8 categories. Then fragmentation characteristics of different classes of mycotoxins were rapidly extracted by a Python program \"Fragmentation pattern screener (FPScreener)\" and nontarget screening rules were determined by analyzing the frequencies and average intensities of fragmentation characteristics. Using the suspect and nontarget screening strategy, we successfully identified six parent mycotoxins and eight modified mycotoxins with different confidence levels in contaminated wheat and flour samples. This strategy enables screening of unknown parents and modified mycotoxins in food matrices with corresponding fragmentation characteristics.

A novel pH-responsive magnetic graphene oxide composite (MGO@PEI-BA) is proposed for the first time as an adsorbent for the rapid capture and detection of nucleosides (cytidine, uridine, guanosine, and adenosine). The morphology, structure, and magnetic properties of the composite were evaluated using various characterization techniques. The results indicated that the composite was successfully fabricated. A series of parameters that affect extraction and elution were optimized through one-factor-at-a-time and Box-Behnken design of response surface methodology (BBD-RSM). The unique layered structures and easily accessible active sites of the composite facilitated molecular transport, resulting in instantaneous equilibrium of nucleosides adsorption within 5 min. Based on this study, a magnetic dispersive micro-solid-phase extraction (MD-μ-SPE) method assisted by the MGO@PEI-BA was developed in combination with UHPLC-UV analysis for the determination of nucleosides in rat urine. Under the optimum conditions, a wide linear range (10-2000 ng mL-1), good linearity (r > 0.99), low detection limits (1-3 ng mL-1), low relative standard deviations (RSDs ≤ 3.9%), and satisfactory recoveries (82.7-96.3%) were achieved. These results demonstrate that the MGO@PEI-BA is an excellent adsorbent for extracting nucleosides from biological samples.

An ultrahigh-performance liquid chromatography coupled with ion mobility quadrupole time-of-flight mass spectrometry method was developed for the separation and identification of phenols, organic acids, flavonoids and curcumin in different species of ginger. The parameters affecting the separation and response of liquid chromatography, including the stationary phase and mobile phase, were systematically investigated and optimized. To further identify the differential metabolites in the six types of samples, a chemometric approach was introduced. Principal component analysis, cluster analysis and partial least squares discriminant analysis were used to identify the major components in the samples and to compare the compositional differences between the various samples. In addition, antioxidant experiments were designed to investigate the differences in antioxidant activity among the six ginger samples. The method showed good linearity (R2 ≥0.9903), satisfactory precision (RSD% ≤ 4.59 %), low LOD (0.35-25.86 ng/mL) and acceptable recovery (78-109 %) and reproducibility (RSD% ≤ 4.20 %). Therefore, the method has great potential for application in the compositional analysis and quality control of ginger.

Food contamination with pesticides poses significant risks to consumer safety and undermines confidence in food supply chains. Detecting pesticides in food samples is a challenging task that requires efficient extraction techniques. This study aims to compare and validate two microextraction techniques, μSPEed and μQuEChERS-dSPE, for the simultaneous extraction of eight pesticides (paraquat, thiabendazole, asulam, picloram, ametryn, atrazine, linuron, and cymoxanil) from wastewater samples. A good analytical performance was obtained for both methodologies, with selectivity, linearity in the range 0.5-150 mg L-1 with coefficients of determination up to 0.9979, limits of detection (LODs) and limits of quantification (LOQs) ranging from 0.02 to 0.05 mg L-1 and from 0.06 to 0.17 mg L-1, respectively, precision below 14.7 mg L-1, and recoveries from wastewater samples in the range of 66.1-99.9%. The developed methodologies are simpler, faster, and require less sample and solvent volumes than conventional methodologies, having a lower impact on the environment. Nevertheless, the μSPEed approach was found to be more efficient, easier to perform, and with a higher greener profile. This study highlights the potential of microextraction techniques for the analysis of pesticide residues in food and environmental samples. Overall, it presents a fast and efficient method for the analysis of pesticides in wastewater samples, which can be useful for monitoring and controlling pesticide contamination in the environment.