BackgroundFunctional dyspepsia (FD) is a prevalent gastrointestinal disorder with limited long-term efficacy of conventional treatments. Ganhai Weikang Capsules (GHWKCs), a classic traditional Chinese medicine formula, has shown remarkable clinical efficacy in alleviating FD symptoms, but its underlying mechanisms, particularly on the gut microbiota-host metabolism axis, remain unclear.MethodsWe performed chemical profiling of GHWKCs using UPLC-LTQ-Orbitrap-MS and established an FD rat model mimicking the TCM pathogenesis of “spleen deficiency and qi stagnation”. We evaluated gastrointestinal motility, histopathology, serum gastrointestinal hormones, and tissue inflammatory factors. Integrated duodenal metabolomics, cecal 16S rRNA sequencing, and network pharmacology were applied to explore the regulatory mechanisms, with Spearman correlation analysis conducted between differential metabolites and gut microbiota.ResultsA total of 63 chemical metabolites were identified in GHWKCs. GHWKCs significantly improved gastric emptying and intestinal propulsion, normalized gastrointestinal hormone levels, and inhibited pro-inflammatory cytokine secretion in FD rats. Metabolomics revealed 18 differential metabolites mainly involved in fatty acid β-oxidation and primary bile acid biosynthesis. Gut microbiota analysis showed GHWKCs counteracted dysbiosis, particularly increasing Coriobacteriales and Actinobacteriota abundance. Correlation analysis confirmed a positive correlation between Coriobacteriales and glycocholic acid. Network pharmacology identified 31 active metabolites, 213 intersection targets, and key pathways including PI3K-Akt, MAPK, and Toll-like receptor signaling.ConclusionGHWKCs ameliorate FD through multi-target mechanisms involving improved motility, reduced inflammation, and restored microbiota-metabolic homeostasis. The Coriobacteriales-glycocholic acid axis represents a putative core regulatory pathway bridging TCM theory and modern molecular mechanisms, providing a preclinical mechanistic framework for TCM-based therapy of FD.

BACKGROUND:Vascular dementia (VD), a prevalent neurodegenerative disorder that stems from chronic cerebral hypoperfusion, poses a substantial clinical challenge given the scarcity of efficacious treatment options. While ginsenoside Rg1 (Rg1) has demonstrated neuroprotective and antioxidative effects in various models of neurodegenerative disease, the mechanisms underlying its therapeutic potential in VD pathogenesis have yet to be systematically elucidated. PURPOSE:This study investigate the therapeutic potential of Rg1 in VD using a bilateral common carotid artery occlusion (2-VO) rat model and simultaneously explored the molecular mechanisms underlying its pharmacological effects. METHODS:To systematically assess the therapeutic efficacy of Rg1 on VD, we employed a well-established rat model of 2-VO. Behavioral outcomes were evaluated using standardized tests, histopathological changes were analyzed following histologic staining, and oxidative stress markers were quantified through biochemical analyses. Additionally, untargeted metabolomic profiling of serum and brain tissues was performed using UPLC-LTQ-Orbitrap MS, followed by targeted metabolomics to quantify essential amino acids and neurotransmitters. Additionally, integrated network pharmacology, transcriptomics, molecular docking, microscale thermophoresis (MST), qRT-PCR and western blotting were performed to facilitate a detailed investigation of the therapeutic potential of Rg1 and its molecular mechanisms in VD. RESULTS:Rg1 significantly ameliorated cognitive deficits and neuronal damage in rats with VD. Metabolomics revealed its unique ability to restore amino acid homeostasis and rebalance key neurotransmitters, including acetylcholine and glutamate. Mechanistically, Rg1 activated Adcy1 and Kdr, in turn enhancing cholinergic synapse integrity, and modulating the PI3K-AKT pathway to attenuate oxidative stress. Notably, molecular docking simulations displayed robust binding interactions between Rg1 and target proteins (all binding energies <-7 kcal/mol), and microscale thermophoresis (MST), qRT-PCR and western blotting findings revealed high consistency with multi-omics predictions. CONCLUSION:Thie findings of this reveals novel evidence that Rg1 alleviates VD by restoring amino acid homeostasis and neurotransmitter equilibrium, thereby activating Adcy1/Kdr-mediated cholinergic synapse and PI3K-AKT signaling pathway. These results position Rg1 as a promising phototherapeutic candidate for VD treatment.

Respiratory tract infection (RTI) continues to be a non-negligible cause of global incidence rate and mortality. Shufeng Jiedu formula (SFJD), a traditional Chinese remedy, is used for treating RTI, though its mechanisms are not well understood. The objective of this research was to uncover the underlying molecular mechanisms responsible for the effectiveness of SFJD on RTI. Using UHPLC-Q-Orbitrap HRMS assays, the chemical compounds in SFJD's ethanol and aqueous extracts fractions were identified. The targets of these herbal compounds and RTI-related targets were acquired from various database. Key SFJD-RTI targets were analyzed using Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and tissue/organ-specific analyses. Molecular docking, data mining and dynamic simulations were employed to evaluate interactions between major targets and the main compounds. A total of 94 and 31 compounds were determined in the aqueous and ethanol extract fraction, respectively. Many of these compounds demonstrated good oral bio-availability (60/99). GO analysis revealed significant involvement in inflammatory response and gene silencing processes, while KEGG pathways highlighted connections to viral infections and signaling pathways. The lung was the primary target organ screed out by Biogps databases and 15 compounds as well as eight key genes were associated. A set of 15 compounds were virtually fitted into the active site of eight critical targets. Isaindigotone and resveratrol could form hydrogen bond interactions with FOS and JUN, respectively. FOS and JUN were classified as differential genes after the COVID-2019, HCoV-229E and H1N1 infected. The comprehensive results obtained from MD simulation, MMPBSA, and SASA analysis highlighted the compounds' potential to serve as effective inhibitors of FOS and JUN proteins. Lastly, resveratrol exerted a high inhibitory effect on the influenza virus based on MDCK cells. In summary, we have provided a framework that integrated network pharmacology with multiple virus infection models to identify synergistic compounds based on network topology.

Bioenergetic therapy based on tumor glucose metabolism is emerging as a promising therapeutic modality. To overcome the poor bioavailability and toxicity of arenobufagin (ArBu), a MOF-derived intelligent nanosystem, ZIAMH, was designed to facilitate energy deprivation by simultaneous interventions of glycolysis, OXPHOS and TCA cycle. Herein, zeolitic imidazolate framework-8 was loaded with ArBu and indocyanine green, encapsulated within metal-phenolic networks for chemodynamic therapy and hyaluronic acid modification for tumor targeting. ZIAMH nanoparticles can release ArBu in the tumor microenvironment for chemtherapy, and ICG enables photothermal therapy under near-infrared laser irradiation. In vitro and in vivo mechanism studies revealed that the ZIAMH nanoplatform downregulated glucose metabolism related genes, resulting in the reduction of energy substances and metabolites in tumors. Additionally, it significantly promoted cell apoptosis by upregulating pro-apoptotic proteins such as Bax, Bax/Bcl-2, cytochrome C. Animal studies have shown that the tumor inhibition efficiency of ZIAMH nanomedicines was three fold higher than that of free drugs. Therefore, this study provides a new strategy for glucose metabolism-mediated bioenergetic therapy and PTT/CDT/CT combined therapy for tumors.

BackgroundJianwei Xiaoshi oral liquid (JWXS), a classical traditional prescription comprising various edible medicinal plants, has demonstrated significant efficacy in treating paediatric indigestion. It originates from Jianpi Pill, which is developed in the Ming Dynasty and nourishes the spleen and regulates gastrointestinal function. However, the specific molecular mechanisms involved remain unclear.MethodsTo elucidate the material base of JWXS and its underlying mechanism in treating dyspepsia, the UHPLC-Q-Orbitrap HRMS method and network pharmacology were utilized. This was followed by pharmacological experiments, transcriptomics analyses and gut microbiota studies to further investigate the effects of JWXS on dyspepsia.ResultsA total of 105 compounds, mainly flavonoids, alkaloids, organic acids and cyclic peptides, were identified. According to the five principles of generic drug properties, 43 candidate compounds were screened out. Their efficacy was verified through gastric emptying and intestinal propulsion experiments. Transcriptomic analysis revealed that JWXS primarily alleviated dyspepsia symptoms by regulating the secretion of 8 key proteins in the pancreatic secretion pathway. The differences in the gut microbiota, as identified through 16S rRNA and ITS2 sequencing, were subsequently more pronounced than those observed in the bacterial microbiota of the model group. In total, 15 differential bacteria and 16 differential fungi were identified. Targeted metabolomics analysis of SCFAs revealed a significant decrease in valeric acid (VA), acetic acid (AA), and isovaleric acid (IVA) levels in the model group, which were restored to the corresponding levels after the administration of JWXS. Correlation analysis revealed that VA, AA, and IVA were positively correlated with Lactobacillus and Bacteroides, and negatively correlated with Aspergillus and Candida. This further suggested that JWXS might alleviate symptoms of indigestion by regulating the composition of the microbiota, increasing the variety and quantity of beneficial bacteria, reducing fungal contamination, and further increasing the levels of SCFAs in the body.ConclusionJWXS improved functional dyspepsia in immature rats via a mechanism involving the regulation of the secretion of 8 key proteins in the pancreatic secretion pathway and the amelioration of flora disorders.

The quality and flavor of Shanxi Aged Vinegar (SAV) are closely related to its aging time. In this study, for the first time, UPLC-Q-TOF/MS combined with non-targeted metabolomics techniques were employed to analyze SAV samples from 5 different years. The antioxidant activities were evaluated using ABTS and DPPH assays. The non-targeted metabolomics results identified a total of 99 metabolites, among which 24 were significantly associated with the aging time of SAV and were identified as potential markers of vintage, including key components such as organic acids, amino acids, and polyphenols. They contribute significantly to the unique flavor and nutritional value of SAV. Specifically, in SAV aged for 10 years, the contents of citric acid, L-malic acid, L-glutamic acid, protocatechuic acid, and hydroferulic acid were significantly higher than in other vintages. The results of ABTS and DPPH antioxidant activities indicated a positive correlation between the antioxidant activity of SAV and its aging time. Among the 12 identified polyphenols, chlorogenic acid may be the primary contributor to the antioxidant results of SAV. This study provides important references for further exploration of the health value and antioxidant mechanisms of SAV.

Background Bufonis Venenum (BV) is a traditional animal-based Chinese medicine with therapeutic effects against cancer. However, its clinical use is significantly restricted due to associated cardiovascular risks. BV's value in China's market is typically assessed based on “content priority,” focusing on indicator components. However, these components of BV possess both antitumor activity and toxicity, and the correlation between the antitumor activity and toxicity of BV has not yet been elucidated. Purpose This study employs an integrated multi-omics approach to identify bufadienolide Q-markers and explore the correlation between BV's antitumor activity and toxicity. The aim is to establish a more comprehensive method for BV's quality. Methods Normal zebrafish and HepG2 xenograft zebrafish were chosen as activity and toxicity evaluation models. Ultra-high performance liquid chromatography (UHPLC) coupled with a linear ion trap orbitrap (LTQ-Orbitrap) mass spectrometry was used to quantify eight batches of BV and key “toxic and effective” components were screened out. Transcriptomic and metabolomic analyses were performed to elucidate the regulatory mechanisms underlying the antitumor activity and cardiovascular toxicity of the key components in BV. Results Eight key “toxic and effective” compounds were identified: resibufogenin, cinobufagin, arenobufagin, bufotalin, bufalin, gamabufotalin, desacetylcinobufagin, and telocinobufagin. The findings showed that bufalin and cinobufagin interfered with calcium homeostasis through CaV and CaSR, induced cardiotoxicity, and upregulated CASP9 to activate myocardial cell apoptosis. However, desacetylcinobufagin exhibited greater potential in terms of anti-tumor effects. Combining the results of untargeted and targeted metabolomics revealed that desacetylcinobufagin could have a callback effect on differential lipids and correct abnormal energy and amino acid metabolism caused by cancer, similar to cinobufagin and bufalin. Microscale thermophoresis (MST) ligand binding measurements also showed that the binding of desacetylcinobufagin to GPX4 has a more potent ability to induce ferroptosis in tumor cells compared to cinobufagin. Conclusion An innovative evaluation method based on the zebrafish was developed to investigate the relationship between the toxicity and efficacy of BV. This study identified toxicity and activity Q-markers and explored the mechanism between the two effects of BV. The research data could offer valuable insights into the efficacy of BV. Additionally, desacetylcinobufagin, an active ingredient with low toxicity, was found to enhance the quality of BV.

Plucking position and harvest time influence the metabolites of vine tea which are closely related to the taste quality and antioxidant activity. In this study, 75 vine tea samples produced by three plucking positions and harvested in five months were analyzed by untargeted metabolomics, and in vitro and in vivo antioxidant activities of different vine teas were evaluated. 37 compounds were tentatively identified through various methods. Compared with the high-concentration results, VIP scores of the major components in the low-concentration model were much higher and were filtered as characteristic markers. Chemometrics analysis showed that plucking position had a stronger influence on vine tea quality than harvest time. 16 identified metabolites, including flavonoids, organic acids and catechins, were found to be responsible for the recognition of the plucking position. Vine tea infusion made of young green leaves exhibited higher scavenge radical activity in vitro, and tender tip leaves exhibited higher antioxidant activities in vivo zebrafish, which may be due to the presence of a higher concentration of flavonoids in their phytochemical profile. This research is anticipated to provide useful enlightenment for the effective utilization of vine tea resources.

BACKGROUND:Panax notoginseng (PN) was an edible Chinese herbal medicine. PN's current quality control standard cannot precisely match the traditional grading experience.PURPOSE:In this study, under the guidance of the traditional grading experience, the combined metabolomics and biological effect evaluation were used to reveal the distinct chemical quality of PN.METHODS:The quality of PN was evaluated by traditional experience and characterized by the electronic tongue. A zebrafish myocardial ischemia model was developed to verify the grading experience. The untargeted metabolomics method was used to identify and validate the grading markers of PN.RESULTS:The taste was the critical indicator for classifying the quality. Based on the experience sensory scores (ranged from 47.0 to 87.8), PNs could be divided into two grades. The experience scores were significantly associated with umami and richness of the electronic tongue(p<0.01). Besides, superior PN showed substantially stronger anti-myocardial ischemia activity(p<0.001). Thirty-nine differential components were found using UHPLC-LTQ-Orbitrap MS, of which 22 were identified. A new kind of grading quality markers alkynols in PN-associated efficacy was identified, which revealed stronger anti-myocardial ischemia activities than saponin.CONCLUSION:This study evaluated PN through untargeted metabolomics and anti-myocardial ischemia evaluation of zebrafish and proposed the critical role of alkynols in PN's quality classification.

The new trend of adulteration detection of virgin olive oil (VOO) is exploiting a rapid, real-time analytical method to make on-site screening possible. Therefore, the main objective of this work is to develop an analytical method capable of detection and quantifying adulteration of VOO by combining the rapid evaporative ionization mass spectrometry (REIMS) with multivariate statistical analysis. With the sampling by electric soldering iron, this method can effectively distinguish VOO from other plant oils (blended olive oil, sunflower oil, peanut oil, colza oil, corn oil, and soybean oil) with an accuracy of 99.29%. Moreover, principal component analysis-linear discriminate analysis (PCA-LDA) shows that the method is able to identify the adulterated VOO with other oils in different levels. The quantitative prediction models were established using partial least squares (PLS) for the adulteration of VOO with four types of different oils. High correlation coefficients above than 0.97 were achieved in an approximated linear relationship. The result indicates that REIMS combined with appropriate multivariate statistics can be used for VOO adulteration analysis with detection limits as low as 5%.This method could serve as a rapid and real-time approach for authentication and adulteration analysis of VOO. This is also the first application of REIMS coupled with electric soldering iron as sampling tool in the analysis of plant oil samples without the assistance of any additional ionizing medium.

Herpetospermum pedunculosum (Ser.) C. B. Clarke (Family Cucurbitaceae) is a dioecious plant and has been used as a traditional Tibetan medicine for the treatment of hepatobiliary diseases. The component, content, and difference in volatile compounds in the female and male buds of H. pedunculosum were explored by using headspace solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) technology and multivariate statistical analysis. The results showed that isoamyl alcohol was the main compound in both female and male buds and its content in males was higher than that in females; 18 compounds were identified in female buds including 6 unique compounds such as (E)-4-hexenol and isoamyl acetate, and 32 compounds were identified in male buds, including 20 unique compounds such as (Z)-3-methylbutyraldehyde oxime and benzyl alcohol. (Z)-3-methylbutyraldehyde oxime and (E)-3-methylbutyraldehyde oxime were found in male buds, which only occurred in night-flowering plants. In total, 9 differential volatile compounds between female and male buds were screened out, including isoamyl alcohol, (Z)-3-methylbutanal oxime, and 1-nitropentane based on multivariate statistical analysis such as principal component analysis (PCA) and orthogonal partial least squares discrimination analysis (OPLS-DA). This is the first time to report the volatile components of H. pedunculosum, which not only find characteristic difference between female and male buds, but also point out the correlation between volatile compounds, floral odor, and plant physiology. This study enriches the basic theory of dioecious plants and has guiding significance for the production and development of H. pedunculosum germplasm resources.

A high performance liquid chromatography-diode array detector (HPLC-DAD) was used to establish the HPLC fingerprint. Chemometrics methods were used to discriminate against the gender of flower buds of Populus tomentosa based on areas of common peaks calibrated with the HPLC fingerprint. The score plot of principal component analysis (PCA) showed a clear grouping trend (R2X, 0.753; Q2, 0.564) between female and male samples. Two groups were also well discriminated with orthogonal partial least squares-discriminant analysis (OPLS-DA) (R2X, 0.741; R2Y, 0.980; Q2, 0.970). As the hierarchical clustering analysis (HCA) heatmap showed, all samples were separated into two groups. Four compounds were screened out by S-plot and variable importance in projection (VIP > 1.0). Two of them were identified as siebolside B and tremulacin. This study demonstrated that HPLC fingerprints combined with chemometrics can be applied to discriminate against dioecious plants and screen differences, providing a reference for identifying the gender of dioecious plants.

The differences in metabolites in male and female individuals of dioecious Morus alba L. (Moraceae) are usually ignored and lack study. In the present study, 58 leaves and 61 stem barks from male and female individuals were analyzed by untargeted metabolomics via headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS) coupled with chemometrics, including principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA). A total of 66 and 44 metabolites were identified from leaves and stem barks, respectively. Four and eight differential metabolites among candidate metabolites in leaves and stem barks from male and female individuals were identified. Moreover, females possessed stronger antioxidant activity than males. This is the first report where untargeted metabolomics coupled with chemometrics was used to analyze the different metabolites and to discriminate the gender of leaves and stem barks of dioecious M. alba. It provided the basis for further study of M. alba and reference value for researching dioecious plants.

Perfluoroalkyl substances (PFASs) have been identified as global pollutants and raise considerable food safety concerns. However, the development of an analytical method with satisfied pretreatment performance for PFASs with varying alkyl chain length in the fatty samples remains a challenge. We describe herein the preparation of superparamagnetic nanofluid, based on a new choline chloride/1-(o-tolyl)biguanide deep eutectic solvent (DES) system, for direct extraction of perfluoroalkyl carboxylic acids from edible oils. Target PFASs, especially the short-chain one, all possessed high recoveries (90-109% with intra-day and inter-day precision below 10%). This was achieved by adjusting the constituent ratio of DES for balancing the anion-exchange (electrostatic) interactions and hydrogen-bond interactions. Employing the prepared nanofluid in magnetic microextraction, followed by high-resolution mass spectrum analysis, resulted in a rapid (15 min for pretreatment), simple, sensitive (detection limit: 0.3-1.6 pg g(-1)), and efficient method for the enrichment and determination of trace PFASs. Furthermore, the introducing of N-H center dot center dot center dot F weak force increased the pretreatment selectivity to effectively reduce the matrix effect. At the end of the study, the proposed methodology was successfully applied to the analysis of target analytes in real samples.

The quality and flavor of green tea can be affected by various factors, which are closely related to the metabolite composition of tea. In this study, 66 Xinyang Maojian tea (XYMJ) samples produced by four cultivars, grown in different elevations and manufactured by different processing methods were analyzed by untargeted ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry and chemometrics. 1912 ion features were detected and 95 metabolites were identified tentatively through a customized in-house library. Projection to latent structures discriminate analysis showed high capability to explain the cultivar variation. 54 metabolites were found to be responsible for the differentiation of the four cultivars. 27 metabolites including epigallocatechin gallate, epicatechin gallate, theanine, theogallin showed close correlation with elevation, resulting enhanced umami flavor of the high elevation tea. The differences between manual and mechanical tea were not significant. This comprehensive study is of great reference value for other types of tea.

Traditional Chinese Medicine (TCM) is widely used and tested in ethnic communities for a long history. A large number of TCM formulae, as the centralized expression of “theory, cure, recipe, and medicine”, were preserved and recorded in way of relatively completed ingredients and lack of well-noted efficacies. The main objective of this paper is to translate dosage of each herb in a TCM formula into intensity of its efficacies in a quantitative and visual way. In this study, firstly, we estimated the intensity between a herb and its efficacies quantitatively based on clinical case data, which can reflect the difference between main and secondary efficacies of a herb. Then, to calculate and visualize the interaction between different efficacies from different herbs, we have proposed a method based on the Efficacy Semantic Network (ESN). It includes three steps: (1) Simple cumulation; (2) Adjustments; (3) Main efficacies selection. We tested on a dataset of 70 TCM formulae, results show that maximum performance measure reaches 0.818 which prove that the proposed approach could help standardize existing and predict potential efficacies of TCM in a quantitative way. It shows a great potential of semantic technologies to represent, calculate and visualize the TCM knowledge. However, more efforts are still required to investigate how to represent other fuzzy and rule-based knowledge and bring more data of TCM formulae for extensive tests.

A Rapid Evaporative Ionization Mass Spectrometry (REIMS) method, innovatively combined with electric soldering iron, was developed for the identification of honey authentication and adulteration. Monofloral honey (acacia, rape, chaste, jujube, citrus, medlar), syrup (corn and rice), and simulated polyfloral honey and adulterated syrup-honey samples were evaluated. The classification of botanical origins of honey samples were achieved with a total correct rate of 99.66% through the proposed principal component analysis-linear discriminate analysis (PCA-LDA). The real-time recognition of honey samples was obtained near-instantaneously. The monofloral honey samples (acacia honey, chaste honey, rape honey) could be well distinguished from polyfloral honey (their binary and ternary mixtures). High-value honey (acacia honey) adulterated with low-value honey (rape honey) could be detected from the level of 40% down to 5%. C3 syrup (rice syrup) and C4 syrup (corn syrup) and their adulterated syrup-honey samples in different levels (5, 10, 15, 20, 30, 40%) were also appropriately identified in relation to the adulteration level in a near linear tendency. The detection limit can reach 5–10% adulteration level which can meet the requirement of real adulteration identification. The results of this study indicate that REIMS can serve as a promising rapid and real-time approach for authentication and adulteration analysis of more diverse food samples with the application of contact heating by using a soldering iron.

以“新大坪”马铃薯为试材,采用液化—糖化—发酵—蒸馏工艺制备马铃薯蒸馏酒,利用顶空固相微萃取法(HS-SPME)提取马铃薯及马铃薯蒸馏酒中的香气成分、气相色谱-质谱联用(GC-MS)检测、结合计算机检索对其中香气成分进行分离鉴定,并应用色谱峰面积归一法计算各成分的相对含量.结果表明,醇类和酯类是马铃薯蒸馏酒挥发性物质的主要成分,醇类相对含量为64.39％,酯类为24.27％;而马铃薯的主要香气成分是醇,羰基化合物及其他成分,分别占13.29％、17.53％、17.48％,说明将马铃薯制备成蒸馏酒后产生了更多的酯类和醇类,而羰基化合物显著减少.

In this study, a simple and rapid multi-residue screening method for the quantification and confirmation of 100 multi-class veterinary drugs in milk powder by the Quick, Easy, Cheap, Effective, Rugged, and Safe (QuEChERS) method combined with liquid chromatography coupled to quadrupole time-of-flight mass spectrometry was developed. The veterinary drugs investigated belonged to several families such as macrolides, sulfonamides, quinolones, tetracyclines, nitroimidazoles, benzimidazoles, β-agonists, hormones, and tranquilizers. The samples were extracted using a modified QuEChERS procedure: (i) dissolved in a solution of Na2EDTA–McIlvaine buffer (0.1 M, pH 4), (ii) extracted with 5% acetic acid in acetonitrile, and (iii) purified using a C18 sorbent. The linearity, sensitivity, accuracy, repeatability, and reproducibility of the method were fully validated. The average recoveries for the majority of analytes (92.9%) ranged from 70 to 120% at four concentration levels, while the repeatability and reproducibility ranged from 1.1 to 20.1%. A coefficient of correlation (γ2) of 0.995 or above was obtained for all the compounds. The range of the limit of quantification for these compounds in the milk powder was from 0.1 to 25 μg kg−1. More than 93.0% of the analytes were detected when present at 10 μg kg−1 or less in the milk powder samples. For the screening method, the data of the precursor and product ions of the target analytes were simultaneously acquired under the All Ions MS/MS mode in a single run. A homemade library including the elemental composition, retention time, and full-scan product ions was utilized for the confirmation and identification of the target compounds. The applicability of the screening method was verified by applying to real milk powder samples, and certain veterinary drugs were detected in some cases.