Biomedical Chromatography

Ali, Imran; Mimouni, Fatima Zohra; Belboukhari, Nasser; Sekkoum, Khaled; Locatelli, Marcello; Demir, Ersin; Yusuf, Kareem

doi: 10.1002/bmc.6004pmid: 39237855

Thirteen flavanone racemates were successfully separated using a Chiralpak® IA column and isopropanol‐hexane (50:50, v/v). The mobile phase flow rate and detection wavelength were 0.5 mL/min and 254 nm. The retention times values ranged from 5.50 and 56.45 min. The values of the retention, separation, and resolution factors ranged from 0.63 to 21.67, 1.12 to 2.45, and 0.13 to 11.94. The docking binding energies ranged from −6.2 to −8.2 kcal/mol, showing enthalpy‐determined host‐guest complex formation. The molecular docking results and the experimental data were agreed well. The results showed that S‐enantiomers had stronger bindings with chiral selectors compared to R‐enantiomers. Consequently, the R‐enantiomers eluted first followed by S‐enantiomers. The reported method is highly useful to determine the enantiomeric composition of the reported flavanone in any sample.

Zhang, Kejun; Wang, Haizhou

doi: 10.1002/bmc.6011pmid: 39300201

Huperzine A is a reversible and selective cholinesterase inhibitor and has been approved for the treatment of Alzheimer's diseases. In this study, we developed a highly sensitive and specific ulta‐high‐performance liquid chromatography–tandem mass spectrometry method for the determination of Huperzine A in rat plasma. An aliquot of 50 μL of rat plasma sample was pretreated with 200 μL of acetonitrile‐methanol (v/v; 1:1) containing 0.2% formic acid followed by solid phase extraction. The resulting sample was separated on a Waters ACQUITY BEH C18 column using acetonitrile and water containing 0.2% formic acid as mobile phase, at a flow rate of 0.3 mL/min. Multiple‐reaction monitoring (MRM) mode was used for quantitative analysis of Huperzine A in positive electrospray ionization. In the concentration range of 0.01–10 ng/mL, Huperzine A showed excellent linearity with correlation coefficient > 0.998. The intra‐ and inter‐day RSD% were less than 9.7%, while the RE% ranged from −6.7% to 10.0%. The mean recovery was >84.5%. The validated method was demonstrated to be selective, sensitive, and reliable, which has been successfully applied to pharmacokinetic study of Huperzine A in rat plasma. Huperzine A displayed a long half‐life in rat plasma and high oral bioavailability.

Tao, Jiayue; Li, Hanyi; Jin, Mingxuan; Shen, Wenchao; Liu, Siqi; Li, Dan; Hou, Jincai; Wang, Rufeng

doi: 10.1002/bmc.5997pmid: 39225114

Yigong San (YGS) is a traditional Chinese medicine formula used for pediatric anorexia, chronic atrophic gastritis, and irritable bowel syndrome. In this study, the excretion of eight main compounds, including liquiritin; isoliquiritin; hesperidin; ginsenosides Rb1, Re, and Rg1; and atractylenolides I and II, in rat urine, feces, and bile, was investigated by ultra‐high performance liquid chromatography–tandem mass spectrometry. The results showed that the cumulative excretion rates of the compounds in rat urine, feces, and bile were 0.018–1.15%, 0.024–19.89%, and 0.0025–0.72%, respectively. Among the eight compounds detected, liquiritin was the richest in urine, and ginsenosides Re and Rg1 and atractylenolide I were mainly found in feces and bile. In summary, the main components of YGS are excreted via multiple approaches. Liquiritin is mainly through urine, whereas isoliquiritin; hesperidin; ginsenosides Rb1, Re, and Rg1; and atractylenolides I and II are mainly through feces. The excretion of these compounds in bile is usually positively correlated with that in feces. This study lays a foundation for further pharmacological research and application of YGS.

Aliyeva, Nilufer; Akgönüllü, Semra; Erdem, Arzum; Denizli, Adil

doi: 10.1002/bmc.5995pmid: 39189513

Activated protein C (APC), a serine protease produced from zymogen protein C (PC), is the key enzyme of the protein C pathway. APC has anticoagulant, anti‐inflammatory, and cytoprotective features. APC has recently been shown to significantly reduce coagulation as well as mortality in patients with severe sepsis. Herein, we aimed to develop an affinity support material that allows the purification of plasma APC for the first time. In this research, a novel APC‐specific DNA aptamer–based poly(2‐hydroxyethyl methacrylate‐glycidyl methacrylate) (poly(HEMA‐GMA/DNA‐Apt)) macroporous cryogel membrane at different molar ratios was prepared using affinity binding method and their potential for purification and identification of APC was investigated. The DNA aptamer–immobilized cryogels were characterized to examine their structural and morphological properties. The effect of pH, initial concentration, temperature, ionic strength difference, and flow rate changes was examined. Selectivity studies were performed in the presence of APC and competitive proteins, and cryogel support materials were shown to have a very high affinity for APC. Adsorption capacity was found to be 89.02 mg/g. Finally, NaCl revealed efficiency for APC desorption and the reuse of cryogels was successfully tested for ten cycles.

Sa, Yuping; Ding, Shuqin; Zhang, Yue; Wang, Weibiao; Wilson, Gidion; Ma, Feng; Zhang, Weiman; Ma, Xueqin

doi: 10.1002/bmc.5998pmid: 39193838

Given the limitations of untargeted metabolomics in precise metabolite quantification, our current research employed a novel approach by integrating untargeted and targeted metabolomics utilizing ultra‐high‐performance liquid chromatography quadrupole time‐of‐flight tandem mass spectrometry (UHPLC‐QTOF‐MS/MS) to analyze the metabolic profile and potential biomarkers for tuberculosis (TB). A cohort of 36 TB patients and 36 healthy controls (HC) was enlisted to obtain serum samples. Multivariate pattern recognition and univariate statistical analysis were employed to screen and elucidate the differential metabolites, whereas dot plots and receiver operating characteristic (ROC) curves were established for the identification of potential biomarkers of TB. The results indicated a distinct differentiation between the two groups, identifying 99 metabolites associated with five primary metabolic pathways in relation to TB. Of these, 19 metabolites exhibited high levels of sensitivity and specificity, as evidenced by the area under curve values approaching 1. Following targeted quantitative analysis, three potential metabolites, namely, L‐asparagine, L‐glutamic acid, and arachidonic acid, were demonstrated excellent discriminatory ability as evidenced by the results of the ROC curve, dot plots, and random forest model. Particularly noteworthy was the enhanced diagnostic efficacy of the combination of these three metabolites compared to singular biomarkers, suggesting their potential utility as serum biomarkers for TB diagnosis.

Guo, Jun Jie; Lu, Wen Tian; Liu, Yue Tao

doi: 10.1002/bmc.6014pmid: 39299923

Xiao–Jian–Zhong–Tang (XJZT) has the effect of warming the middle and tonifying the deficiency, easing the urgency and relieving pain according to the theory of traditional Chinese medicine (TCM), and is able to treat spleen deficiency type chronic atrophic gastritis (CAG). Metabolites of TCM in cecum contents are common metabolites of intestinal bacteria and hosts, which can reflect the metabolic status in disease states. The present work was performed to study the effect of XJZT against CAG coupled with the cecal metabolites analysis and bioinformatics. A total of nine prototypical components and 144 metabolites were firstly identified in the cecum metabolites of XJZT using ultra‐high performance liquid chromatography added to the quadrupole‐time of flight mass spectrometry (UHPLC‐Q‐TOF/MS), which underwent the metabolism of oxidation, reduction, methylation, and glucuronic acid reaction Furthermore, different prototypical compounds might metabolize into identical metabolites in the presence of intestinal flora. Bioinformatics was further used to correlate these metabolites with the disease and intestinal flora. Components and targets were screened by Cytoscape, and molecular docking of key targets and core components showed good binding ability. This study provided important information for exploring the mechanism of TCM formulae.

Mahmoud, Osama A.; Omran, Ahmed A.; Gomaa, Hosni A.; Binsaleh, Ammena Y.; Mohamed, Mahmoud A.

doi: 10.1002/bmc.6006pmid: 39275959

The efficacious treatment of muscle and joint pain relies heavily on etofenamate (ETO) and benzyl nicotinate (BN), which possess robust anti‐inflammatory and pain‐relieving properties when paired with methylparaben (MP) or benzyl alcohol (BA). In this study, we have established and validated innovative RP‐UPLC methods for assessing ETO and BN in the presence of MP or BA in their dosage forms, employing eight green tools to evaluate their eco‐friendliness and effectiveness. Reversed phase‐ultra‐performance liquid chromatography (RP‐UPLC) technique employs a flow rate of 0.3 mL/min on Waters Acquity UPLC BEH Column (C18, 1.7 μm, 100 mm × 2.1 mm), detection at 254 nm using a photo diode array (PDA) detector and mobile phase of 0.05 M KH2PO4 buffer, acetonitrile, and methanol (50:15:35, v/v/v) adjusted pH 6.0 with 0.2% triethylamine. For ETO, BN, MP, and BA, the calibration curves were linear and ranged from 0.005 to 1.0, from 0.001 to 0.2, from 0.002 to 0.08, and from 0.0001 to 0.1 mg/mL, respectively. The correlation value was 0.9999, and the accuracy findings ranged from 98.81% to 100.56%. Consequently, the methodology has been successfully implemented in assay testing for the pharmaceuticals in the presence of the MP or BA, demonstrating the high selectivity of these approaches. The present study presents the Blue Applicability Grade Index (BAGI), an innovative approach that complements green metrics in practical white analytical chemistry. According to the International Council for Harmonisation (ICH) criteria, the procedures were effectively validated.

Zhang, Jian‐Jia; Guan, Wei; Wang, Yue; Wang, Yu‐Xuan; An, Dong‐Qi; Hao, Zhi‐Chao; Li, Meng‐Meng; Kuang, Hai‐Xue; Chen, Qing‐Shan; Zhang, Li‐Li; Liu, Yan; Yang, Bing‐You

doi:

Chen, Jiahui; Chen, Liqiong; Li, Bin; Zhao, Qi; Cheng, Yan; Yan, Dongmei; Liu, Hongning; Li, Fei

doi: 10.1002/bmc.5996pmid: 39175367

Molnupiravir (MO) is a pyrimidine nucleoside anti‐SARS‐CoV‐2 drug. MO treatment could cause mild liver injury. However, the underlying mechanism of MO‐induced liver injury and the metabolic pathway of MO in vivo are unclear. In this study, metabolomics analysis and molecular biology methods were used to explore these issues. Through metabolomics analysis, it was found that the homeostasis of pyrimidine, purine, lysophosphatidylcholine (LPC), and amino acids in mice was destroyed after MO treatment. A total of 80 changed metabolites were detected. Among these changed metabolites, 4‐ethylphenyl sulfate, dihydrouracil, and LPC 20:0 was related to the elevation of alkaline phosphatase (ALP), interleukin‐6 (IL6), and nuclear factor kappa‐B (NF‐κB). The levels of 4‐ethylphenyl sulfate, dihydrouracil, and LPC 20:0 in plasma were positively correlated with their levels in the liver, suggesting that these metabolites were associated with MO‐induced liver injury. MO treatment could increase NHC and cytidine levels, activate cytidine deaminase (CDA), and increase LPC levels. CDA and LPC could increase the mRNA expression level of toll‐like receptor (TLR). The current study indicated that the elevation of hepatic TLR may be an important reason for MO leading to the liver injury.

Guo, Mingxin; Chu, Yujiao; Zhu, Wenjiao; Sun, Miaomiao; Lv, Qiang; Tang, Ruijie; Jiang, Xuping; Zhao, Jiahao; Tang, Zhian; Ma, Tieliang

doi: 10.1002/bmc.5967pmid: 39189519

This study aims to identify potential efficacy‐related biomarkers and investigate the mechanism of Youjing granule (YG) in improving spermatogenic function in rats based on metabolomics combined with network pharmacology. We obtained YG‐containing serum from Sprague–Dawley rats, compared it with control group serum and analyzed it using gas chromatography–mass spectroscopy to identify potential biomarkers and investigate the mechanism of YG in improving spermatogenic function in rats. Six important differential biomarkers, comprising putrescine, amidine, arginine, d‐fructose‐6‐phosphate, l‐proline and galactose, were identified in the YG‐containing serum and then used to explore the potential mechanisms. The ultra‐high‐performance liquid chromatography–high‐resolution mass spectrometry technology was adopted for the rapid separation, identification and analysis of chemical components of YG in blood. A total of 69 detected chromatographic peaks were revealed. The binding energy between core compounds and key proteins is low, among which dipsacoside B is the best. The outcomes suggest that YG may improve spermatogenic function in rats by facilitating the development of spermatogonial stem cells, counteracting oxidative stress and controlling cellular apoptosis. Youjing granule may also affect the energy required for sperm production or influence sperm growth and maturation.