Biomedical Chromatography

Jiang, Jun; He, Jinjin; Xiao, Shichang; Shenyuan, Jiayi; Chen, Tong; Pei, Dan

doi: 10.1002/bmc.5686pmid: 37277120

Iron overload is a risk factor for postmenopausal osteoporosis (PMOP) and lowering iron levels to regulate the labile plasma iron is the preferred therapy. Icariin (ICA), baohuoside I (BHS) and icaritin (ICT) are three flavonoids obtained from Epimedii Folium that are efficient in facilitating osteogenesis. In this study, an active flavonoid with dual effects of reversing iron overload and promoting osteogenesis was screened based on pharmacokinetics, iron complexation properties and the potential to downregulate iron overload, reversing PMOP. As a result, the in vivo absorption of three compounds was ICA > ICT > BHS, while the exposure in muscle and bone was BHS > ICT > ICA. In vitro complexation showed that only ICT complexed with Fe (III) at a 1:1 ratio on 3‐OH and the ICT–Fe (III) complex (m/z 424.3750) was identified by UPLC–Q‐TOF–MS. In vivo dynamic detection also showed that the concentration of ICT–Fe (III) complexes varied with the concentration of ICT in plasma. The behavioral blunting and bone loss in zebrafish induced by Fe (III) were significantly reversed by ICT in a dose‐dependent manner. Pharmacokinetic–pharmacodynamic analysis showed that ICT was negatively correlated with serum ferritin and positively correlated with osteogenic markers including alkaline phosphatase, osteocalcin and osteoprotegerin. Bone loss in ovariectomized rats was significantly altered after ICT intervention, with reduced serum ferritin levels and improved osteogenic marker levels. These results demonstrated that ICT had favorable musculoskeletal penetration and iron complexation capability to shrink labile plasma iron, showing superior performance in anti‐PMOP through the dual effects of reversing iron overload and promoting osteogenesis.

Xin, Ling; Liu, Shuo; Lou, Yanwei; Zhang, Jing; Lu, Qing; Zhao, Longshan; Wei, Xiuyan; Xiong, Zhili

doi: 10.1002/bmc.5693pmid: 37403411

Gushudan (GSD) has the effect of strengthening bones and nourishing kidneys. However, its specific intervention mechanism still remains unclear. In this study, to investigate the pathogenesis of glucocorticoid‐induced osteoporosis (GIOP) and the preventive mechanism of GSD on GIOP, fecal metabolomics based on 1H‐NMR and ultra‐high‐performance liquid chromatography‐quadrupole time‐of‐flight‐mass spectrometry method was established. The changes in endogenous metabolites and the relevant metabolic pathways in the control group, model group, and GSD treatment group were investigated via multivariate statistical analysis. As a result, a total of 39 differential metabolites were identified. Of these, 22 metabolites, such as L‐methionine, guanine, and sphingosine, were newly discovered as differential metabolites of GIOP. Amino acid metabolism, energy metabolism, intestinal flora metabolism, and lipid metabolism were significantly changed in the fecal profiles of GIOP rats, and GSD could play an anti‐osteoporosis role by regulating these metabolic pathways. Finally, compared with our previous study of the GSD to prevent kidney yang deficiency syndrome, this study suggested that there were some identical differential metabolites and metabolic pathways. It showed that there was some correlation among the metabolic profiles of the intestine, kidney, and bone in GIOP rats. Therefore, this study offered new insights into the in‐depth understanding of the pathogenesis of GIOP and the intervention mechanism of GSD.

doi: 10.1002/bmc.5404pmid: N/A

Jia, Xinming; Sun, Shilin; Yang, Mengxin; Zhang, Qian; Wang, Nan; Jin, Yiran; Du, Yingfeng

doi: 10.1002/bmc.5682pmid: 37158044

Isodon excisoides (Y.Z.Sun ex C.H.Hu) H. Hara has been often used to treat liver diseases in folk medicine. However, the potential hepatoprotective mechanism of I. excisoides remains unclear. In this study, the mechanism of I. excisoides in alleviating drug‐induced liver injury (DILI) was explored using a strategy combining metabolomics with network pharmacology for the first time. First, serum metabolomics was applied to identify differential metabolites and enrich metabolic pathways. The potential targets of I. excisoides for the treatment of DILI were investigated by network pharmacology. Subsequently, a comprehensive network of network pharmacology and metabolomics was established to find the key genes. Finally, molecular docking technology was used to further verify the key targets. As a result, four key genes including TYMS, IMPDH2, DHODH, and ASAH1 were identified. The proteins produced by these genes had high affinity with the corresponding diterpenoids. These results indicate that the components of I. excisoides play a liver‐protective role by affecting the aforesaid key genes and key proteins. Our results offer a novel strategy for determining the pharmacological effects and potential targets of natural compounds.

Li, Jufang; Chen, Congyan; Sun, Xingchao; Hu, Zhineng; Wu, Chaochao; Gao, Qiang; Ying, Guoqing

doi: 10.1002/bmc.5666pmid: 37139579

A green and inexpensive pretreatment known as dispersive liquid–liquid microextraction (DLLME) was developed in this assay coupled with the LC–MS/MS method for routine analysis of fat soluble vitamins (FSVs). The technique was performed with methanol as the dispersive solvent and dichloromethane as the extraction solvent. The extraction phase containing FSVs was evaporated to dryness and reconstituted in a mixture of acetonitrile and water. The influence variables concerning the DLLME procedure were optimized. After that, the method was investigated for its applicability in LC–MS/MS analysis. As a result, the parameters were settled for the optimal conditions during the DLLME process. A cheap and lipid‐free substance was found as an alternative to serum to eliminate the matrix effect while preparing the calibrators. The method validation indicated that it was suitable for determining FSVs in serum. Moreover, this method was applied successfully to determine serum samples, which was consistent with the literature. In summary, the DLLME method developed in this report was reliable and more cost‐effective than the traditional LC–MS/MS method, and could be applied in the future.

Dai, Bingling; Cao, Hanbing; Hu, Yu; Wang, Xuemei; Huang, Xiaoyue; Su, Xinyue; Ma, Weina; Chen, Yanbin; Peng, Xiujuan; Gong, Zhengyan; Liu, Feng; Zhang, Yanmin

doi: 10.1002/bmc.5692pmid:

Júnior, Gilberto Quirino; Britto‐Júnior, José; Magalhaes, Tainah Babadopulos; Campos, Rafael; Nyamkondiwa, Kudzai L.; Klugh, Kameron L.; Peterson, Larryn W.; Corvino, Angela; Sparaco, Rosa; Frecentese, Francesco; Caliendo, Giuseppe; De Nucci, Gilberto

Menda, Jyothsna; Kanuparthy, Phani Raja; Katari, Naresh Kumar; Kowtharapu, Leela Prasad; Ettaboina, Santhosh Kumar; Pydimarry, Surya Prakash Rao

doi: 10.1002/bmc.5687pmid: 37392152

Ritonavir and darunavir were examined using a ultra‐performance liquid chromatography (UPLC) approach in pharmaceutical dosage forms. The small number of analytical studies that are currently available do not demonstrate the method's stability or nature. The study sought to assess both chemicals using a stability‐indicating approach with a relatively short run time. The HSS C18 (100 × 2.1 mm), 2‐mm column was used for the chromatographic separation, and isocratic elution was used to achieve this. In the mobile phase, methanol and 0.01 M phosphate buffer (pH 4.0) were included in a 60:40 (v/v) ratio. Throughout the analysis, the flow rate was kept at 0.2 mL min−1, and a photodiode array detector set to 266 nm was used to find the major components. The proposed method showed a linear response (r2 > 0.999), and the accuracy was between 98.0% and 102.0%. The precision data showed relative standard deviation ≤1.0%. The UPLC method for quantification of ritonavir and darunavir in pharmaceutical dosage forms using a very short run time of under a minute is the subject of the proposed article. To meet current regulatory criteria, the quality by design idea was used in the method performance verification.

Vadagam, Niroja; Haridasyam, Sharath Babu; Venkatanarayana, Muvvala

doi: 10.1002/bmc.5688pmid: 37325866

Lacosamide (LA) is an antiepileptic medicine that is used to treat tonic–clonic seizures, partial‐onset seizures, mental problems, and pain. A simple, effective, and reliable normal‐phase liquid chromatographic technique was developed and validated to separate and estimate the enantiomer of (S‐enantiomer) LA in pharmaceutical drug substance and drug product. Normal‐phase LC was performed using USP L40 packing material (250 × 4.6 mm, 5 μm) and a mobile phase of n‐hexane and ethanol at 1.0 ml min−1. The detection wavelength, column temperature, and injection volume used were 210 nm, 25°C, and 20 μl, respectively. The enantiomers (LA and S‐enantiomer) were completely separated using a minimum resolution of 5.8 and accurately quantified without any interference in a 25‐min run time. Accuracy study for stereoselective and enantiomeric purity trials was conducted between 10 and 200%, with recovery values ranging from 99.4 to 103.1% and linear regression values >0.997. The stability‐indicating characteristics were assessed using forced degradation tests. The proposed normal‐phase HPLC technique is an alternate approach to the official monograph methods (USP and Ph.Eur.) of LA, and it was successfully used in the evaluation of release and stability samples for both tablet dosage forms and pharmaceutical substances.

Li, Linjie; Tan, Jingxuan; Du, Yujing; Li, Xixuan; Lv, Yongning; Zhai, Xuejia

doi: 10.1002/bmc.5668pmid: 37125701

A sensitive and specific high‐performance liquid chromatography–tandem mass spectrometry method has been developed to determine the pharmacokinetic interactions of the antiplatelet agents aspirin and clopidogrel combined with dl‐3‐n‐butylphthalide. For the determination of aspirin metabolite salicylic acid, clopidogrel inactive metabolite SR26334 and NBP prototype drug in rat plasma, plasma samples were prepared by precipitation of proteins using methanol containing 0.1% formic acid, followed by centrifugation. Chromatography was performed on a C18 column, eluting with a gradient of acetonitrile (with 0.1% formic acid)–water (with 0.1% formic acid). The detection adopted electrospray ion source and positive ion multiple reaction monitoring modes. The linear detection response range of salicylic acid is 80–80,000 ng/ml, and the linear detection response range of SR26334 and dl‐3‐n‐butylphthalide is 10–10,000 ng/ml. Our study revealed that dl‐3‐n‐butylphthalide affected the pharmacokinetics of aspirin and clopidogrel when administered to rats.