Journal of Pharmaceutical Analysis

2023, 13(2): ii-ii. doi: 10.1016/S2095-1779(23)00009-6

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Microneedle-based interstitial fluid extraction for drug analysis: Advances, challenges, and prospects

Shuwen Ma, Jiaqi Li, Lixia Pei, Nianping Feng, Yongtai Zhang

2023, 13(2): 111-126. doi: 10.1016/j.jpha.2022.12.004

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Similar to blood, interstitial fluid (ISF) contains exogenous drugs and biomarkers and may therefore substitute blood in drug analysis. However, current ISF extraction techniques require bulky instruments and are both time-consuming and complicated, which has inspired the development of viable alternatives such as those relying on skin or tissue puncturing with microneedles. Currently, microneedles are widely employed for transdermal drug delivery and have been successfully used for ISF extraction by different mechanisms to facilitate subsequent analysis. The integration of microneedles with sensors enables in situ ISF analysis and specific compound monitoring, while the integration of monitoring and delivery functions in wearable devices allows real-time dose modification. Herein, we review the progress in drug analysis based on microneedle-assisted ISF extraction and discuss the related future opportunities and challenges.

Novel insights into histone lysine methyltransferases in cancer therapy: From epigenetic regulation to selective drugs

Qili Liao, Jie Yang, Shengfang Ge, Peiwei Chai, Jiayan Fan, Renbing Jia

2023, 13(2): 127-141. doi: 10.1016/j.jpha.2022.11.009

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The reversible and precise temporal and spatial regulation of histone lysine methyltransferases (KMTs) is essential for epigenome homeostasis. The dysregulation of KMTs is associated with tumor initiation, metastasis, chemoresistance, invasiveness, and the immune microenvironment. Therapeutically, their promising effects are being evaluated in diversified preclinical and clinical trials, demonstrating encouraging outcomes in multiple malignancies. In this review, we have updated recent understandings of KMTs' functions and the development of their targeted inhibitors. First, we provide an updated overview of the regulatory roles of several KMT activities in oncogenesis, tumor suppression, and immune regulation. In addition, we summarize the current targeting strategies in different cancer types and multiple ongoing clinical trials of combination therapies with KMT inhibitors. In summary, we endeavor to depict the regulation of KMT-mediated epigenetic landscape and provide potential epigenetic targets in the treatment of cancers.

Progress and prediction of multicomponent quantification in complex systems with practical LC-UV methods

Xi Chen, Zhao Yang, Yang Xu, Zhe Liu, Yanfang Liu, Yuntao Dai, Shilin Chen

2023, 13(2): 142-155. doi: 10.1016/j.jpha.2022.11.011

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Complex systems exist widely, including medicines from natural products, functional foods, and biological samples. The biological activity of complex systems is often the result of the synergistic effect of multiple components. In the quality evaluation of complex samples, multicomponent quantitative analysis (MCQA) is usually needed. To overcome the difficulty in obtaining standard products, scholars have proposed achieving MCQA through the “single standard to determine multiple components (SSDMC)” approach. This method has been used in the determination of multicomponent content in natural source drugs and the analysis of impurities in chemical drugs and has been included in the Chinese Pharmacopoeia. Depending on a convenient (ultra) high-performance liquid chromatography method, how can the repeatability and robustness of the MCQA method be improved? How can the chromatography conditions be optimized to improve the number of quantitative components? How can computer software technology be introduced to improve the efficiency of multicomponent analysis (MCA)? These are the key problems that remain to be solved in practical MCQA. First, this review article summarizes the calculation methods of relative correction factors in the SSDMC approach in the past five years, as well as the method robustness and accuracy evaluation. Second, it also summarizes methods to improve peak capacity and quantitative accuracy in MCA, including column selection and two-dimensional chromatographic analysis technology. Finally, computer software technologies for predicting chromatographic conditions and analytical parameters are introduced, which provides an idea for intelligent method development in MCA. This paper aims to provide methodological ideas for the improvement of complex system analysis, especially MCQA.

Celastrol targeting Nedd4 reduces Nrf2-mediated oxidative stress in astrocytes after ischemic stroke

Zexuan Hong, Jun Cao, Dandan Liu, Maozhu Liu, Mengyuan Chen, Fanning Zeng, Zaisheng Qin, Jigang Wang, Tao Tao

2023, 13(2): 156-169. doi: 10.1016/j.jpha.2022.12.002

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Stroke is the second leading cause of death worldwide, and oxidative stress plays a crucial role. Celastrol exhibits strong antioxidant properties in several diseases; however, whether it can affect oxidation in cerebral ischemic-reperfusion injury (CIRI) remains unclear. This study aimed to determine whether celastrol could reduce oxidative damage during CIRI and to elucidate the underlying mechanisms. Here, we found that celastrol attenuated oxidative injury in CIRI by upregulating nuclear factor E2-related factor 2 (Nrf2). Using alkynyl-tagged celastrol and liquid chromatography-tandem mass spectrometry, we showed that celastrol directly bound to neuronally expressed developmentally downregulated 4 (Nedd4) and then released Nrf2 from Nedd4 in astrocytes. Nedd4 promoted the degradation of Nrf2 through K48-linked ubiquitination and thus contributed to astrocytic reactive oxygen species production in CIRI, which was significantly blocked by celastrol. Furthermore, by inhibiting oxidative stress and astrocyte activation, celastrol effectively rescued neurons from axon damage and apoptosis. Our study uncovered Nedd4 as a direct target of celastrol, and that celastrol exerts an antioxidative effect on astrocytes by inhibiting the interaction between Nedd4 and Nrf2 and reducing Nrf2 degradation in CIRI.

SUMO1 regulates post-infarct cardiac repair based on cellular heterogeneity

Zhihao Liu, Xiaozhi Liu, Li Liu, Ying Wang, Jie Zheng, Lan Li, Sheng Li, Han Zhang, Jingyu Ni, Chuanrui Ma, Xiumei Gao, Xiyun Bian, Guanwei Fan

2023, 13(2): 170-186. doi: 10.1016/j.jpha.2022.11.010

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Small ubiquitin-related modifier (SUMOylation) is a dynamic post-translational modification that maintains cardiac function and can protect against a hypertrophic response to cardiac pressure overload. However, the function of SUMOylation after myocardial infarction (MI) and the molecular details of heart cell responses to SUMO1 deficiency have not been determined. In this study, we demonstrated that SUMO1 protein was inconsistently abundant in different cell types and heart regions after MI. However, SUMO1 knockout significantly exacerbated systolic dysfunction and infarct size after myocardial injury. Single-nucleus RNA sequencing revealed the differential role of SUMO1 in regulating heart cells. Among cardiomyocytes, SUMO1 deletion increased the Nppa⁺ Nppb⁺ Ankrd1⁺ cardiomyocyte subcluster proportion after MI. In addition, the conversion of fibroblasts to myofibroblasts subclusters was inhibited in SUMO1 knockout mice. Importantly, SUMO1 loss promoted proliferation of endothelial cell subsets with the ability to reconstitute neovascularization and expressed angiogenesis-related genes. Computational analysis of ligand/receptor interactions suggested putative pathways that mediate cardiomyocytes to endothelial cell communication in the myocardium. Mice preinjected with cardiomyocyte-specific AAV-SUMO1, but not the endothelial cell-specific form, and exhibited ameliorated cardiac remodeling following MI. Collectively, our results identified the role of SUMO1 in cardiomyocytes, fibroblasts, and endothelial cells after MI. These findings provide new insights into SUMO1 involvement in the pathogenesis of MI and reveal novel therapeutic targets.

PKM2-mediated neuronal hyperglycolysis enhances the risk of Parkinson's disease in diabetic rats

Ya Zhao, Yanwei Wang, Yuying Wu, Cimin Tao, Rui Xu, Yong Chen, Linghui Qian, Tengfei Xu, Xiaoyuan Lian

2023, 13(2): 187-200. doi: 10.1016/j.jpha.2022.11.006

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Epidemiological and animal studies indicate that pre-existing diabetes increases the risk of Parkinson's disease (PD). However, the mechanisms underlying this association remain unclear. In the present study, we found that high glucose (HG) levels in the cerebrospinal fluid (CSF) of diabetic rats might enhance the effect of a subthreshold dose of the neurotoxin 6-hydroxydopamine (6-OHDA) on the development of motor disorders, and the damage to the nigrostriatal dopaminergic neuronal pathway. In vitro, HG promoted the 6-OHDA-induced apoptosis in PC12 cells differentiated to neurons with nerve growth factor (NGF) (NGF-PC12). Metabolomics showed that HG promoted hyperglycolysis in neurons and impaired tricarboxylic acid cycle (TCA cycle) activity, which was closely related to abnormal mitochondrial fusion, thus resulting in mitochondrial loss. Interestingly, HG-induced upregulation of pyruvate kinase M2 (PKM2) combined with 6-OHDA exposure not only mediated glycolysis but also promoted abnormal mitochondrial fusion by upregulating the expression of MFN2 in NGF-PC12 cells. In addition, we found that PKM2 knockdown rescued the abnormal mitochondrial fusion and cell apoptosis induced by HG+6-OHDA. Furthermore, we found that shikonin (SK), an inhibitor of PKM2, restored the mitochondrial number, promoted TCA cycle activity, reversed hyperglycolysis, enhanced the tolerance of cultured neurons to 6-OHDA, and reduced the risk of PD in diabetic rats. Overall, our results indicate that diabetes promotes hyperglycolysis and abnormal mitochondrial fusion in neurons through the upregulation of PKM2, leading to an increase in the vulnerability of dopaminergic neurons to 6-OHDA. Thus, the inhibition of PKM2 and restoration of mitochondrial metabolic homeostasis/pathways may prevent the occurrence and development of diabetic PD.

Quantitative estimation of enzymatic released specific oligosaccharides from Hericium erinaceus polysaccharides using CE-LIF

Yong Deng, Jing Zhao, Shaoping Li

2023, 13(2): 201-208. doi: 10.1016/j.jpha.2022.11.004

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Polysaccharides exhibit multiple pharmacological activities which are closely related to their structural features. Therefore, quantitatively quality control of polysaccharides based on their chemical characteristics is important for their application in biomedical and functional food sciences. However, polysaccharides are mixed macromolecular compounds that are difficult to isolate and lack standards, making them challenging to quantify directly. In this study, we proposed an improved saccharide mapping method based on the release of specific oligosaccharides for the assessment of Hericium erinaceus polysaccharides from laboratory cultured and different regions of China. Briefly, a polysaccharide from H. erinaceus was digested by β-(1-3)-glucanase, and the released specific oligosaccharides were labeled with 8-aminopyrene-1,3,6-trisulfonic-acid (APTS) and separated by using micellar electrokinetic chromatography (MEKC) coupled with laser induced fluorescence (LIF), and quantitatively estimated. MEKC presented higher resolution compared to polysaccharide analysis using carbohydrate gel electrophoresis (PACE), and provided great peak capacity between oligosaccharides with polymerization degree of 2 (DP2) and polymerization degree of 6 (DP6) in a dextran ladder separation. The results of high performance size exclusion chromatography coupled with multi-angle laser light scattering and refractive index detector (HPSEC-MALLS-RI) showed that 12 h was sufficient for complete digestion of polysaccharides from H. erinaceus. Laminaritriose (DP3) was used as an internal standard for quantification of all the oligosaccharides. The calibration curve for DP3 showed a good linear regression (R² > 0.9988). The limit of detection (LOD) and limit of quantification (LOQ) values were 0.05 μg/mL and 0.2 μg/mL, respectively. The recovery for DP3 was 87.32 (±0.03)% in the three independent injections. To sum up, this proposed method is helpful for improving the quality control of polysaccharides from H. erinaceus as well as other materials.

A benzenesulfonic acid-modified organic polymer monolithic column with reversed-phase/hydrophilic bifunctional selectivity for capillary electrochromatography

Yikun Liu, Ning He, Yingfang Lu, Weiqiang Li, Xin He, Zhentao Li, Zilin Chen

2023, 13(2): 209-215. doi: 10.1016/j.jpha.2022.10.006

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Here, a styrene-based polymer monolithic column poly(VBS-co-TAT-co-AHM) with reversed-phase/hydrophilic interaction liquid chromatography (RPLC/HILIC) bifunctional separation mode was successfully prepared for capillary electrochromatography by the in situ polymerization of sodium p-styrene sulfonate (VBS) with cross-linkers 3-(acryloyloxy)-2-hydroxypropyl methacrylate (AHM) and 1,3,5-triacryloylhexahydro-1,3,5-triazine (TAT). The preparation conditions of the monolith were optimized. The morphology and formation of the poly(VBS-co-TAT-co-AHM) monolith were confirmed by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FT-IR). The separation performances of the monolith were evaluated systematically. It should be noted that the incorporation of VBS functional monomer can provide π-π interactions, hydrophilic interactions, and ion-exchange interactions. Hence, the prepared poly(VBS-co-TAT-co-AHM) monolith can achieve efficient separation of thiourea compounds, benzene series, phenol compounds, aniline compounds and sulfonamides in RPLC or HILIC separation mode. The largest theoretical plate number for N,N'-dimethylthiourea reached 1.7×10⁵ plates/m. In addition, the poly(VBS-co-TAT-co-AHM) monolithic column showed excellent reproducibility and stability. This novel monolithic column has great application value and potential in capillary electrochromatography (CEC).

Comparison of different approaches for direct coupling of solid-phase microextraction to mass spectrometry for drugs of abuse analysis in plasma

Wei Zhou, Martyna N. Wieczorek, Runshan Will Jiang, Janusz Pawliszyn

2023, 13(2): 216-222. doi: 10.1016/j.jpha.2022.10.004

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Abstract:
The direct coupling of solid-phase microextraction (SPME) to mass spectrometry (MS) (SPME-MS) has proven to be an effective method for the fast screening and quantitative analysis of compounds in complex matrices such as blood and plasma. In recent years, our lab has developed three novel SPME-MS techniques: SPME-microfluidic open interface-MS (SPME-MOI-MS), coated blade spray-MS (CBS-MS), and SPME-probe electrospray ionization-MS (SPME-PESI-MS). The fast and high-throughput nature of these SPME-MS technologies makes them attractive options for point-of-care analysis and anti-doping testing. However, all these three techniques utilize different SPME geometries and were tested with different MS instruments. Lack of comparative data makes it difficult to determine which of these methodologies is the best option for any given application. This work fills this gap by making a comprehensive comparison of these three technologies with different SPME devices including SPME fibers, CBS blades, and SPME-PESI probes and SPME-liquid chromatography-MS (SPME-LC-MS) for the analysis of drugs of abuse using the same MS instrument. Furthermore, for the first time, we developed different desorption chambers for MOI-MS for coupling with SPME fibers, CBS blades, and SPME-PESI probes, thus illustrating the universality of this approach. In total, eight analytical methods were developed, with the experimental data showing that all the SPME-based methods provided good analytical performance with R² of linearities larger than 0.9925, accuracies between 81% and 118%, and good precision with an RSD%≤13%.

2023 Vol. 13, No. 2