Journal of pharmaceutical and biomedical analysis最新文献_第3页

LC-MS peptide mapping of monoclonal antibodies using the mirror proteases trypsin and Tryp-N 使用镜像蛋白酶胰蛋白酶和胰蛋白酶n的单克隆抗体的LC-MS肽定位

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-20 DOI: 10.1016/j.jpba.2026.117361

Domantas Sargautis, Bernd Thiede

Accurate characterization of the amino acid sequence and post translational modifications (PTMs) of monoclonal antibodies (mAbs) is essential for evaluating product quality. Peptide mapping through bottom-up LC/MS analysis is a key methodology for this purpose. While trypsin is commonly the first choice for mAb digestion, it typically yields high but incomplete sequence coverage. As a result, supplementary endoproteases such as Asp-N, chymotrypsin, Glu-C or Lys-C are often employed to enhance coverage. In this report, we evaluated another endoprotease, Tryp-N, which serves as an effective alternative to trypsin for mAb analysis. The sequence coverages achieved for bevacizumab, cetuximab, NISTmAb, and trastuzumab with Tryp-N were comparable to that of trypsin, and the combination of both enzymes slightly improved overall sequence coverage. Notably, both trypsin and Tryp-N generated identical peptides beside the N- and C-terminal ends. The presence of a basic amino acid at opposite ends of the peptide often resulted in complementary sequence coverage of the MS2 of the same peptide sequences. These complementary ion series can be leveraged for precise localization of PTMs, as demonstrated in detail for deamidation, and oxidation sites as well as single amino acid variants (SAVs).

准确鉴定单克隆抗体（mab）的氨基酸序列和翻译后修饰（PTMs）是评价单克隆抗体（mab）产品质量的关键。通过自下而上LC/MS分析的肽图谱是实现这一目的的关键方法。虽然胰蛋白酶通常是单抗消化的首选，但它通常产生高但不完整的序列覆盖率。因此，补充的内源性蛋白酶如Asp-N、凝乳胰蛋白酶、Glu-C或Lys-C通常被用来增强覆盖。在本报告中，我们评估了另一种内源性蛋白酶，胰蛋白酶- n，它可以作为胰蛋白酶的有效替代品进行单抗分析。贝伐单抗、西妥昔单抗、NISTmAb和曲妥珠单抗与胰蛋白酶的序列覆盖率相当，两种酶的组合略微提高了总体序列覆盖率。值得注意的是，胰蛋白酶和胰蛋白酶-N在N端和c端产生相同的肽。在肽的相反端存在碱性氨基酸通常会导致相同肽序列的MS2的互补序列覆盖。这些互补离子系列可以用于精确定位PTMs，如脱酰胺、氧化位点以及单氨基酸变体（sav）的详细演示。

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引用次数: 0

Biopharmaceutical assessment of naloxone permeation through human respiratory epithelial tissues: A chromatographic-mass spectrometric approach with cloud based aerosol dosing and delivery 纳洛酮通过人呼吸道上皮组织渗透的生物制药评估：基于云的气溶胶给药和给药的色谱-质谱方法

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-20 DOI: 10.1016/j.jpba.2026.117366

Tasmin Ara Sultana , Diaa Shakleya , Dustin G. Brown , Patrick J. Faustino , Muhammad Ashraf , Ahmed Zidan

Understanding naloxone permeation is important for optimizing nasal delivery and supporting comparative assessment of nasal drug products. In this study, a stability-indicating LC–MS/MS method was developed and validated for the simultaneous quantification of naloxone and its related impurities, naloxone N-oxide and noroxymorphone, in in vitro permeation test receptor media. The validated method was applied to characterize naloxone permeation following cloud-based aerosol dosing across a synthetic Nuclepore Track-Etched membrane and a differentiated human EpiAirway™ mucociliary tissue model under finite-dose conditions. The analytical procedure demonstrated linearity over 0.25–20.0 ng/mL in Dulbecco’s Phosphate-Buffered Saline and Krebs–Ringer Bicarbonate Buffer, with acceptable accuracy and precision. No degradation products or additional impurities were detected in permeation samples, confirming the stability-indicating capability of the method. Naloxone exhibited rapid early-time permeation across the synthetic membrane, whereas transport across the epithelial tissue model was attenuated and plateaued, reflecting physiological barrier function. Integration of cloud-based aerosol delivery with a validated LC–MS/MS platform enables mechanistic evaluation of nasal naloxone permeation and provides a supportive in vitro framework for formulation characterization and comparative assessments.

了解纳洛酮的渗透对优化鼻腔给药和支持鼻腔药物产品的比较评估是重要的。本研究建立了一种稳定性指示的LC-MS /MS方法，用于同时定量体外渗透试验受体介质中纳洛酮及其相关杂质n -氧化物纳洛酮和去甲氧吗啡酮。在有限剂量条件下，将经过验证的方法应用于基于云的气溶胶给药，通过合成核孔轨道蚀刻膜和分化的人EpiAirway™粘膜纤毛组织模型，来表征纳洛酮的渗透。在Dulbecco的磷酸盐缓冲盐水和Krebs-Ringer碳酸氢盐缓冲液中，分析方法在0.25-20.0 ng/mL范围内呈线性，具有可接受的准确度和精密度。在渗透样品中未检测到降解产物或额外的杂质，证实了该方法的稳定性指示能力。纳洛酮在合成膜上表现出快速的早期渗透，而在上皮组织模型上的转运则减弱并趋于稳定，这反映了生理屏障功能。将基于云的气溶胶给药与经过验证的LC-MS /MS平台相结合，可以对纳洛酮鼻腔渗透进行机理评估，并为制剂表征和比较评估提供一个支持性的体外框架。

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引用次数: 0

Integrated metabolomic and proteomic analysis of cardiac tissues in a murine model of Kawasaki disease 川崎病小鼠模型心脏组织的综合代谢组学和蛋白质组学分析

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-20 DOI: 10.1016/j.jpba.2026.117364

Panpan Liu , Yiyi Shen , Mingyang Zhang , Zhiyuan Liu , Shuhui Wang , Zhiheng Liu , Nana Wang , Jiaying Zhang , Jing Li , Guanghui Qian , Chengyao Ma , Haitao Lv , Ying Liu

Kawasaki disease (KD) is an acute systemic vasculitis in children that can result in severe cardiac complications. This study utilized an integrated proteomic and metabolomic approach to explore molecular dynamics in cardiac tissue from a Candida albicans cell wall extract (CAWS)-induced KD mouse model. Echocardiography demonstrated significant left ventricular dysfunction in the CAWS group, as evidenced by reduced ejection fraction and fractional shortening, alongside histopathological signs of inflammatory cell infiltration. Multi-omics analysis revealed 206 differentially expressed metabolites (DEMs) and 155 differentially expressed proteins (DEPs) compared to PBS controls. Bioinformatics analysis highlighted substantial disturbances in glycerophospholipid metabolism, amino acid metabolism, fatty acid synthesis, and cofactor biosynthesis pathways, with concurrent upregulation of immune- and inflammation-related proteins. Integrated analysis revealed co-enrichment in cofactor biosynthesis, amino acid metabolism, and purine metabolism pathways, and a regulatory network of key molecules was established. These findings suggest that KD-induced cardiac injury involves significant metabolic reprogramming and immune-inflammatory activation, offering new insights into the pathogenesis and providing a theoretical basis for the development of biomarkers and therapeutic targets.

川崎病是一种儿童急性全身性血管炎，可导致严重的心脏并发症。本研究利用综合蛋白质组学和代谢组学方法探索白色念珠菌细胞壁提取物（CAWS）诱导的KD小鼠模型心脏组织的分子动力学。超声心动图显示CAWS组有明显的左心室功能障碍，射血分数降低和分数缩短，同时有炎症细胞浸润的组织病理学征象。多组学分析显示，与PBS对照组相比，有206种差异表达代谢物（DEMs）和155种差异表达蛋白（DEPs）。生物信息学分析强调了甘油磷脂代谢、氨基酸代谢、脂肪酸合成和辅因子生物合成途径的实质性干扰，并伴有免疫和炎症相关蛋白的上调。综合分析发现，在辅助因子生物合成、氨基酸代谢和嘌呤代谢途径中共富集，并建立了关键分子的调控网络。这些发现表明，kd诱导的心脏损伤涉及显著的代谢重编程和免疫炎症激活，为其发病机制提供了新的认识，并为开发生物标志物和治疗靶点提供了理论基础。

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引用次数: 0

Analysis of gut microbiota and intestinal secondary bile acids metabolism in rats after short-term antibiotic treatment 短期抗生素治疗后大鼠肠道菌群及肠道次级胆汁酸代谢分析

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-19 DOI: 10.1016/j.jpba.2026.117365

Zhuan Yang , Xiaoping Zhang , Bo Lv , Yunli Yu

Antibiotics have a profound impact on the overall taxonomic composition of gut microbiota. As gut microbiota influence host bile acids metabolism, changes in the composition of gut microbiota induced by antibiotics are certain to alter host bile acids profile. However, the differences in the effects of various antibiotics on bile acids metabolism and their associations with the impact on intestinal microbiota remain unelucidated. Here, investigation was conducted into how different antibiotics impact host intestinal bile acids metabolism via gut microbiota using in vivo study and multi-omics approaches. Four antibiotics (cefixime, clarithromycin, moxifloxacin and metronidazole) were used to treat normal rats for a short period (3 days), and then gut microbiota, intestinal bile acids profile, BSH and 7α-dehydroxylases activity mediated bile acid metabolism were measured. The results showed that bile acid metabolism in intestine was significantly altered along with the abundance change in bile acid-producing microbiota. It was found that moxifloxacin and metronidazole inhibited the transformation of primary BAs to secondary BAs in the intestine as compared to cefixime and clarithromycin, which was exposed to be associated with its regulation of Ruminococcaceae by Spearman’s correlation analysis (correlation with CDCA/LCA is r = -0.89, FDR<0.001 and CA/DCA is r = -0.708, FDR<0.01). The abundance of Ruminococcaceae in the gut decreased by 87 % after moxifloxacin intervention, while metronidazole completely suppressed the abundance of intestinal Ruminococcaceae. Further analysis revealed that Ruminococcaceae were most strongly associated with the activity of 7α-dehydroxylases (r = 0.701, FDR<0.001). Specifically, we found that Ruminococcus flavefaciens had the closest association with alterations in secondary bile acids (r < -0.68, FDR<0.01). Collectively, our research demonstrated that different antibiotics exerted varying impacts on the production of intestinal secondary bile acids, which was related to their differential effects on the gut microbiota. This work provides novel insights into the interplay between antibiotics and microbial metabolites.

抗生素对肠道微生物群的整体分类组成有深远的影响。由于肠道菌群影响宿主胆汁酸代谢，抗生素引起的肠道菌群组成的变化必然会改变宿主胆汁酸谱。然而，各种抗生素对胆汁酸代谢的影响差异及其与肠道微生物群影响的关联尚不清楚。本研究采用体内研究和多组学方法研究了不同抗生素如何通过肠道微生物群影响宿主肠道胆汁酸代谢。采用头孢克肟、克拉霉素、莫西沙星、甲硝唑4种抗生素短时间（3 d）治疗正常大鼠，测定其肠道菌群、肠道胆汁酸谱、BSH和7α-去羟化酶活性介导的胆汁酸代谢。结果表明，肠道内胆汁酸代谢随着产胆汁酸微生物群丰度的变化而显著改变。与头孢克肟和克拉霉素相比，莫西沙星和甲硝唑抑制了肠道内初级BAs向次级BAs的转化，并通过Spearman相关分析发现其对瘤胃球菌科的调节作用与其相关（与CDCA/LCA的相关性为r = -0.89,FDR<0.001， CA/DCA的相关性为r = -0.708,FDR<0.01）。莫西沙星干预后，肠道Ruminococcaceae丰度下降了87 %，而甲硝唑完全抑制了肠道Ruminococcaceae的丰度。进一步分析发现，Ruminococcaceae与7α-去羟化酶活性相关性最强（r = 0.701,FDR<0.001）。具体来说，我们发现黄瘤球菌与次级胆汁酸的改变密切相关（r <； -0.68,FDR<0.01）。总的来说，我们的研究表明，不同的抗生素对肠道次级胆汁酸的产生有不同的影响，这与它们对肠道微生物群的不同影响有关。这项工作为抗生素和微生物代谢物之间的相互作用提供了新的见解。

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引用次数: 0

Identification and characterization of trametinib degradation product employing Orbitrap LC-HRMS, and development of a robust, eco-friendly stability indicating method of analysis 利用Orbitrap LC-HRMS对曲美替尼降解产物进行鉴定和表征，并开发了一种稳健、环保的稳定性指示分析方法。

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-17 DOI: 10.1016/j.jpba.2026.117355

Sohan G. Jawarkar , Gayatri Amliyar , Nasir Khan, Pooja Dhakne, Megha Pillai, Shankha Dey, Jinal Ajabiya, Pinaki Sengupta

Trametinib is a selective MEK1/2 inhibitor approved for the treatment of various cancers. Stress testing of a drug is essential for understanding the stability profile and ensuring compliance with regulatory requirements during pharmaceutical development. To the best of our knowledge, only a few reports on stress testing of trametinib are available to date. Moreover, comprehensive reports on the stress testing of trametinib using LC-HRMS are not available in any literature. Therefore, the present study aimed to conduct stress studies on trametinib in accordance with ICH guidelines. The RP-HPLC stability-indicating analytical method was developed, and the stress degradation condition for trametinib was optimised. Chromatographic separation of trametinib and its degradation products was achieved on a Phenomenex Luna C18 column (250 × 4.6 mm, 5 µm) using a mobile phase consisting of 0.1 % formic acid in Milli-Q water and acetonitrile using gradient elution. The proposed method demonstrated excellent specificity, linearity, precision (RSD ≤ 0.09 %), and accuracy (99.78–101.38 %). Results indicate that trametinib exhibited high susceptibility to acidic and basic stress conditions, whereas it remained relatively stable under oxidative and thermal stress conditions. All degradation products were characterised using Orbitarp LC-HRMS in positive HESI mode (m/z 50–800 Da) for accurate mass determination and fragmentation pathway elucidation. In-silico toxicity prediction suggested that among the degradation products, neurotoxicity was the most consistently predicted endpoint with high confidence (≥70 %), followed by respiratory toxicity and immunotoxicity. Greenness evaluation using AGREE, Eco-Scale, and BAGI tools indicated a moderate environmental impact.

曲美替尼是一种选择性MEK1/2抑制剂，被批准用于治疗各种癌症。在药物开发过程中，药物压力测试对于了解药物的稳定性和确保符合法规要求至关重要。据我们所知，到目前为止，关于曲美替尼压力测试的报道很少。此外，在文献中没有使用LC-HRMS对曲美替尼进行压力测试的全面报道。因此，本研究旨在根据ICH指南对曲美替尼进行应激研究。建立了反相高效液相色谱稳定性指示分析方法，优化了曲美替尼的应力降解条件。采用Phenomenex Luna C18色谱柱（250 × 4.6 mm, 5 µm）对曲美替尼及其降解产物进行色谱分离，流动相为0.1 %甲酸- milliq水和乙腈，采用梯度洗脱。该方法具有良好的特异性、线性度、精密度（RSD≤0.09 %）和准确度（99.78 ~ 101.38 %）。结果表明，曲美替尼对酸性和碱性胁迫条件表现出较高的敏感性，而在氧化和热胁迫条件下保持相对稳定。所有降解产物均使用Orbitarp LC-HRMS在正HESI模式下（m/z 50-800 Da）进行表征，以准确测定质量和裂解途径。硅毒性预测表明，在降解产物中，神经毒性是最一致的预测终点，置信度高（≥70 %），其次是呼吸毒性和免疫毒性。使用AGREE、Eco-Scale和BAGI工具进行的绿色评价表明环境影响中等。

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引用次数: 0

Investigation of the stability profile of therapeutic α-MSH analogue: Insights from liquid chromatography-high resolution mass spectrometry analysis of afamelanotide 治疗性α-MSH类似物的稳定性研究：液相色谱-高分辨质谱分析阿麦兰肽的见解

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-14 DOI: 10.1016/j.jpba.2026.117362

Ashwini Chawathe , Nitish Sharma

Afamelanotide, also known as melanotan-1, is a synthetic 13-amino acid peptidomimetic of α-melanocyte stimulating hormone (α-MSH), and is a critical peptide orphan drug used for the management of erythropoietic protoporphyria. It contains norleucine and D-phenylalanine at positions 4 and 7, in place of methionine and L-phenylalanine, respectively as found in endogenous peptide. Therapeutic peptide stability profiling is crucial in drug development because chemical and physical degradation during storage alters structural properties, potentially reducing efficacy and compromising safety by preventing target engagement. Stability testing for synthetic peptides is performed by following the International Council for Harmonisation (ICH) guidelines Q1A(R2) and Q5C. The current work endeavours to explore afamelanotide’s degradation pathways under various chemical and physical stress conditions: acidic, basic, neutral, and oxidative stress, UV light exposure, and increased temperature at 60⁰C. The study demonstrated that afamelanotide undergoes degradation under all applied stress conditions with the generation of fourteen different degradation products (DPs) which were separated by gradient reversed-phase HPLC on a Zorbax SB C18 column (300 Å, 4.6*150 mm, 3.5 µm) and the method was validated according to the ICH Q2(R1) guideline. To enable comprehensive characterization, the analysis was coupled with ultra-high-performance liquid chromatography-high resolution tandem mass spectrometry (UHPLC-HRMS/MS), where collision-induced dissociation yielded abundant and accurate fragmentation patterns, enabling the detailed structural elucidation of the products. While this work has identified several degradation pathways such as truncation, methylation, deacetylation, and oxidation, it also establishes complete stability profile of α-MSH analogue, thus offering key insights for the rational design of robust drug formulations.

Afamelanotide，又称melanotan-1，是一种合成的具有13个氨基酸的α-促黑素细胞激素（α-MSH）类肽，是一种用于治疗红细胞原卟啉症的关键肽孤儿药。在4和7位分别含有去甲亮氨酸和d -苯丙氨酸，代替内源性肽中的蛋氨酸和l -苯丙氨酸。治疗性肽稳定性分析在药物开发中至关重要，因为储存过程中的化学和物理降解会改变结构特性，可能会降低疗效，并通过阻止靶标接触而损害安全性。合成肽的稳定性测试遵循国际统一委员会（ICH）指南Q1A（R2）和Q5C进行。目前的工作努力探索afamelanotide在各种化学和物理应激条件下的降解途径：酸性、碱性、中性和氧化应激、紫外线照射和60⁰C温度升高。研究表明，在所有施加的应力条件下，afamelanotide都能被降解，产生14种不同的降解产物（dp），在Zorbax SB C18色谱柱（300 Å， 4.6*150 mm, 3.5 µm）上用梯度反相高效液相色谱分离，并根据ICH Q2（R1）指南对该方法进行验证。为了进行全面的表征，分析结合了超高效液相色谱-高分辨率串联质谱（UHPLC-HRMS/MS），其中碰撞诱导解离产生了丰富而准确的碎片模式，从而能够详细阐明产品的结构。虽然这项工作已经确定了几种降解途径，如截断、甲基化、去乙酰化和氧化，但它也建立了α-MSH类似物的完整稳定性谱，从而为合理设计稳健的药物配方提供了关键见解。

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引用次数: 0

Determination of fifteen compounds in rat plasma by UHPLC-QTRAP-MS/MS for pharmacokinetic study after oral administration of Euonymi herba extract UHPLC-QTRAP-MS/MS法测定大鼠口服卫矛提取物后血浆中15种化合物的药代动力学研究

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-14 DOI: 10.1016/j.jpba.2026.117358

Fanjiao Zuo , Peng Zhao , Xueyu Liu, Huining Geng, Lu Chen, Jinyue Ma, Yameng Zhu, Zhenguo Lv, Ye Shang, Huizi Ouyang, Jun He

Euonymi herba (EH) was one of the traditional characteristic national medicines of Guangxi, which had various pharmacological activities such as hemostasis, anti-myocardial hypoxia, and anti-aging. This study established a reliable ultra-high performance liquid chromatography-quadrupole linear ion trap tandem mass spectrometry (UHPLC-QTRAP-MS/MS) method for the simultaneous quantification of multiple components in rat plasma following oral administration of EH extract. The method demonstrated satisfactory specificity, excellent linearity (r ≥ 0.9938), acceptable precision (RSD ≤ 10.34 %), accuracy (ranging from −9.73–9.00 %), recovery (between 65.95 % and 111.75 %), matrix effect (between 60.49 % and 116.11 %) and stability (RSD ≤ 8.36 %), with all key parameters meeting acceptance criteria for bioanalytical method validation. The pharmacokinetic study yielded the profiles and parameters for ten components, revealing dulcitol as the compound with the highest exposure and fastest absorption. This work provides critical data for elucidating the pharmacologically active constituents of EH.

卫矛是广西传统特色国药之一，具有止血、抗心肌缺氧、抗衰老等药理作用。本研究建立了一种可靠的超高效液相色谱-四极杆线性离子阱串联质谱（UHPLC-QTRAP-MS/MS）同时定量大鼠口服EH提取物后血浆中多种成分的方法。方法演示了满意的特异性,出色的线性度(r ≥0.9938 ),可接受的精度(RSD为≤10.34 %),精度(从9.73−-9.00 %),恢复  % 65.95和111.75之间(%),基体效应  % 60.49和116.11之间(%)和稳定性(RSD为≤8.36 %),与所有关键参数满足验收标准为生物分析方法验证。药动学研究得到了10种成分的药动学特征和参数，结果表明dulcitol是暴露量最大、吸收速度最快的化合物。这项工作为阐明EH的药理活性成分提供了重要的数据。

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引用次数: 0

Identification of leprosy reactions using Fourier transform infrared (FTIR) spectroscopy supervised by clinical evaluation 用傅立叶变换红外光谱识别麻风病反应，并进行临床评价

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-12 DOI: 10.1016/j.jpba.2026.117354

Paulo Cezar de Moraes , Alessandra Koehler , Letícia Maria Eidt , Cristiane Almeida Soares Cattani , Valeriano Antonio Corbellini , Maria Lúcia Scroferneker

Leprosy is a chronic granulomatous, infectious and disabling disease, whose etiological agents are Mycobacterium leprae and M. lepromatosis. Leprosy reactions are the main causes of peripheral nerve damage and sequelae of the disease. There is no laboratory test that alone allows the identification of these episodes, causing delays in their diagnosis and treatment. The objective of this study was to propose a methodology based on Fourier transform infrared (FTIR) spectroscopy to detect leprosy reactions in the saliva of leprosy patients, supervised by the clinical diagnosis of these episodes. A total of 131 saliva samples were included and analyzed by attenuated total reflection (ATR)-FTIR; a supervised analysis with partial least squares discriminant analysis (PLS-DA), after orthogonal signal correction (OSC), was used to classify the samples into two groups: with and without leprosy reactions. The PLS-DA model with one latent variable and one OSC component showed 100 % sensitivity and specificity both in the calibration and prediction sets. Thus, all samples were correctly classified, allowing the diagnosis of leprosy reactions in saliva with high accuracy. Therefore, the FTIR-based chemometric model proved promising for the rapid and early identification of these episodes, contributing to clinical management of patients with leprosy.

麻风病是一种慢性肉芽肿性、传染性和致残性疾病，其病原是麻风分枝杆菌和麻风分枝杆菌病。麻风病反应是周围神经损伤和该病后遗症的主要原因。没有单独的实验室测试可以识别这些发作，导致其诊断和治疗的延误。本研究的目的是提出一种基于傅立叶变换红外（FTIR）光谱的方法来检测麻风患者唾液中的麻风反应，并以这些事件的临床诊断为指导。采用衰减全反射(ATR)-FTIR分析131份唾液样本；经正交信号校正（OSC）后，采用偏最小二乘判别分析（PLS-DA）的监督分析将样本分为两组：有麻风反应和无麻风反应。具有一个潜在变量和一个盐含量成分的PLS-DA模型在校准集和预测集上的灵敏度和特异性均为100 %。因此，所有样本都被正确分类，使得唾液中麻风病反应的诊断具有很高的准确性。因此，基于ftir的化学计量模型有望快速和早期识别这些事件，有助于麻风患者的临床管理。

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引用次数: 0

Unraveling excretion behavior of polyethylene glycol 1000 with 13–26 subunits in rats by UHPLC-MS/MS UHPLC-MS/MS揭示聚乙二醇1000 13-26亚基在大鼠体内的排泄行为

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-12 DOI: 10.1016/j.jpba.2026.117353

Xiaoyan Zhang , Shuang Feng , Ziluo Zhang , Chuya Wang , Yongshan Ai , Yalin Xi , Lei Yin , Meiyun Shi

This study established a robust and selective ultra-high-performance liquid chromatography–tandem mass spectrometry (UHPLC-MS/MS) method to investigate the excretion behavior of polyethylene glycol 1000 (PEG1K) oligomers (N = 13–26) in rats. PEG is widely used in pharmaceutical applications, particularly in drug delivery systems, yet detailed excretion profiles of lower molecular weight PEGs remain insufficiently characterized. The developed method utilized ammonium adducts ([M+NH₄]⁺) as precursor ions in multiple reaction monitoring mode, enabling sensitive and high-throughput quantification of individual oligomers in biological matrices. Following intravenous administration in rats, excretion kinetics were monitored over 48 h. Cumulative recovery analysis revealed that renal excretion was the dominant elimination pathway, accounting for 47.30–68.52 % of the administered dose in urine, while fecal excretion represented only 0.66–3.38 %. These findings provide critical insights into the in vivo disposition of PEG1K, supporting its safety evaluation and rational application in drug formulation. The presented analytical platform offers a reliable tool for polymer pharmacokinetic studies.

本研究建立了一种稳健、选择性的超高效液相色谱-串联质谱（UHPLC-MS/MS）方法来研究聚乙二醇1000 （PEG1K）低聚物（N = 13-26）在大鼠体内的排泄行为。聚乙二醇广泛用于制药应用，特别是在药物输送系统中，但低分子量聚乙二醇的详细排泄谱仍然没有充分表征。该方法利用铵盐加合物（[M+NH₄]+）作为前体离子，采用多反应监测模式，实现了对生物基质中单个低聚物的灵敏、高通量定量。在大鼠静脉给药后，在48 h内监测排泄动力学。累积恢复分析显示，肾脏排泄是主要的消除途径，占尿液给药剂量的47.30-68.52 %，而粪便排泄仅占0.66-3.38 %。这些发现为PEG1K在体内的分布提供了重要的见解，支持其安全性评估和在药物配方中的合理应用。该分析平台为聚合物药代动力学研究提供了可靠的工具。

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引用次数: 0

Development of a simple labeling method using fluorescent protein fusion proteins targeting the membrane lipids of small extracellular vesicles 利用荧光蛋白融合蛋白靶向小细胞外囊泡膜脂的简单标记方法的发展

IF 3.1 3区医学 Q2 CHEMISTRY, ANALYTICAL

Journal of pharmaceutical and biomedical analysis

Pub Date : 2026-01-12 DOI: 10.1016/j.jpba.2026.117356

Yuki Kobayashi, Yuki Takahashi, Hiroto Otera, Yuriko Higuchi, Yoshinobu Takakura

Extracellular vesicles (EV) are lipid-based nanoparticles naturally released by cells, exhibiting considerable heterogeneity in size, surface charge, and biomolecular composition. Recently, increasing attention has been directed toward the characterization of distinct EV-subpopulations, particularly based on unique surface antigen expression profiles. Therefore, a method for analyzing EV-subpopulations using versatile equipment would be highly valuable. In this study, we developed a labeling method for analyzing specific populations in small EVs (sEVs) distinguished by their levels phosphatidylserine (PS) exposure. For visualization, sEVs were labeled with two fusion proteins (enhanced green fluorescent protein [EGFP] linked to lactadherin [LA] and mCherry-Vn96) comprising a PS-binding protein or sEV-tropic peptide (Vn96) combined with fluorescent proteins. Using ultracentrifugation, bulk sEVs were collected, and a fraction of PS⁽⁻⁾ sEVs (PS⁽⁺⁾ sEV-depleted fraction) were isolated by depleting PS⁽⁺⁾ sEVs from bulk sEVs. In bulk sEVs, the colocalization of EGFP-LA and mCherry-Vn96-derived signals was detected. In contrast, the PS⁽⁺⁾ sEV-depleted fraction exhibited reduced EGFP-LA fluorescence signal, with only mCherry-Vn96 fluorescence remaining detectable. In conclusion, our labeling technique facilitates the identification and analysis of sEV-subpopulations using fluorescence microscopy in small sample volumes. This platform can also be adapted for broader applications by incorporating additional protein markers.

细胞外囊泡（EV）是由细胞自然释放的脂质纳米颗粒，在大小、表面电荷和生物分子组成上表现出相当大的异质性。最近，人们越来越关注不同ev亚群的特征，特别是基于独特的表面抗原表达谱。因此，一种使用多功能设备分析ev亚群的方法将是非常有价值的。在这项研究中，我们开发了一种标记方法，用于分析小型电动汽车（sev）中由磷脂酰丝氨酸（PS）暴露水平区分的特定群体。为了可视化，sev用两种融合蛋白（与乳酸粘附素（LA）连接的增强型绿色荧光蛋白[EGFP]和mCherry-Vn96）标记，其中包括ps结合蛋白或sEV-tropic peptide （Vn96）与荧光蛋白结合。利用超离心的方法，收集大量的sev，并通过从大量sev中耗尽PS(+) sev分离出一部分PS(−)sev （PS(+) sev）。在批量sev中，检测到EGFP-LA和mcherry - vn96衍生信号的共定位。相比之下，PS(+) sev缺失部分的EGFP-LA荧光信号减弱，仅残留mCherry-Vn96荧光。总之，我们的标记技术有助于荧光显微镜在小样本量下对sev亚群进行鉴定和分析。该平台还可以通过加入额外的蛋白质标记物来适应更广泛的应用。

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