For the resolution of isomer-rich lipid extracts, it is common practice during RPLC method development to evaluate column efficiency and mobile phase composition. However, further gains can be made by tuning other parameters, such as the microstructure of the packing material. Today, lipid separations are mostly performed with columns packed with particles having narrow pore sizes (80–120 Å). This is unsurprising, given the expected hydrodynamic radius of lipids up to of 23 Å. However, specific investigations of pore size influence on lipid separations are scarce. Therefore, a systematic comparison of 150 mm columns packed with C18 particles having pore sizes of 130 and 300 Å was performed with respect to selectivity, peak capacity and chromatographic resolution. An RPLC/HRMS-based lipidomic analysis was developed on 150 mm columns. Similar loadability, selectivity, sensitivity and average peak capacity were demonstrated by 300 Å column compared to its narrow-pore counterpart. Overall, current experimental results indicate that wide-pore columns are equally applicable for RPLC/HRMS-based discovery lipidomics and may be even beneficial for the analysis of lipids with high structural complexity. In addition, these columns appear to be better suitable for high-throughput analysis due to lower lipid retention via smaller surface area.
{"title":"Applicability of wide-pore columns in reversed-phase liquid chromatography for human blood lipid profiling","authors":"Sergey Girel , Davy Guillarme , Serge Rudaz , Isabel Meister","doi":"10.1016/j.jcoa.2025.100295","DOIUrl":"10.1016/j.jcoa.2025.100295","url":null,"abstract":"<div><div>For the resolution of isomer-rich lipid extracts, it is common practice during RPLC method development to evaluate column efficiency and mobile phase composition. However, further gains can be made by tuning other parameters, such as the microstructure of the packing material. Today, lipid separations are mostly performed with columns packed with particles having narrow pore sizes (80–120 Å). This is unsurprising, given the expected hydrodynamic radius of lipids up to of 23 Å. However, specific investigations of pore size influence on lipid separations are scarce. Therefore, a systematic comparison of 150 mm columns packed with C<sub>18</sub> particles having pore sizes of 130 and 300 Å was performed with respect to selectivity, peak capacity and chromatographic resolution. An RPLC/HRMS-based lipidomic analysis was developed on 150 mm columns. Similar loadability, selectivity, sensitivity and average peak capacity were demonstrated by 300 Å column compared to its narrow-pore counterpart. Overall, current experimental results indicate that wide-pore columns are equally applicable for RPLC/HRMS-based discovery lipidomics and may be even beneficial for the analysis of lipids with high structural complexity. In addition, these columns appear to be better suitable for high-throughput analysis due to lower lipid retention via smaller surface area.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100295"},"PeriodicalIF":3.2,"publicationDate":"2025-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145684633","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-25DOI: 10.1016/j.jcoa.2025.100293
Claudia Desiderio , Alexandra Muntiu
The application of Capillary Zone Electrophoresis (CZE) in proteomic analysis has increased significantly since the technique was hyphenated with high resolution mass spectrometry (HR-MS). The coupling was successful in combining the analytical features of high separation efficiency and resolving power of CZE with the precision of molecular mass measurements and sequencing of HR-MS analyzers for characterization of proteins and peptides and of their proteoforms, even in pre-digested, intact and native form. In this regard, CZE has made a strong contribution in the field of analysis and characterization of monoclonal antibodies (mAb) biotherapeutics, due to its high resolution power and separation mechanism based on differential migration, demonstrating challenging features for discriminating variants with rapid and high-efficiency analyses and for assessment of manufacturing quality control and safety of the final product. This review aims to highlight the most recent applications of CZE-MS in separation and characterization of mAb biotherapeutics based on the most recent literature (2022-2025). The various analytical approaches used in proteins MS characterization will be illustrated, with particular emphasis on the top-down approach, widely applied in this field of analysis and offering comprehensive characterization of proteoforms and exploration of protein interactions under native conditions. CZE-MS demonstrated an effective analytical strategy for the detailed characterization of mAb proteoforms with high potential for application in the challenging task of monitoring mAb release in advanced drug delivery systems.
{"title":"The challenges and prospects of capillary zone electrophoresis-mass spectrometry for characterization of monoclonal antibodies biotherapeutics","authors":"Claudia Desiderio , Alexandra Muntiu","doi":"10.1016/j.jcoa.2025.100293","DOIUrl":"10.1016/j.jcoa.2025.100293","url":null,"abstract":"<div><div>The application of Capillary Zone Electrophoresis (CZE) in proteomic analysis has increased significantly since the technique was hyphenated with high resolution mass spectrometry (HR-MS). The coupling was successful in combining the analytical features of high separation efficiency and resolving power of CZE with the precision of molecular mass measurements and sequencing of HR-MS analyzers for characterization of proteins and peptides and of their proteoforms, even in pre-digested, intact and native form. In this regard, CZE has made a strong contribution in the field of analysis and characterization of monoclonal antibodies (mAb) biotherapeutics, due to its high resolution power and separation mechanism based on differential migration, demonstrating challenging features for discriminating variants with rapid and high-efficiency analyses and for assessment of manufacturing quality control and safety of the final product. This review aims to highlight the most recent applications of CZE-MS in separation and characterization of mAb biotherapeutics based on the most recent literature (2022-2025). The various analytical approaches used in proteins MS characterization will be illustrated, with particular emphasis on the top-down approach, widely applied in this field of analysis and offering comprehensive characterization of proteoforms and exploration of protein interactions under native conditions. CZE-MS demonstrated an effective analytical strategy for the detailed characterization of mAb proteoforms with high potential for application in the challenging task of monitoring mAb release in advanced drug delivery systems.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100293"},"PeriodicalIF":3.2,"publicationDate":"2025-11-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145618376","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The reliable identification of anaesthetic and opioid drugs in biological matrices is of critical importance in forensic toxicology, particularly in cases of drug misuse and suspected medical malpractice. Given the typically low concentrations of these analytes in postmortem samples, an efficient extraction and preconcentration strategy is indispensable. Capsule phase microextraction (CPME), a recently developed sol-gel–based sample preparation technology that encapsulates sorbents within a porous tubular polymeric membrane, was employed for the selective isolation of anaesthetics (bupivacaine, lidocaine, ropivacaine, and prilocaine) and the opioid (tramadol) from postmortem blood and tissue samples (liver, kidney, stomach, and spleen). Subsequent analysis was performed using gas chromatography–mass spectrometry (GC–MS). The optimized CPME-GC–MS method exhibited excellent analytical performance, achieving linear calibration ranges of 0.01–1.0 μg mL⁻¹ in blood and 0.01–1.0 μg g⁻¹ in tissue samples, with limits of quantification as low as 0.0060–0.0079 μg mL⁻¹ (blood) and 0.0080–0.0099 μg g⁻¹ (tissues). Precision studies confirmed strong method reliability, with intra-day repeatability below 5% and inter-day reproducibility below 10%. The method was successfully applied to authentic postmortem specimens from a suspected fatal intoxication case attributed to medical negligence involving anaesthetic and opioid drugs. Beyond its forensic applicability, the environmental and operational sustainability of the protocol was critically evaluated using ComplexMoGAPI, AGREE and CACI assessment tools, confirming the method’s greenness and practicality. Overall, the CPME-GC- MS workflow offers a robust, sensitive, and eco-friendly analytical platform, representing a significant advancement for drug monitoring in medico-legal investigations.
{"title":"Leveraging the potential of capsule phase microextraction for simultaneous determination of anaesthetic and opioid drugs in postmortem samples of its fatal intoxication","authors":"Bharti Jain , Rajeev Jain , Abuzar Kabir , Torki Zughaibi , Shweta Sharma","doi":"10.1016/j.jcoa.2025.100291","DOIUrl":"10.1016/j.jcoa.2025.100291","url":null,"abstract":"<div><div>The reliable identification of anaesthetic and opioid drugs in biological matrices is of critical importance in forensic toxicology, particularly in cases of drug misuse and suspected medical malpractice. Given the typically low concentrations of these analytes in postmortem samples, an efficient extraction and preconcentration strategy is indispensable. Capsule phase microextraction (CPME), a recently developed sol-gel–based sample preparation technology that encapsulates sorbents within a porous tubular polymeric membrane, was employed for the selective isolation of anaesthetics (bupivacaine, lidocaine, ropivacaine, and prilocaine) and the opioid (tramadol) from postmortem blood and tissue samples (liver, kidney, stomach, and spleen). Subsequent analysis was performed using gas chromatography–mass spectrometry (GC–MS). The optimized CPME-GC–MS method exhibited excellent analytical performance, achieving linear calibration ranges of 0.01–1.0 μg mL⁻¹ in blood and 0.01–1.0 μg g⁻¹ in tissue samples, with limits of quantification as low as 0.0060–0.0079 μg mL⁻¹ (blood) and 0.0080–0.0099 μg g⁻¹ (tissues). Precision studies confirmed strong method reliability, with intra-day repeatability below 5% and inter-day reproducibility below 10%. The method was successfully applied to authentic postmortem specimens from a suspected fatal intoxication case attributed to medical negligence involving anaesthetic and opioid drugs. Beyond its forensic applicability, the environmental and operational sustainability of the protocol was critically evaluated using ComplexMoGAPI, AGREE and CACI assessment tools, confirming the method’s greenness and practicality. Overall, the CPME-GC- MS workflow offers a robust, sensitive, and eco-friendly analytical platform, representing a significant advancement for drug monitoring in medico-legal investigations.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100291"},"PeriodicalIF":3.2,"publicationDate":"2025-11-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145618377","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-21DOI: 10.1016/j.jcoa.2025.100292
Sanka N. Atapattu , Colin F. Poole
Many partition processes of interest have been characterised using Abraham's solvation parameter model. However, to estimate physicochemical, environmental, or biophysical properties of compounds requires reliable solute descriptors. Exploitation of the correlation of partition processes and chromatographic retention data is an alternative approach to estimate properties of interest without relying on solute descriptors. The system constants of the solvation parameter model, along with screening tools such as Cos θ, d-parameter, D-parameter, principal component analysis, and hierarchical cluster analysis, facilitate the identification of similarities between partitioning and chromatographic systems. In this review we discuss various screening tools to identify similarities between partitioning and chromatographic systems as well as applications of surrogate chromatographic models to estimate physicochemical, environmental, and biophysical properties of interest.
{"title":"Surrogate chromatographic models and the solvation parameter model","authors":"Sanka N. Atapattu , Colin F. Poole","doi":"10.1016/j.jcoa.2025.100292","DOIUrl":"10.1016/j.jcoa.2025.100292","url":null,"abstract":"<div><div>Many partition processes of interest have been characterised using Abraham's solvation parameter model. However, to estimate physicochemical, environmental, or biophysical properties of compounds requires reliable solute descriptors. Exploitation of the correlation of partition processes and chromatographic retention data is an alternative approach to estimate properties of interest without relying on solute descriptors. The system constants of the solvation parameter model, along with screening tools such as Cos θ, <em>d</em>-parameter, <em>D</em>-parameter, principal component analysis, and hierarchical cluster analysis, facilitate the identification of similarities between partitioning and chromatographic systems. In this review we discuss various screening tools to identify similarities between partitioning and chromatographic systems as well as applications of surrogate chromatographic models to estimate physicochemical, environmental, and biophysical properties of interest.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100292"},"PeriodicalIF":3.2,"publicationDate":"2025-11-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145618381","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
This study developed a method to determine perfluorooctanoic acid in real water samples using ultra-high performance liquid chromatography-electrospray-tandem mass spectrometry after dispersive magnetic solid phase extraction. The adsorbent used in dispersive magnetic solid phase extraction was tea waste activated carbon modified with magnetic nanoparticles. Successful synthesis of the adsorbent was confirmed by Fourier Transform infrared spectroscopy, X-ray diffraction, and energy-dispersive X-ray spectroscopy. The morphological and surface properties were investigated using scanning and transmission electron microscopy. Various experimental factors affecting the extraction and preconcentration efficiency were optimised using the design of experiments. Under the optimised conditions, the developed method had a wide linear range (39-5000 ng/L) with a correlation coefficient of 0.9915. The limit of detection was 11.7 ng/L, whereas the limit of quantification was 39.0 ng/L. The precision in terms of repeatability (intraday, n = 4 replicates) and reproducibility (interday, n = 3 days) expressed as %RSD were 2.40% and 3.10% respectively, which indicated that the developed method had good precision. The trueness of the developed method was investigated using a spike recovery test, and percentage recoveries ranged from 88.2% to 98.4 %. The magnetic activated carbon adsorbent could be reused up to 4 cycles. The developed method was successfully applied to detect perfluorooctanoic acid in three river water samples. Finally, the extent of greenness of the developed method was evaluated using six metric tools, and the findings showed that the method was environmentally friendly, cost-effective, highly practical and applicable.
{"title":"Magnetic extraction and preconcentration of perfluorooctanoic acid in aqueous solutions using magnetic tea waste activated carbon","authors":"Nompumelelo Malatji , Anele Mpupa , Boris Mizaikoff , Philiswa Nosizo Nomngongo","doi":"10.1016/j.jcoa.2025.100290","DOIUrl":"10.1016/j.jcoa.2025.100290","url":null,"abstract":"<div><div>This study developed a method to determine perfluorooctanoic acid in real water samples using ultra-high performance liquid chromatography-electrospray-tandem mass spectrometry after dispersive magnetic solid phase extraction. The adsorbent used in dispersive magnetic solid phase extraction was tea waste activated carbon modified with magnetic nanoparticles. Successful synthesis of the adsorbent was confirmed by Fourier Transform infrared spectroscopy, X-ray diffraction, and energy-dispersive X-ray spectroscopy. The morphological and surface properties were investigated using scanning and transmission electron microscopy. Various experimental factors affecting the extraction and preconcentration efficiency were optimised using the design of experiments. Under the optimised conditions, the developed method had a wide linear range (39-5000 ng/L) with a correlation coefficient of 0.9915. The limit of detection was 11.7 ng/L, whereas the limit of quantification was 39.0 ng/L. The precision in terms of repeatability (intraday, n = 4 replicates) and reproducibility (interday, n = 3 days) expressed as %RSD were 2.40% and 3.10% respectively, which indicated that the developed method had good precision. The trueness of the developed method was investigated using a spike recovery test, and percentage recoveries ranged from 88.2% to 98.4 %. The magnetic activated carbon adsorbent could be reused up to 4 cycles. The developed method was successfully applied to detect perfluorooctanoic acid in three river water samples. Finally, the extent of greenness of the developed method was evaluated using six metric tools, and the findings showed that the method was environmentally friendly, cost-effective, highly practical and applicable.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100290"},"PeriodicalIF":3.2,"publicationDate":"2025-11-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145618378","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-19DOI: 10.1016/j.jcoa.2025.100289
Giovanni D’Orazio , Salvatore Fanali
This study investigated the effect of mobile phase composition, especially water content in a mixture of protic and non-protic organic solvents, on chiral recognition and separation efficiency using a cellulose tris(3‑chloro-4-methylphenylcarbamate)-based stationary phase in the enantioseparation of selected fungicide compounds. The chiral stationary phase, coated on porous native silica particles, was packed into fused silica capillaries (250 mm x 100 µm I.D.) via the "slurry" packing method. The capillary column was utilized for the enantiomeric separation of five azole compounds through capillary electrochromatography. The primary objective was to thoroughly investigate how the composition of the mobile phase affects enantioresolution. This was achieved by systematically varying the ratios of methanol, acetonitrile, and water while keeping the buffer concentration, type, and pH constant. The role of water content was studied in relation to chiral interactions by evaluating its effect on key chromatographic parameters such as retention factor, resolution, and efficiency.
By selecting an appropriate acetonitrile to methanol ratio at a fixed water content, chiral resolutions (Rs) ranging from approximately 2.0 to 3.5 were achieved for the tested analytes.
本研究利用纤维素三（3 -氯-4-甲基苯基氨基甲酸酯）固定相对选定的杀菌剂对映体分离进行了手性识别和分离，研究了流动相组成，特别是在质子和非质子有机溶剂混合物中含水量对手性识别和分离效率的影响。手性固定相涂覆在多孔的天然二氧化硅颗粒上，通过“浆液”填料法填充到熔融二氧化硅毛细管（250 mm x 100 μ m id）中。采用毛细管电色谱法对五种唑类化合物的对映体进行分离。主要目的是深入研究流动相的组成如何影响对映体分辨率。这是通过系统地改变甲醇、乙腈和水的比例来实现的，同时保持缓冲液的浓度、类型和pH恒定。通过评价水含量对保留系数、分辨率和效率等关键色谱参数的影响，研究了水含量与手性相互作用的关系。通过在固定的含水量下选择合适的乙腈与甲醇的比例，所测分析物的手性分辨率（Rs）范围约为2.0至3.5。
{"title":"Water and organic content tuning of mobile phase for Enantioseparation of fungicides by capillary electrochromatography using cellulose Tris(3‑chloro-4-methylphenylcarbamate) based stationary phase","authors":"Giovanni D’Orazio , Salvatore Fanali","doi":"10.1016/j.jcoa.2025.100289","DOIUrl":"10.1016/j.jcoa.2025.100289","url":null,"abstract":"<div><div>This study investigated the effect of mobile phase composition, especially water content in a mixture of protic and non-protic organic solvents, on chiral recognition and separation efficiency using a cellulose tris(3‑chloro-4-methylphenylcarbamate)-based stationary phase in the enantioseparation of selected fungicide compounds. The chiral stationary phase, coated on porous native silica particles, was packed into fused silica capillaries (250 mm x 100 µm I.D.) via the \"slurry\" packing method. The capillary column was utilized for the enantiomeric separation of five azole compounds through capillary electrochromatography. The primary objective was to thoroughly investigate how the composition of the mobile phase affects enantioresolution. This was achieved by systematically varying the ratios of methanol, acetonitrile, and water while keeping the buffer concentration, type, and pH constant. The role of water content was studied in relation to chiral interactions by evaluating its effect on key chromatographic parameters such as retention factor, resolution, and efficiency.</div><div>By selecting an appropriate acetonitrile to methanol ratio at a fixed water content, chiral resolutions (<em>Rs</em>) ranging from approximately 2.0 to 3.5 were achieved for the tested analytes.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100289"},"PeriodicalIF":3.2,"publicationDate":"2025-11-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145618379","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-19DOI: 10.1016/j.jcoa.2025.100288
Rahul Patel , Dhruvrajsinh Rathod , Kyle Jackson , Mrunal Ingawale , Yating Xu , Zeinab Hosseinidoust , Raja Ghosh
The rise in the number of antibiotic-resistant bacteria has created the need for alternative therapeutic approaches, including bacteriophage therapy. Bacteriophages are viruses that infect and lyse bacteria and therefore offer a highly targeted and adaptable solution to combat multidrug-resistant pathogens. However, the scalable purification of bacteriophages, while achieving high yield of product and efficient removal of impurities such as endotoxins, remains a significant barrier to their therapeutic adoption. This study presents a two-step purification process combining size-exclusion chromatography (SEC) and ultrafiltration as an efficient and scalable method for bacteriophage purification. T7 was used as the model bacteriophage. Preparative SEC using a cuboid device served as the first step for removing endotoxin and other impurities from bacterial culture filtrate. The SEC-purified phage sample was then processed using a carrier phase ultrafiltration and backflow recovery technique to further purify and enrich the bacteriophage. Phage titer, endotoxin level, and protein concentration were quantified at each stage to evaluate purification performance. The endotoxin level was substantially reduced, decreasing from 1.44 EU/mL in the initial culture filtrate to 0.49 EU/mL in the final purified phage sample, the FDA's acceptable limit for products that directly or indirectly contact the cardiovascular and lymphatic systems being 0.50 EU/mL. The phage titer was enhanced from 5.33×10⁹ PFU/mL (in the cell culture filtrate) to 9.33×10⁹ PFU/mL in the final product. The overall protein removal achieved was 95.8 % while the overall endotoxin removal was 92.5 %. These findings suggest that the proposed two-step purification process could be developed further and adopted for scalable production of high-purity bacteriophage preparations suitable for therapeutic applications.
{"title":"A size exclusion chromatography- and ultrafiltration-based two-step process for purifying phage for therapeutic applications","authors":"Rahul Patel , Dhruvrajsinh Rathod , Kyle Jackson , Mrunal Ingawale , Yating Xu , Zeinab Hosseinidoust , Raja Ghosh","doi":"10.1016/j.jcoa.2025.100288","DOIUrl":"10.1016/j.jcoa.2025.100288","url":null,"abstract":"<div><div>The rise in the number of antibiotic-resistant bacteria has created the need for alternative therapeutic approaches, including bacteriophage therapy. Bacteriophages are viruses that infect and lyse bacteria and therefore offer a highly targeted and adaptable solution to combat multidrug-resistant pathogens. However, the scalable purification of bacteriophages, while achieving high yield of product and efficient removal of impurities such as endotoxins, remains a significant barrier to their therapeutic adoption. This study presents a two-step purification process combining size-exclusion chromatography (SEC) and ultrafiltration as an efficient and scalable method for bacteriophage purification. T7 was used as the model bacteriophage. Preparative SEC using a cuboid device served as the first step for removing endotoxin and other impurities from bacterial culture filtrate. The SEC-purified phage sample was then processed using a carrier phase ultrafiltration and backflow recovery technique to further purify and enrich the bacteriophage. Phage titer, endotoxin level, and protein concentration were quantified at each stage to evaluate purification performance. The endotoxin level was substantially reduced, decreasing from 1.44 EU/mL in the initial culture filtrate to 0.49 EU/mL in the final purified phage sample, the FDA's acceptable limit for products that directly or indirectly contact the cardiovascular and lymphatic systems being 0.50 EU/mL. The phage titer was enhanced from 5.33×10⁹ PFU/mL (in the cell culture filtrate) to 9.33×10⁹ PFU/mL in the final product. The overall protein removal achieved was 95.8 % while the overall endotoxin removal was 92.5 %. These findings suggest that the proposed two-step purification process could be developed further and adopted for scalable production of high-purity bacteriophage preparations suitable for therapeutic applications.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100288"},"PeriodicalIF":3.2,"publicationDate":"2025-11-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145618380","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-14DOI: 10.1016/j.jcoa.2025.100287
Valentina Quarta, Francesca Merlo, Chiara Milanese, Antonella Profumo, Andrea Speltini
This work presents a novel application of biochar as a new-generation sorbent to bioanalytical sample preparation. The carbonaceous material produced by pyrolysis of vegetal wastes (orange and pumpkin peels) at 650°C was easily attached (∼ 4 mg) to a small magnetic stir bar (8 × 3 mm), enabling a novel stir bar sorptive microextraction protocol to be performed directly in the HPLC-MS vial. The device was first tested in synthetic saliva samples (1 mL) observing sorption equilibrium in 1 h contact time and full elution was achieved using 300 µL 1 % v/v acetic acid ethanol solution after 30 min. Under the selected conditions, recovery in real saliva, spiked with each analyte in the range 0.5-5 ng mL-1, was in the range 88%-112%, with good inter-day inter-batch device-to-device precision (RSD < 18%, n=3), and no need for additional clean-up. Reusability tests confirmed that the device retains its sorption/desorption efficiency over at least 20 consecutive cycles. Additionally, the greenness of this biochar-based “in-vial” approach as a new stir bar sorptive microextraction was proven by using different dedicated software. The practical applicability of the procedure was demonstrated in real saliva samples from healthy volunteers.
{"title":"Biochar-based “in-vial” stir bar sorptive microextraction for a greener determination of steroid hormones in saliva","authors":"Valentina Quarta, Francesca Merlo, Chiara Milanese, Antonella Profumo, Andrea Speltini","doi":"10.1016/j.jcoa.2025.100287","DOIUrl":"10.1016/j.jcoa.2025.100287","url":null,"abstract":"<div><div>This work presents a novel application of biochar as a new-generation sorbent to bioanalytical sample preparation. The carbonaceous material produced by pyrolysis of vegetal wastes (orange and pumpkin peels) at 650°C was easily attached (∼ 4 mg) to a small magnetic stir bar (8 × 3 mm), enabling a novel stir bar sorptive microextraction protocol to be performed directly in the HPLC-MS vial. The device was first tested in synthetic saliva samples (1 mL) observing sorption equilibrium in 1 h contact time and full elution was achieved using 300 µL 1 % v/v acetic acid ethanol solution after 30 min. Under the selected conditions, recovery in real saliva, spiked with each analyte in the range 0.5-5 ng mL<sup>-1</sup>, was in the range 88%-112%, with good inter-day inter-batch device-to-device precision (RSD < 18%, <em>n=3</em>), and no need for additional clean-up. Reusability tests confirmed that the device retains its sorption/desorption efficiency over at least 20 consecutive cycles. Additionally, the greenness of this biochar-based “in-vial” approach as a new stir bar sorptive microextraction was proven by using different dedicated software. The practical applicability of the procedure was demonstrated in real saliva samples from healthy volunteers.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100287"},"PeriodicalIF":3.2,"publicationDate":"2025-11-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145572015","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-13DOI: 10.1016/j.jcoa.2025.100275
Marijana Sokolovic , Natasa P. Kalogiouri , Victoria F. Samanidou , Camelia Tulcan , Camen Dorin , Roberta Tripon , Snježana Kazazić , Fani Krstulović , Mirta Balenović , Tajana Amšel Zelenika , Ana Vulić , Claudia Daniela Serban , Marija Berendika
Nutritional, microbiological and toxicological quality of plants like Pimpinella anisum (anise) and Foeniculum vulgare Mill (fennel) are important factors in assuring their beneficial characteristics and usage. Besides defining basic nutritional and safety status, understanding the profile of compounds like polyphenols provides insights into their potential phytochemical, antioxidant and antimicrobial effects. The aim of this study was to evaluate chemical composition and polyphenol profile, mycotoxicological and microbiological quality and assess the antimicrobial potential of fennel extracts against bacterial pathogens isolated from cats.
In our study, the proximate chemical compositions of both seeds were in line with previous researches. In fennel and anise seeds, the most abundant group of fatty acids were monosaturated fatty acids (82.15%, 67.16%), followed by polysaturated fatty acids (10.81%, 23.96%) and saturated fatty acids (7.04%, 8.88%). Among individual fatty acids, the most abundant were oleic acid (81.02%, 65.05%), followed by linoleic acid (10.29%, 23.06%), respectively. Polyphenol profiling of plant extracts showed the highest amounts of Hydroxytyrosol (1069.0 mg/kg), Naringenin (420.3 mg/kg) and Epicatechin G (555.4 mg/kg) in fennel, while Naringenin (667.3 mg/kg), Apigenin 360.4 mg/kg), Luteolin (360.4 mg/kg), Coutaric Acid (328.9 mg/kg) and Caftaric acid (317.7 mg/kg) were detected in anise. Microbiological analysis of both plants showed presence of bacteria, fungi, yeasts and tested mycotoxins. Antimicrobial activity of both plants was demonstrated on wild bacterial isolates from cats, with noticed differences for tested extraction solvents. The results of our study show the promising potential of plant extracts for treatment of infections caused by tested pathogenic bacteria.
{"title":"Determination of chemical components, phytochemical (polyphenol) profile, microorganisms and mycotoxins and evaluation of antimicrobial potential of Pimpinella anisum (Anise) and Foeniculum vulgare (Fennel) against bacteria isolated from cats","authors":"Marijana Sokolovic , Natasa P. Kalogiouri , Victoria F. Samanidou , Camelia Tulcan , Camen Dorin , Roberta Tripon , Snježana Kazazić , Fani Krstulović , Mirta Balenović , Tajana Amšel Zelenika , Ana Vulić , Claudia Daniela Serban , Marija Berendika","doi":"10.1016/j.jcoa.2025.100275","DOIUrl":"10.1016/j.jcoa.2025.100275","url":null,"abstract":"<div><div>Nutritional, microbiological and toxicological quality of plants like <em>Pimpinella anisum</em> (anise) and <em>Foeniculum vulgare</em> Mill (fennel) are important factors in assuring their beneficial characteristics and usage. Besides defining basic nutritional and safety status, understanding the profile of compounds like polyphenols provides insights into their potential phytochemical, antioxidant and antimicrobial effects. The aim of this study was to evaluate chemical composition and polyphenol profile, mycotoxicological and microbiological quality and assess the antimicrobial potential of fennel extracts against bacterial pathogens isolated from cats.</div><div>In our study, the proximate chemical compositions of both seeds were in line with previous researches. In fennel and anise seeds, the most abundant group of fatty acids were monosaturated fatty acids (82.15%, 67.16%), followed by polysaturated fatty acids (10.81%, 23.96%) and saturated fatty acids (7.04%, 8.88%). Among individual fatty acids, the most abundant were oleic acid (81.02%, 65.05%), followed by linoleic acid (10.29%, 23.06%), respectively. Polyphenol profiling of plant extracts showed the highest amounts of Hydroxytyrosol (1069.0 mg/kg), Naringenin (420.3 mg/kg) and Epicatechin G (555.4 mg/kg) in fennel, while Naringenin (667.3 mg/kg), Apigenin 360.4 mg/kg), Luteolin (360.4 mg/kg), Coutaric Acid (328.9 mg/kg) and Caftaric acid (317.7 mg/kg) were detected in anise. Microbiological analysis of both plants showed presence of bacteria, fungi, yeasts and tested mycotoxins. Antimicrobial activity of both plants was demonstrated on wild bacterial isolates from cats, with noticed differences for tested extraction solvents. The results of our study show the promising potential of plant extracts for treatment of infections caused by tested pathogenic bacteria.</div></div>","PeriodicalId":93576,"journal":{"name":"Journal of chromatography open","volume":"9 ","pages":"Article 100275"},"PeriodicalIF":3.2,"publicationDate":"2025-11-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145537589","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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