The LC for Pharma Resource Hub is a comprehensive platform designed to support professionals in the pharmaceutical industry by providing valuable resources related to liquid chromatography (LC). This hub offers a variety of materials, including webinars, technical articles, application notes, and case studies, all aimed at enhancing knowledge and proficiency in LC methodologies within pharmaceutical applications.​<h3>Key features of the LC for Pharma Resource Hub include:</h3><h4>Educational Webinars</h4>Access on-demand webinars presented by industry experts, covering topics such as method development, validation, and troubleshooting in LC.​<ul><li>Episode 1: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3437">USP 621 Harmonization - Method Scaling</a><ul><li>Guest Speakers: Amanda Guiraldelli, Ph.D. &amp; Catharine Layton</li></ul></li><li>Episode 2: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3438">USP 621 Revisions - S/N Calculation, Sensitivity and Peak Symmetry</a><ul><li>Guest Speakers: Edmond Biba, Ph.D. &amp; Neil Lander</li></ul></li><li>Episode 3: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3439">Enhanced Method Development</a><ul><li>Guest Speakers: Amanda Guiraldelli, Ph.D. &amp; Richard Verseput</li></ul></li><li>Episode 4: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3440">aQbD for OTC Products</a><ul><li>Guest Speakers: Joshua Bhattacharaya &amp; Fadi Alkhateeb, Ph.D.</li></ul></li><li>Episode 5: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3441">Impurity Profile Characterization</a><ul><li>Guest Speakers: Christian Zeine, Henry Foddy &amp; Margaret Maziarz</li></ul></li><li>Episode 6: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/4123">USP Chapter 129 Updates and SEC-MALS</a></li><li>Episode 7: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/4167">Dietary Supplement Quality</a></li><li>Episode 8: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3444">Lipid Nanoparticles</a></li><li>Episode 9: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3445">Nitrosamines</a></li><li>Episode 10: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/4169">Dissolution</a></li><li>Episode 11: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/4170">LG Polymers</a></li><li>Episode 12: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/4669">Method Validation</a></li><li>Episode 13: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/4670">Multi-Attribute Method</a></li></ul><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Waters_Solving_Problems_That_Matter_a_Waters_webinar_series_featuring_USP_speakers_ecf3502756/Waters-Solving-Problems-That-Matter-a-Waters-webinar-series-featuring-USP-speakers.jpg" alt="Waters Corporation: Solving Problems That Matter - a Waters webinar series featuring USP speakers" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Waters_Solving_Problems_That_Matter_a_Waters_webinar_series_featuring_USP_speakers_ecf3502756/thumbnail_Waters-Solving-Problems-That-Matter-a-Waters-webinar-series-featuring-USP-speakers.jpg 245w," sizes="100vw" width="800" height="418"><h4>Waters 2024 LC Symposium - Pushing the boundaries of LC to address today's challenges</h4><ul><li>Waters 2024 LC Symposium - Day 1 - <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3935">Advanced LC Technologies for Increasingly Complex Molecules</a><ul><li>Guest Speakers: Davy Guillarme, Ph.D., Guilherme (Gui) Guimaraes, Ph.D., &amp; Michelle Chen, Ph.D.</li></ul></li><li>Waters 2024 LC Symposium - Day 2 - <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/webinar/3936">Next Generation HPLC: the Quest for Performance and Simplicity</a><ul><li>Guest Speakers: Amanda Guiraldelli, Ph.D., Jennifer Simeone, &amp; Andrea Gheduzzi</li></ul></li></ul><h4>Brochures and specifications</h4><ul><li>eBook: Improving Chromatographic Separations of Biopharmaceuticals With MaxPeak High Performance Surfaces (HPS) Technology<ul><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/labrulez-bucket-strapi-h3hsga3/720008011en_dd0b025cd5/720008011en.pdf">Full PDF for download</a></li></ul></li><li>White Paper: MaxPeak Premier Solutions: Improving Consumer Safety Through Innovative Science<ul><li><a target="_blank" rel="noopener noreferrer" href="https://www.waters.com/waters/library.htm?locale=en_US&amp;lid=135123293&amp;cid=511436&amp;srsltid=AfmBOoripwfSGOjTFXNWqjDHknCTbtr0G9sKgb9BsUD-L9avphhNaGuH">Full PDF for download</a></li></ul></li><li>Application Notebook: Alliance iS<ul><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/labrulez-bucket-strapi-h3hsga3/720008280en_e9d283fb41.pdf">Full PDF for download</a></li></ul></li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.cz/products/210">Alliance iS </a>Product Information</li><li>ICH Q14 Enhanced Approach: Tools in Empower Software to Support Your Analytical Procedure Development<ul><li><a target="_blank" rel="noopener noreferrer" href="https://cdn.technologynetworks.com/TN/resources/PDF/Tools-in-Empower-Software-to-Support-Your-Analytical-Procedure-Development.pdf">Full PDF for download</a></li></ul></li><li><a target="_blank" rel="noopener noreferrer" href="https://www.wyatt.com/products/instruments/multi-angle-light-scattering-detectors-sec-mals.html">MALS Detectors Product Information</a></li></ul><h4>Guides</h4><ul><li>LC Adjustments Made Simple: A Practical Guide<ul><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/labrulez-bucket-strapi-h3hsga3/720008065en_88843a7860/720008065en.pdf">Full PDF for download</a></li></ul></li><li>Guidebook: Characterization of New Modalities</li><li>Multi-Attribute Methods for Biopharmaceutical Analysis<ul><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/labrulez-bucket-strapi-h3hsga3/waters_mam_ebook_2023_84c6ae3f04/waters_mam_ebook_2023.pdf">Full PDF for download</a></li></ul></li><li><a target="_blank" rel="noopener noreferrer" href="https://www.waters.com/waters/library.htm?lid=135083997&amp;icid=hm-inno_00326&amp;srsltid=AfmBOopUZ0Cbncgz3zb34VIy1ysiVknnwF8Bca5aLAmt2KFOLtB3ZGhc">AQbD Method Development Infographic - Achieve Consistent Reportable Results</a></li></ul>By offering these resources, the LC for Pharma Resource Hub serves as a valuable tool for professionals seeking to enhance their understanding and application of liquid chromatography in the pharmaceutical industry.<blockquote>For complete information, please visit the <a target="_blank" rel="noopener noreferrer" href="https://gateway.on24.com/wcc/experience/elitewatersregional/2426651/4494230/method-management-campaign?partnerref=CZLabrulez">LC Regulated Workflows: Access insights from Pharma industry experts website</a></blockquote>

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Evaluating the potential of hydrophilic interaction liquid chromatography for collagen peptide mapping analysis

This study presents novel LC-TQ-MS/MS and cost-effective UPLC methods for quantifying N-nitroso drug substance-related impurity (N-MFA) in mefenamic acid tablets and pediatric suspensions. Both methods were successfully validated according to ICH guidelines, demonstrating excellent sensitivity, linearity, and accuracy. LC-MS/MS achieved quantification at 0.01 ng/mL (1 mg/mL MFA), while UPLC quantified down to 0.036 μg/mL (10 mg/mL MFA). Linearity was confirmed across 0.01–125 ppm for LC-MS/MS and 3.6–125 ppm for UPLC with R² = 0.999.Accuracy recoveries ranged from 90.43% to 101.34% for LC-MS/MS and 92.12% to 101.21% for UPLC. Both methods were applied to commercial formulations and demonstrated suitability for routine use in R&amp;D and quality control labs. Additionally, sustainability and environmental impact were evaluated using tools like GAPI, AGREE, and RGB12, confirming their green analytical profile.<blockquote><h4>The original article</h4><h4><a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597">A novel LC-TQ-MS/MS method for quantifying mefenamic acid-NDSRI (N-nitroso drug substance-related impurity) in mefenamic acid tablet and pediatric suspension dosage forms: a comparative study with a cost-effective white, green, and blue UPLC method</a></h4>Srinivas Nakka, Naresh Kumar Katari, Siva Krishna Muchakayala, Sreekantha Babu Jonnalagadda, Surendra Babu Manabolu Surya&nbsp;RSC Advances, Volume 15, Issue 3, 2025, 1957-1969<a target="_blank" rel="noopener noreferrer" href="https://doi.org/10.1039/d4ra08425j">https://doi.org/10.1039/d4ra08425j</a><h6>licensed under <a target="_blank" rel="noopener noreferrer" href="https://creativecommons.org/licenses/by-nc/3.0/">CC-BY 3.0</a></h6></blockquote>Selected sections from the article follow. Formats and hyperlinks were adapted from the original.Regulatory agencies have underscored the significance of predicting toxicity and precisely measuring the levels of nitrosamine contaminants in certain pharmaceuticals. These impurities, which include nitrosamines derived from active pharmaceutical ingredients, are referred to as Nitrosamine Drug Substance Related Impurities (NDSRIs). Nitrosamines, or N-nitrosamines, are molecules characterized by their N-nitroso functional group. They are classified as a group of concern due to their status as human carcinogens, with evidence showing that long-term intake above acceptable levels can increase cancer risks.These NDSRI substances are categorized into one of three “Cohort-of-Concern” (CoC) groups: aflatoxin-like, alkyl-azoxy, and N-nitroso. “CoC” denotes a collection of highly potent mutagenic compounds requiring special attention due to potential health risks. The focus of pharmaceutical regulatory authorities has shifted from common nitrosamines to NDSRIs because of their potential carcinogenicity and their presence in drug products at unacceptable levels.1–6 As a result, regulatory authorities recalled various drug products worldwide. Furthermore, the detection and quantification of NDSRIs are of significant concern to analytical researchers.7–11Nonsteroidal anti-inflammatory drugs (NSAIDs) are a class of medications that relieve pain and inflammation, including in renal transplants and chronic non-cancer pain in children.12,13 About 30 million people worldwide use NSAIDs every day, according to a report by Medical News Today. The worldwide market for NSAIDs was valued at USD 15.58 billion in 2019 and is envisioned to reach USD 24.35 billion by 2027.Mefenamic acid (MFA) is chemically known as 2-[(2,3-dimethylphenyl)amino]benzoic acid, with the chemical formula C15H15NO2 and a molecular weight of 241.28 g mol−1. MFA is classified as an anthranilic acid (phenolate) NSAID that exhibits both antipyretic and analgesic properties. This medication treats moderate pain and inflammation associated with osteoarthritis, post-operative pain, menstrual pain, rheumatoid arthritis, toothaches, and severe back and muscle pain. Additionally, it is also used to manage menorrhagia.14,15To our knowledge, this is the first UPLC-MS/MS and environmentally friendly UPLC method for quantifying the target compound N-MFA. As a result, there are no existing methods available to compare with the proposed UPLC-TQ-MS/MS and the cost-effective green UPLC method. Various regulatory agencies, including the US FDA and Health Canada, have proposed that the acceptable intake (AI) value for N-MFA is 78 000 ng per day.16,17 The interim control limit for N-MFA is 62.4 ppm, which can be derived from the AI value of the impurity and the maximum daily dose of the MFA. Hence, simple and highly sensitive UPLC-MS/MS and UPLC methods have been developed for the accurate quantification of N-MFA in MFA drug formulations, including tablets and pediatric suspensions.<div>The current study aimed to assess genotoxicity using (Q)-SAR (Quantitative Structure–Activity Relationship) models and to develop new LC-TQ-MS/MS and UPLC methods for quantifying N-MFA in both the MFA drug substance and its formulation dosage forms. Recent regulatory guidelines demonstrate that setting the quantitation limit (QL) at 30% is considered reasonably good and acceptable. If the values consistently fall below the QL across all process validation batches and corresponding stability samples (both accelerated and long-term), it is acceptable to proceed with skip lot testing. UPLC can detect the N-MFA impurity below the 30% control limit (∼19 ppm). Therefore, this cost-effective UPLC method is proposed alongside the LC-TQD-MS/MS for use in quality control laboratories. Both methods have been successfully validated regarding the QL, linearity, precision, accuracy, and robustness. The sustainability and eco-friendliness of the UPLC method were evaluated using greenness and whiteness assessment metrics, including GAPI, the analytical eco-scale, AGREE, AGREEprep, BAGI, and RGB12 tools.18–20 The results indicated that the proposed method exhibits a high level of greenness and whiteness. <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#fig1">Fig. 1</a> illustrates the chemical structures of MFA, N-DPA (N-nitroso diphenylamine), and N-MFA. The applicability of these methods was successfully verified through the analysis of commercial samples of APIs and dosage forms of MFA.<h4>Materials and method</h4><h5>Instrument and software</h5>The chromatographic investigation was accomplished on the <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/198">Waters H class UPLC system</a>, which includes a PDA-photo diode array detector model (L20UPD127A) modules, an Auto Sampler Manager with a Flow-Through Needle (FTN-L20FTP352G), Quaternary Solvent Manager (QSM-L20QSP358A) (Waters Corporation, USA). For the LC-TQ-MS/MS method (MRM analysis) a <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/195">Waters Xevo TQ-XS MS</a> system equipped with an ESI ion source was utilized. A Mettler-Toledo (Model: XPE205, Columbus, Ohio, USA) analytical balance was employed to weigh the samples and the impurity standard. The analysis utilized high-purity water obtained from the Millipore Milli-Q purification system (Bedford, MA, USA). Centrifuge (Model: 5810R, Hemburg, Germany) from Eppendorf. ChemDraw Professional 15.0 sketched MFA, N-DPA, and N-MFA molecular structures. The two primary (Q)-SAR methodologies employed were the Derek Nexus (version 6.2.0, featuring the Derek KB 2022 1.0 knowledge base) and the Sarah Nexus (version 3.2.0, which includes the Nexus version 1.9 and the Sarah Model 2022.1). <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?mainauthorItems=5&amp;search=TargetLynx">Target Lynx software</a> v 4.2 was used to compute all LC-TQ-MS/MS acquisition and process parameters.<h4>Results and discussion</h4><h5>Development and optimization of mass spectrometry conditions</h5>To develop an accurate quantification of N-MFA in MFA tablets and pediatric suspensions, optimizing mass parameters is a decisive step. We intended to attain a sensitive, precise, and accurate method. For N-MFA quantification, the MRM technique was utilized. For initial mass conditions optimization by injecting the 1000 ng mL−1 concentration of N-MFA into mass spectrometry using infusion mode to identify the prominent product ion in electron spray ionization in positive ion mode (m/z 271.13 → 241.14). Due to the pump flow rate being set at 0.7 mL min−1 and the recommended desolvation gas temperature and gas flow being 450 °C and 900 L per Hr, hence performed the analysis with the values mentioned earlier. Employing the Intellistart software, which is built into Waters mass lynx software, to fine-tune MS/MS parameters, including cone voltage, capillary voltage, and collision energy values with different mass transitions. Intellistart identified three major MRM transitions, including m/z 271.13 → 241.14, m/z 271.13 → 223.12, and m/z 271.13 → 208.10 (<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#fig2">Fig. 2</a>). After careful optimization of cone voltage and collision energies finalized, the quantifier ion transition as m/z 271.13 → 241.14 and the qualifier ion transition as m/z 271.13 → 223.12 (Fig. S4<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#fn1">†</a>). A mass spectrometer was installed, a 50 μL loop, which improves the peak shape when injecting the samples containing a high percentage of organic solvents. The highest intensity of quantifier and qualifier ions are detected by using the below MS/MS optimized conditions: cone voltage is 18 V, capillary voltage is 2.48 kV, collision energy is 8 V and 18 V for qualifier and quantifier respectively, desolvation gas temperature is 450 °C, and gas flow 900 L h−1. In the MS/MS method, the events program flow state was directed from 0 to 5.4 min set to mass to detect the N-MFA and the flow state from 5.5 to 10 minutes to avoid the high concentration of MFA contamination triggered by formulation tablet and pediatric suspension samples 1 mg mL−1 concentration. The optimized MS/MS parameters are shown in <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#tab2">Table 2</a>.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/RSC_Advances_Volume_15_Issue_3_2025_Pages_1957_1969_Fig_2_A_Mass_spectrum_of_NMFA_and_B_MSMS_fragmentation_pattern_of_N_MFA_09e3975748.jpg" alt="RSC Advances, Volume 15, Issue 3, 2025, Pages 1957-1969: Fig. 2. (A) Mass spectrum of NMFA, and (B) MS/MS fragmentation pattern of N-MFA." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_RSC_Advances_Volume_15_Issue_3_2025_Pages_1957_1969_Fig_2_A_Mass_spectrum_of_NMFA_and_B_MSMS_fragmentation_pattern_of_N_MFA_09e3975748.jpg 182w," sizes="100vw" width="1500" height="1286"><h5>Method validation</h5>The present proposed LC-TQ-MS/MS and UPLC methods have been fully validated by ICH Q2R2, USP 〈1225〉 guidelines,<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#cit23">23</a> and published journals,24–26 following the method validation parameters, such as specificity, limit of detection, limit of quantitation, linearity, precision, accuracy, robustness, and solution stability. <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#tab3">Table 3</a> presents the results of the validation study, demonstrating that the method is specific, precise, linear, and accurate.<h5>Specificity</h5>The current analytical method's specificity is demonstrated by the liquid chromatography system's capability to differentiate between the diluent, placebo, MFA, N-MFA, and N-MFA spiked at the specification level. Injected each 10 μL for LC-MS/MS and 50 μL for UPLC of diluent, placebo, N-MFA standard solution, and sample solution of MFA. The analysis showed that no co-eluting peaks were apparent at the retention time (tR) of N-MFA, allowing for the specific and accurate estimation of N-MFA in MFA tablet and pediatric suspension formulations. Overlay UV spectra of N-MFA and MFA are shown in Fig. S4,<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#fn1">†</a> and the typical LC-MS/MS and UPLC chromatograms indicating the specificity of the method can be found in <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#fig3">Fig. 3</a> and <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/org/science/article/pii/S2046206925001597#fig4">4</a>, respectively.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/RSC_Advances_Volume_15_Issue_3_2025_Pages_1957_1969_Fig_3_Typical_MRM_chromatograms_of_A_placebo_B_N_MFA_standard_22464e21b1.jpg" alt="RSC Advances, Volume 15, Issue 3, 2025, Pages 1957-1969: Fig. 3. Typical MRM chromatograms of (A) placebo, (B) N-MFA standard at the specification level, and (C) UV spectra of N-MFA spiked to the MFA sample at the specification level, and (D) MRM of N-MFA spiked to the MFA sample at the specification level." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_RSC_Advances_Volume_15_Issue_3_2025_Pages_1957_1969_Fig_3_Typical_MRM_chromatograms_of_A_placebo_B_N_MFA_standard_22464e21b1.jpg 137w," sizes="100vw" width="1500" height="1710"><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/RSC_Advances_Volume_15_Issue_3_2025_Pages_1957_1969_Fig_4_Typical_chromatograms_of_A_blank_B_placebo_C_N_MFA_standard_at_specification_level_1ba78e4737.jpg" alt="RSC Advances, Volume 15, Issue 3, 2025, Pages 1957-1969: Fig. 4. Typical chromatograms of (A) blank, (B) placebo, (C) N-MFA standard at specification level, and (D) N-MFA spiked to the MFA sample at specification level." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_RSC_Advances_Volume_15_Issue_3_2025_Pages_1957_1969_Fig_4_Typical_chromatograms_of_A_blank_B_placebo_C_N_MFA_standard_at_specification_level_1ba78e4737.jpg 125w," sizes="100vw" width="1500" height="1878"><h4>Conclusion</h4>In conclusion, two interdependent (Q)-SAR tools were utilized to assess and classify N-MFA as a Class-3 category, designated as a “CoC” which suggests potential genotoxicity. Diphenylamine is a precursor that can lead to the formation of N-MFA. The acceptable intake (AI) of N-MFA was derived from N-DPA, and the determined AI for N-MFA is 78 000 ng; hence the specification limit is 62.4 ppm. Proposed novel LC-TQ-MS/MS and cost-effective UPLC methods are proposed to accurately quantify N-nitroso mefenamic acid (N-MFA) in mefenamic acid (MFA) tablets and pediatric suspension dosage forms. The proposed method has been successfully validated in compliance with ICH guidelines. The developed methods were further applied to commercially available mefenamic acid formulations to determine accurate and precise quantification of N-MFA. For routine quality control laboratories, this cost-effective UPLC method can be used to detect 30% of the interim control limit (∼19 ppm). Furthermore, sustainability and eco-friendliness were evaluated using greenness and whiteness assessment metrics, including the analytical eco-scale, GAPI, AGREE, AGREEprep, BAGI, and RGB12 tools. The results indicated that the method proposed exhibits a high level of greenness and whiteness. Both methods can be used to determine N-MFA content in mefenamic acid formulation dosage based on the availability of instruments LC-TQ-MS/MS and UPLC in R&amp;D and quality control laboratories.</div>

A novel LC-TQ-MS/MS method for quantifying mefenamic acid-NDSRI (N-nitroso drug substance-related impurity) in mefenamic acid tablet and pediatric suspension dosage forms: a comparative study with a cost-effective white, green, and blue UPLC method

This study presents a novel and eco-friendly LC-QqQ-MS/MS method for the trace-level quantification of N-Nitroso Moxifloxacin (N-MOX) and di-Nitroso pyrrolopiperidine (DNPP) in moxifloxacin drug substances and products. As both NDSRIs fall into potency category 4 with an acceptable intake limit of 1500 ng/day, their control is vital for patient safety. The method achieves efficient chromatographic separation using a Cortecs T3 C18 column and gradient elution, and employs MRM transitions m/z 431.0 → 386.0 for N-MOX and 185.1 → 138.1 for DNPP.Validated per ICH guidelines, the method proved accurate, sensitive, and reproducible. Green assessment tools confirmed its low environmental impact in terms of time, solvent use, waste, and sample volume. Successfully applied to commercial moxifloxacin tablets and eye drops, this method offers a robust solution for routine NDSRI monitoring in pharmaceuticals.<blockquote><h4>The original article</h4><h4><a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971">A facile and eco-friendly simultaneous quantification LC-TQ-MS/MS approach for N-Nitroso Moxifloxacin and di-nitroso pyrrolopiperidine in Moxifloxacin tablets and eye drops</a></h4>Srinivas Nakka, Siva Krishna Muchakayala, Surendra Babu Manabolu Surya&nbsp;Green Analytical Chemistry, Volume 12, 2025, 100188<a target="_blank" rel="noopener noreferrer" href="https://doi.org/10.1016/j.greeac.2024.100188">https://doi.org/10.1016/j.greeac.2024.100188</a><h6>licensed under <a target="_blank" rel="noopener noreferrer" href="https://creativecommons.org/licenses/by/4.0/">CC-BY 4.0</a></h6></blockquote>Selected sections from the article follow. Formats and hyperlinks were adapted from the original.The emergence of nitrosamine impurities in active pharmaceutical ingredients (APIs) is a complex issue influenced by various factors related to chemical synthesis, raw material quality, and manufacturing processes. The concern of nitrosamine contamination in pharmaceuticals is quickly becoming more widespread, extending not only beyond smaller nitrosamines but also nitrosamine drug substance-related impurities (NDSRIs). These NDSRIs are particularly challenging because they are structurally related to the API, making their presence almost unavoidable [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0001">[1]</a>, <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0002">[2]</a>, <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0003">[3]</a>, <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0004">[4]</a>]. Moreover, the lack of carcinogenicity or mutagenicity data for most NDSRIs complicates the establishment of acceptable intake levels (AIs). Based on the GSRS (Global Substance Registration System) database, 41.4 % of APIs and 30.2 % of API impurities are potential nitrosamine precursors. However, when tertiary amines are excluded from this classification, the percentages adjust to 14.7 %, and 12.8 % are for APIs and API impurities, respectively, that could potentially lead to nitrosamine formation [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0005">5</a>].In July 2023, the European Medicines Agency (EMA) introduced a novel assessment method known as the Carcinogenic Potency Categorization Approach (CPCA) for N-nitrosamines. This approach was designed to facilitate the determination of AI for nitrosamines, thereby simplifying the complexities associated with setting AI [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0006">6</a>,<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0007">7</a>]. Developing an analytical method with high sensitivity for regular analysis presents a considerable challenge, given the outstandingly low acceptable intake thresholds for these impurities and is fraught with sensitivity issues, while reactivity and stability concerns further complicate the analysis.Moxifloxacin (MOX) is a broad-spectrum antibiotic that acts against both Gram-positive and Gram-negative bacteria's from the fourth-generation fluoroquinolone class of medication and is prescribed for numerous bacterial ailments such as pneumonia, conjunctivitis, endocarditis, tuberculosis, and sinusitis. It can be administered orally, intravenously, or as eye drops. It obstructs the bacteria's DNA replication process, vital for their growth and proliferation [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0008">[8]</a>, <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0009">[9]</a>, <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0010">[10]</a>]. It is acknowledged worldwide for its efficacy in treating numerous bacterial infections and is listed among the World Health Organization (WHO)’s essential medicines.Green analytical chemistry (GAC) is dedicated to developing and applying methods that minimize environmental impact and improve sustainability. The core goal of this approach is to reduce the use of hazardous substances, minimize waste production, and encourage energy-efficient analytical processes. In 1998, Paul Anastas and his team introduced twelve principles to assess the environmental impact of chemical processes, which were later adapted for analytical methods [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0011">11</a>]. Gałuszka et al. developed twelve principles specifically for green analytical chemistry, incorporating the core concepts of green chemistry [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0012">12</a>]. It involves analyzing smaller sample quantities and reducing the usage or volume of toxic reagents, solvents, and energy consumption. Experts in green chemistry focus on the use of safe, environmentally friendly chemicals and low-waste assessment methods [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0013">[13]</a>, <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0014">[14]</a>].<div>A comprehensive literature review reveals that, to date, there are no published research articles on the simultaneous trace-level quantification of N-MOX (1-cyclopropyl-6-fluoro-8-methoxy-7-((4aS,7aS)-1-nitrosooctahydro-6H-pyrrolo[3,4-b]pyridin-6-yl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid), and DNPP {(4aS,7aS)-1,6-dinitrosooctahydro-1H-pyrrolo[3,4-b]pyridine}. The current study aimed to assess genotoxicity by utilizing Quantitative structure-activity relationship ((Q)-SAR) models that apply carcinogenic potency category classification, while also developing a straightforward, rapid, and novel UPLC-triple quadrupole (QqQ)-MS/MS method for the simultaneous trace-level quantification of N-MOX and DNPP impurities. The MOX, N-MOX, and DNPP chemical structures are illustrated in <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#fig0001">Fig. 1</a>.<h4>2. Materials and methods</h4><h5>2.1. Instruments and software</h5>The chromatographic study was performed on the <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/198">Waters H class UPLC</a> (Ultra Performance Liquid Chromatography) system, which includes a PDA-photo diode array detector model (L20UPD127A) modules, an Auto Sampler Manager with a Flow-Through Needle (FTN- L20FTP352G), Quaternary Solvent Manager (QSM- L20QSP358A) (Waters Corporation, USA). A <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/195">Waters Xevo TQ-XS LC-MS system</a> equipped with a triple quadrupole (QqQ) mass analyzer and Electrospray Ionization (ESI) ion source with multiple reaction monitoring (MRM) techniques was utilized (Waters Corporation, Milford, Massachusetts, USA). A Mettler-Toledo (Model: XPE205, Columbus, Ohio, USA) analytical balance was employed to weigh the samples and the impurity standard. The analysis utilized high-purity water consists of resistivity &lt;18.2 MΩ˙cm at 25 °C, TOC (Total Oxidizable Carbon) ≤5 ppb, Bacteria &lt;10 CFU (Colony-forming unit) / 100 mL obtained from the Millipore Milli-Q purification system (Bedford, MA, USA). Centrifuge (Model: 5810R, Hemburg, Germany) from Eppendorf. Millex-GV hydrophilic 0.22 µm PVDF filters (Burlington, Massachusetts, USA) have been obtained from Millipore. Using ChemDraw Professional 15.0, sketched the chemical structures of MOX, N-MOX, and DNPP. The two primary (Q)-SAR methodologies employed were the Derek Nexus (version 6.2.0, featuring the Derek KB 2022 1.0 knowledge base) and the Sarah Nexus (version 3.2.0, which includes the Nexus version 1.9 and the Sarah Model 2022.1). <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/635">MassLynx</a> and TargetLynx software (v 4.2) were used for all LC-QqQ-MS/MS acquisition parameters and to process the data, respectively.<h4>3. Results and discussion</h4><h5>3.4. Development and optimization of MS/MS conditions</h5>Multiple Reaction Monitoring (MRM) is an analytical method of high sensitivity and specificity, gaining prominence in different fields, including drug development, clinical research, and metabolomics [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0017">17</a>]. It enables precise detection and accurate quantification of impurities at very low concentrations. During the current study, N-MOX and DNPP impurities were detected and quantified using the MRM approach in an electrospray ionization positive mode (ESI). The capillary voltage (kV), desolvation temperature ( °C), and desolvation gas flow (L/Hr) are the three major ESI source parameters. Concerning the low pump flow rate of 0.3 mL/min, the recommended desolvation gas temperature and gas flow are 450 °C and 900 L/Hr. Both N-MOX and DNPP showed maximum intensity at a capillary voltage of 3.0 kV. The MS parameters for MRM channels, such as collision energy (CE) and cone voltage (CV), were individually optimized to ensure a stable and robust signal for the compounds. Intellistart software, built into Waters mass lynx software, is employed to fine-tune MS/MS parameters, including cone voltage and collision energy values with different mass transitions. MS/MS fragmentation pattern for both impurities shown in <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#fig0003">Fig. 3</a>. After careful optimization of cone voltage and collision energies finalized, the quantifier ion transition as m/z 185.1→ 138.1 and the qualifier ion transition as m/z 185.1 → 96.10 for DNPP; the quantifier ion transition as m/z 431→ 386 and the qualifier ion transition as m/z 431 → 319 for N-MOX (Fig. S2). A mass spectrometer, a 50 µL loop, was installed, which improves the peak shape when injecting samples containing a high percentage of organic solvents. Later, studies of the impact of collision energies varying from 5 to 30 V and optimized collision energy for N-MOX and DNPP were 20 and 15 V, respectively (Fig. S3). The MRM quantifier qualifier and ESI source parameters for both impurities were tabulated in <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#tbl0002">Table 2</a>. In the MS/MS method; the events program was configured to direct the flow from 0 to 2.8 min. for mass detection of DNPP. To prevent contamination from high concentrations of MOX from tablets and eye drops (5 mg/mL concentration), the flow was diverted to waste from 2.9 to 4.5 min. From 4.6 to 8 min, the flow was directed again to detect N-MOX.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Green_Analytical_Chemistry_Volume_12_2025_100188_Fig_3_MS_MS_fragmentation_pattern_for_A_DNPP_and_B_N_MOX_fce810c628.jpg" alt="Green Analytical Chemistry, Volume 12, 2025, 100188: Fig. 3. MS/MS fragmentation pattern for (A) DNPP, and (B) N-MOX." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Green_Analytical_Chemistry_Volume_12_2025_100188_Fig_3_MS_MS_fragmentation_pattern_for_A_DNPP_and_B_N_MOX_fce810c628.jpg 232w," sizes="100vw" width="1338" height="900"><h5>3.9. Method greenness assessment of the proposed method</h5>The environmental impact or “greenness” of the recently developed LC-QqQ-MS/MS method was evaluated using insights from published journals [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0024">[24]</a>, <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0025">[25]</a>, <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0026">[26]</a>] and contemporary tools, including the Analytical Eco-scale, GAPI, AGREE, AGREEprep, and the Blue Applicability Grade Index (BAGI).<h5>3.9.2. Green analytical procedure index (GAPI) assessment tool</h5>In 2018, Płotka-Wasylka introduced the GAPI [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0028">28</a>], which uses a pictogram with red, yellow, and green colors to represent high, medium, and low environmental impacts, respectively. For the developed LC-QqQ-MS/MS method, pictograms 2, 3, 6, 8, 9, and 12 are green, indicating low environmental impact. Pictograms 4, 5, 10, 11, and 14 are yellow, reflecting medium impact, while pictograms 1, 7, 13, and 15 are red, indicating high impact. This visual representation highlights both qualitative and quantitative environmental impacts [<a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#bib0029">29</a>]. An illustration of the GAPI pictogram is provided in <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#fig0007">Fig. 7</a>A. <a target="_blank" rel="noopener noreferrer" href="https://www.sciencedirect.com/science/article/pii/S2772577424000971#tbl0006">Table 6</a> details the principles of GAPI, the selected method condition values, and their corresponding scores.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Green_Analytical_Chemistry_Volume_12_2025_100188_Fig_7_Pictograms_of_A_GAPI_B_AGREE_C_AGRE_Eprep_and_D_BAGI_11db2203ed.jpg" alt="Green Analytical Chemistry, Volume 12, 2025, 100188: Fig. 7. Pictograms of (A) GAPI (B) AGREE, (C) AGREEprep, and (D) BAGI." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Green_Analytical_Chemistry_Volume_12_2025_100188_Fig_7_Pictograms_of_A_GAPI_B_AGREE_C_AGRE_Eprep_and_D_BAGI_11db2203ed.jpg 106w," sizes="100vw" width="1090" height="1600"><h4>4. Conclusion</h4>A novel, highly sensitive, and eco-friendly UPLC-QqQ-MS/MS method has been developed for the simultaneous trace-level quantification of MOX NDSRI and DNPP in MOX drug substances, tablets, and eye drops. The method was rigorously validated according to ICH Q2(R2) guidelines, including parameters such as linearity, LOD, LOQ, accuracy, precision, and robustness, showing excellent performance with high recovery rates and lower % RSD values and detecting N-MOX from 0.1 ng/mL to 37.5 ng/mL and DNPP from 0.5 ng/mL to 37.5 ng/mL. The method's versatility and sensitivity were demonstrated through a successful application to the analysis of commercial MOX tablets and eye drops, thereby ensuring the safety of MOX formulations. Furthermore, the proposed LC-QqQ-MS/MS method was evaluated using advanced green tools such as Analytical Eco-scale, GAPI, AGREE, AGREEprep, and BAGI, which confirmed its sustainability. This method can be employed for the accurate trace-level quantification of N-MOX and DNPP in any of the drug substances and formulations adopting suitable sample preparation. This methodology offers several advantages, such as reduced sample and organic solvent consumption, as well as a shorter run time, allowing for the processing of more batch samples. Overall, this validated UPLC-QqQ-MS/MS method presents a precise, reliable, and environmentally friendly approach for the simultaneous quantification of N-MOX and DNPP, offering a valuable contribution to pharmaceutical research and development.</div>

A facile and eco-friendly simultaneous quantification LC-TQ-MS/MS approach for N-Nitroso Moxifloxacin and di-nitroso pyrrolopiperidine in Moxifloxacin tablets and eye drops

Systematic comparison of wide pore size exclusion chromatography columns for the characterization of gene therapy products

News from LabRulezLCMS Library - Week 48, 2025

Our Library never stops expanding. What are the most recent contributions to LabRulezLCMS Library in the week of 19th January 2026? Check out new documents from the field of liquid phase, especially HPLC and LC/MS techniques!<blockquote>👉 <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library">SEARCH THE LARGEST REPOSITORY OF DOCUMENTS ABOUT LCMS AND RELATED TECHNIQUES</a></blockquote><blockquote>👉 Need info about different analytical techniques? Peek into <a target="_blank" rel="noopener noreferrer" href="https://gcms.labrulez.com/library">LabRulezGCMS</a> or <a target="_blank" rel="noopener noreferrer" href="https://icpms.labrulez.com/library">LabRulezICPMS</a> libraries.</blockquote>This week we bring you application notes by Agilent Technologies, Shimadzu, Waters Corporation and poster by Thermo Fisher Scientific!<h3>1. Agilent Technologies: Automated Bioprocess Monitoring with Agilent 1290 Infinity III Bio Online LC Solutions</h3>Process analytics of L-alanine production using genetically modified Vibrio natriegens<ul><li>Application note</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/labrulez-bucket-strapi-h3hsga3/an_automated_bioprocess_monitoring_1290_infinity_bioonline_solutions_5994_8865en_agilent_9f33de6ddf.pdf">Full PDF for download</a></li></ul>Bioprocess engineering is a challenging field because of the constantly changing nature of bioprocess models which make them difficult to validate.1 Consequently, process control needs to be developed at various levels using strategies tailored to these specific requirements.1 The performance and effectiveness of the bioreactor are determined by key performance indicators such as product yield, titer, production rate, and quality of the product.1 These factors can be quickly influenced by changes in process variables and the raw materials used.1,2 The ability to react quickly to problems is therefore crucial for preserving ongoing production, a factor that can be particularly significant in bioprocesses due to the substantial cost and time investmented.1,2 Such challenges can be tackled by constantly monitoring the bioprocess, which requires the ability to analyze and collect data quickly. Therefore, monitoring is of the utmost importance.&nbsp;This application note describes the advantages of using an Agilent 1290 Infinity III Bio Online LC solution with Agilent Online LC Monitoring software. A bioreactor was interfaced with a Flownamics Seg-Flow S3 sampling system for aseptic withdrawal and an Agilent 1290 Infinity III Bio Online LC to make an hourly analysis.&nbsp;This is a fully automated approach for taking sterile samples from a bioreactor and preparing or diluting them. In addition, the 1290 Infinity III Bio Online LC solution displays data in near real time, which is a crucial element for effective bioprocess&nbsp;monitoring.<h4>Experimental&nbsp;</h4><h5>Equipment&nbsp;</h5>The <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?mainauthorItems=10&amp;search=1290+Infinity+III+Bio">1290 Infinity III Bio </a>Online LC comprised the following&nbsp;modules:&nbsp;<ul><li>Agilent 1290 Infinity III Bio Flexible Pump (G7131A) with an InfinityLab Quick Change Bio inline filter assembly (part&nbsp;number 5067-1607), 2.1 mm id, 0.2 µm pore size&nbsp;</li><li>Agilent 1290 Infinity III Bio Online Sample Manager (G3167B) with Sample Thermostat (option #101) clustered with Agilent 1290 Infinity Valve Drive (G1170A), featuring a reactor valve pod (part number 5067-6680) and Online LC Monitoring Software&nbsp;</li><li>Agilent 1290 Infinity III Multicolumn Thermostat (G7116B) equipped with an Agilent Quick Connect Bio Heat Exchanger Std. (G7116-60071)&nbsp;</li><li>Agilent 1290 Infinity III Diode Array Detector (DAD) (G7117B), equipped with a Bio Max-Light Cartridge Cell (G4212-6008), 10 mm, 1 µL&nbsp;</li><li>Agilent 1260 Infinity High-Performance Degasser (G4225A) is connected between S1 (transport solvent) solvent line and Agilent 1290 Infinity III Bio Online Sample Manager (G3167B)&nbsp;</li></ul><h5>Software&nbsp;</h5><ul><li>Agilent <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?mainauthorItems=10&amp;search=OpenLab+CDS">OpenLab CDS</a> version 2.8 or later versions&nbsp;</li><li>Agilent Online LC Monitoring Software version 1.3 or later&nbsp;versions&nbsp;</li><li>Flownamics FlowWeb control&nbsp;software</li></ul><h5>Instrumental configuration&nbsp;</h5>The 1290 Infinity III Bio Online LC solution was used with a Seg-Flow interface for automatic sampling and online analysis (see Figure 1). The Seg-Flow interface was used to withdraw samples from a bioreactor through a Flownamics F-Series 310 FISP probe with a ceramic membrane filter (0.2&nbsp;µm pore size) from a 5 L bioreactor. The drawn samples were sent through the built-in sample collection cup on the Seg‑Flow to the LC and the samples were retained into vials. The collected samples were then automatically derivatized by using an injector program and subsequently analyzed. A more detailed description of the instrumental configuration has been published.3<h4>Conclusion&nbsp;</h4>For online bioprocess monitoring, a fully automated setup consisting of a bioreactor, a Flownamics sampling system, and an Agilent 1290 Infinity III Bio Online LC solution was successfully established. This setup is capable of taking aseptic samples from a bioreactor, preparing, analyzing, and processing samples without the need for manual interaction. In addition, the added value of the Agilent Online LC Monitoring software was evident in the display method and the ability to track it quickly.<h3>2. Shimadzu: Analysis of Oligonucleotide Impurities Using Single Quadrupole Mass Spectrometer</h3><ul><li>Application note</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/labrulez-bucket-strapi-h3hsga3/an_01_00974_en_c50bb67c7a.pdf">Full PDF for download</a></li></ul><h4>User Benefits</h4><ul><li>Combination use of LCMS-2050 single quadrupole mass spectrometer and LabSolutions Insight Biologics provides comprehensive characterization of oligonucleotides and related impurities.</li><li>Purity calculations for target components can be performed even on poorly separated peaks by utilizing MS spectra simultaneously acquired with UV chromatogram.</li></ul>In recent years, oligonucleotide therapeutics have been paid attention as a novel drug discovery modality because of their rapid progress of development. During the synthesis of oligonucleotide therapeutics, impurities that have differing base chain lengths are generated along with the target oligonucleotide. To ensure the safety and efficacy of the drug, comprehensive detection and identification of these impurities are required.&nbsp;Reversed phase ion-pair chromatography is often employed as a method to separate the target compound from co-existing impurities. However, it is often difficult to establish analytical conditions that provide compete separation of all impurities that are formed in synthesis process due to their similar structures and properties to the principal compound of active ingredient. Against this drawback, liquid chromatography-mass spectrometry (LC-MS) is commonly used for impurity detection and identification. For analysis of oligonucleotide impurities, quadrupole time-of-flight mass spectrometers are mainly used because of their high mass resolution and ability to perform sequence and structure estimations via MS/MS analysis. However, in quality control process, the use of a single quadrupole mass spectrometer is expected to be advantageous because of providing mass information with easy operation when impurity information is already known.&nbsp;This article presents a simulated impurity analysis of synthetic oligonucleotide using the LCMS-2050 high-performance liquid chromatography-mass spectrometer and LabSolutions Insight Biologics.<h4>Analytical conditions</h4>The employed instrument setup was <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?mainauthorItems=1&amp;search=Nexera+XS">Nexera XS inert</a> and <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?mainauthorItems=1&amp;search=LCMS-2050">LCMS-2050</a>. HPLC conditions are shown in Table 2, and MS conditions are shown in Table 3. The conditions were set where n-1(5') co-eluted with FLP, while n-3(5') was separated (Fig. 1) to demonstrate impurity analysis performance.<h4>Conclusion</h4>The simulated sample consisting of mainly FLP and small amounts of n-1(5')/n-3(5') was analyzed using the LCMS-2050 and LabSolutions Insight Biologics software and obtained data were processed appropriately. Detection, identification, and quantitation at low concentrations were accomplished not only for n-3(5') well-separated from FLP but also for n-1(5') co-eluted with FLP. In case that establishing analytical conditions is difficult, Using this instrument setup and software will provide efficient quality confirmation of oligonucleotide therapeutics.<h3>3. Thermo Fisher Scientific: Simultaneous Quantitation and Untargeted Discovery (SQUAD) workflow using multiple fragmentation techniques for quantitation and characterization of trace level nitrosamine in biological matrices</h3><ul><li>Poster</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/labrulez-bucket-strapi-h3hsga3/po_squad_workflow_multiple_fragmentation_techniques_po_en_94a5b27b12.pdf">Full PDF for download</a></li></ul>N-Nitrosamines are a class of carcinogenic chemical compounds that pose threat to humans exogenously through matrices like drinking water, food, cosmetics, and endogenously by nitrosation in gastrointestinal tract, nitrate to nitrite conversion, etc. An accurate, reliable analytical method is critical for detection and quantification of these trace-level, polar and low molecular weight compounds to understand the association between their exposure and related diseases.&nbsp;An optimized SQUAD workflow is presented that discovers unknown compounds in the matrix by using high resolution orbitrap and a sensitive linear ion trap detector that quantifies the low intensity nitrosamines in a single run. The workflow uses multiple fragmentation types – HCD, CID and UVPD and MSn to help characterize isomeric structures.<h4>Methods</h4>Serial dilution of Nitrosamine standards were subjected to Simultaneous Quantitation and Discovery (SQUAD) analysis using <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/744">Thermo Scientific™ Vanquish™ Horizon UHPLC system</a> and the <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/561">Thermo Scientific™ Orbitrap IQ-X™ Tribrid™ mass spectrometer</a> . Nitrosamine spiked matrices of human pooled urine and human Serum were subjected to SQUAD analysis.<h4>Data Analysis&nbsp;</h4>Spectra were evaluated for nitrosamines using Thermo Scientific™ FreeStyle™ software and annotated using Thermo Scientific™ <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1757">Mass Frontier™ 8.0 software</a>. Untargeted data analysis was performed using <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1759">Compound Discoverer 3.4 software</a>. Calibration curves were generated using Thermo Scientific™ <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/77">TraceFinder™ 5.2 software</a>.<h4>Conclusions&nbsp;</h4><ul><li>Detection limits as low ppb levels ( 0.2 ppb –0.5 ppb) and quantitation limits (0.2 ppb- 5 ppb ) in presence of the matrix were achieved for all seven nitrosamines by applying parallel ion trap analysis&nbsp;</li><li>SQUAD analysis was run on Orbitrap IQ-X Tribrid MS and achieved low ppb level quantitation in ion trap and identified unknown compounds through discovery analysis in Orbitrap.&nbsp;</li><li>Employing nano LC helped achieve lower limit of quantitation (5 ppt)</li></ul><h3>4. Waters Corporation: Validated LC-MS/MS Method for Vitamin E Acetate in Cannabis Plants and Derived Products</h3><ul><li>Application note</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/labrulez-bucket-strapi-h3hsga3/720009082_en_c7a6fc6190.pdf">Full PDF for download</a></li></ul><h5>Benefits&nbsp;</h5><ul><li>The validated LC-MS/MS method provides high sensitivity and specificity for VEA quantification in complex cannabis matrices, including flower and concentrates.&nbsp;</li><li>Minimal sample preparation reduces handling time and improves laboratory throughput, supporting efficient routine analysis.&nbsp;</li><li>The method demonstrates robust performance across multiple matrices, ensuring reliable detection even in lipid-rich, chemically diverse samples.</li><li>Use of Waters ACQUITY UPLC H-Class System and Xevo TQ-S micro Mass Spectrometer enables accurate quantitation well below regulatory requirements.&nbsp;</li><li>The validated method complies with Missouri DCR and DHSS regulatory limits, enabling laboratories to confidently monitor VEA in cannabis products.</li></ul>VEA is used as a diluent and viscosity modifier in certain cannabis-derived products, including vaping formulations. Following increased public health awareness during 2019, VEA became a marker of interest for quality control and regulatory surveillance across cannabis supply chains due to its potential impact on lung health. In addition to concentrates and vape liquids, trace residues may be present in infused products and processing intermediates. Therefore, reliable quantification of VEA in these chemically diverse, lipid-rich matrices requires an analytical method that is sensitive, selective, and robust against matrix effects. In February 2024, Missouri’s DCR revised the tolerance limit for VEA in cannabis products from 0.2 ppm to 5 ppm.1&nbsp;Liquid chromatography coupled with tandem quadrupole mass spectrometry (LC-MS/MS) provides the sensitivity and specificity required to distinguish VEA from endogenous cannabinoids, terpenes, and other lipophilic excipients, while also offering the sensitivity to support low-ppm and sub-ppm action limits. Unlike UVbased methods, where co-eluting chromophores may interfere, LC-MS/MS enables targeted multiple reaction monitoring (MRM) for confident identification and accurate quantitation in complex cannabis matrices.&nbsp;This application note describes the validation of an LC-MS/MS method for the quantitation of VEA in two inhalable products, cannabis flower and cannabis concentrates. The method was developed and validated using the Waters ACQUITY UPLC H-Class System coupled with a Xevo TQ-S micro Triple Quadrupole Mass Spectrometer. Chromatographic separation was achieved with a reversed-phase ACQUITY UPLC BEH C18 Column. A streamlined sample preparation approach was applied for cannabis flowers and concentrates, which minimizes handling time and increases sample throughput.&nbsp;The validation study was conducted at GPA. Method performance was evaluated for calibration range, linearity, LOD, LOQ, accuracy, precision, recovery, repeatability, and robustness. The results of these are evaluated against acceptance criteria set by State of Missouri DHSS and presented in this application note.<h4>Experimental&nbsp;</h4><ul><li>LC system: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/198">ACQUITY UPLC H-Class Plus System&nbsp;</a></li><li>MS detection: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/209">Xevo TQ-S micro Mass Spectrometer&nbsp;</a></li><li>Column: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1952">ACQUITY UPLC BEH C18 Column</a>, 2.1 x 50 mm, 1.7 µm (p/n: 186002350)</li><li>Software: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/635">MassLynx™ 4.1 Software&nbsp;</a></li><li>Data processing: <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?mainauthorItems=5&amp;search=TargetLynx">TargetLynx™</a> Application</li></ul><h4>Conclusion&nbsp;</h4>The LC-MS/MS method developed and validated for VEA quantification in cannabis flower and concentrates demonstrates excellent linearity, accuracy, precision, and robustness, and meets the regulatory requirements. Calibration curves achieved R² values greater than 0.999, with LOD and LOQ well below the Missouri action limit of 5 ppm. Matrix-spiked recoveries across flower and concentrate samples were within the 70–130% acceptance range, and QC samples met all performance criteria, confirming method reliability. Robustness testing across analysts and matrices further validated the method’s reproducibility. Overall, this approach provides a streamlined, sensitive, and regulatory-compliant workflow for laboratories, enabling accurate monitoring of VEA to support quality control, consumer safety, and regulatory compliance in cannabis-derived products

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