The Application Notebook
Thermo Fisher Scientific Application Note
Terry Zhang and Guifeng Jiang, Thermo Fisher Scientific, San Jose, California, USA.
Carbonyl compounds from motor vehicle and industrial emissions, precursors to ground-level ozone, are also sources of pollution in indoor living and working environments.
HPLC coupled with UV detection is the most widely recognized technique for the analysis of carbonyl-DNPH derivatives.
LC separations were performed on a Thermo Scientific Accela 1250 UHPLC system which offers the flexibility of performing both HPLC and UHPLC separations on a single platform. The Accela 1250 Pump delivers precise flows and accurate gradients at an expansive range of flow rates (up to 2 mL/min) and pressures (up to 1250 bar); and accelerates method development and maximizes method flexibility through quaternary gradient capabilities. The Accela UHPLC system together with Thermo Scientific 1.9 μm Hypersil GOLD C18 columns enables fast chromatographic separations with high efficiency and resolution.
The use of sub-2 μm particle columns facilitates rapid analysis of challenging samples by improving chromatographic resolution, speed and sensitivity. Using the Accela 1250 UHPLC system, a single Hypersil GOLD C18 column (1.9 μm, 2.1 × 100 mm) and a simple acetonitrile/water gradient, a mixture of the DNPH standards of 12 carbonyls targeted by US EPA Method 8315A Procedure 1 was successfully separated. This analysis was performed using a flow rate of 800 μL/minute, which generated back pressures over 1000 bar. The Accela 1250 pump is the only commercially available LC platform that is capable of handling such high operational pressures due to its very low internal back pressures.
Figure 1 demonstrates UHPLC separation of the 15 carbonyl-DNPH derivatives using the Hypersil GOLD C18 column and a 13-minute gradient. All compounds but the tolualdehyde isomers were baseline resolved.
Figure 1: UHPLC separation of 15 carbonyl-DNPH derivatives at 20 μg/mL concentrations using a Hypersil GOLD C18 column and 13-minute gradient.
Excellent linearity in detector response was observed over the range of 98–50000 ng/mL (ppb) [196–100000 ng/mL (ppb) for m- and p-tolualdehyde combined], with correlation coefficients greater than 0.999 for all analytes.
Reproducibility was investigated by analysing five replicate injections of each analyte. With four channel mixings of the solvents at various viscosities, retention time RSDs ranged from 0.52–2.22% while peak area RSDs ranged from 0.46–4.91%, indicating excellent method reproducibility, particularly of the LC pump.
Quantitative accuracy for all carbonyl-DNPH derivatives was evaluated at two levels of concentrations, 400 ppb and 2000 ppb, using external calibration method. The accuracy of two representative analytes, benzaldehyde-DNPH and o-tolualdehyde-DNPH are reflected by the values of 96.3% and 103.6% at 400 ppb, respectively, and 99.8% and 99.9% at 2000 ppb, respectively and achieved with the UHPLC method.
Thermo Fisher Scientific Inc.
355 River Oaks Parkway, San Jose, California 95134, USA
tel. +1 561 688 8900 fax +1 608 273 6880
E-mail: analyze@thermoscientific.com
Website: www.thermoscientific.com
LC–MS/MS Quantification of Plasma Proteins in Elephant Seals to Advance Wildlife Conservation
November 12th 2024Liquid chromatography and tandem mass spectrometry (LC–MS/MS) was used to show that repeated stress can cause delayed but sustained changes in blood plasma proteins associated with water conservation, immune responses, fat metabolism, insulin sensitivity, iron recycling, and hormone transport in elephant seals. Some of these changes may be novel markers of recent and chronic stress exposure in marine mammals.
AI and GenAI Applications to Help Optimize Purification and Yield of Antibodies From Plasma
October 31st 2024Deriving antibodies from plasma products involves several steps, typically starting from the collection of plasma and ending with the purification of the desired antibodies. These are: plasma collection; plasma pooling; fractionation; antibody purification; concentration and formulation; quality control; and packaging and storage. This process results in a purified antibody product that can be used for therapeutic purposes, diagnostic tests, or research. Each step is critical to ensure the safety, efficacy, and quality of the final product. Applications of AI/GenAI in many of these steps can significantly help in the optimization of purification and yield of the desired antibodies. Some specific use-cases are: selecting and optimizing plasma units for optimized plasma pooling; GenAI solution for enterprise search on internal knowledge portal; analysing and optimizing production batch profitability, inventory, yields; monitoring production batch key performance indicators for outlier identification; monitoring production equipment to predict maintenance events; and reducing quality control laboratory testing turnaround time.