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High Performance Liquid Chromatography Video Lecture | Crash Course for CSIR NET Life Sciences

FAQs on High Performance Liquid Chromatography Video Lecture - Crash Course for CSIR NET Life Sciences

1. What is High Performance Liquid Chromatography (HPLC) and how does it work?

Ans. High Performance Liquid Chromatography (HPLC) is an analytical technique used to separate, identify, and quantify components in a mixture. It works by passing a liquid sample through a column packed with solid adsorbent material. The components of the sample interact differently with the stationary phase and the mobile phase, leading to their separation as they travel through the column. The separated components are then detected, often using UV-Vis spectroscopy.

2. What are the main types of HPLC and their applications?

Ans. The main types of HPLC include Normal Phase HPLC and Reverse Phase HPLC. Normal Phase HPLC uses a polar stationary phase and a non-polar mobile phase, making it suitable for separating polar compounds. In contrast, Reverse Phase HPLC employs a non-polar stationary phase and a polar mobile phase, which is widely used for analyzing pharmaceuticals, biomolecules, and other complex mixtures due to its high efficiency and resolution.

3. What are the advantages of using Reverse Phase HPLC over other techniques?

Ans. Reverse Phase HPLC offers several advantages, including high sensitivity, specificity, and efficiency in separating a wide range of compounds. It allows for better retention of polar compounds and is compatible with various detection methods. Additionally, it requires less sample preparation compared to traditional methods, making it a preferred choice in pharmaceuticals, environmental testing, and biochemical analysis.

4. What factors influence the separation process in Reverse Phase HPLC?

Ans. The separation process in Reverse Phase HPLC is influenced by several factors, including the choice of stationary phase, mobile phase composition, flow rate, temperature, and sample properties. The hydrophobic interactions between the analytes and the stationary phase play a crucial role, as well as the polarity and pH of the mobile phase, which can affect the retention time and resolution of the separated components.

5. How is data interpreted after conducting an HPLC analysis?

Ans. After conducting an HPLC analysis, the data is typically interpreted using a chromatogram, which displays the retention time on the x-axis and the detector response on the y-axis. Peaks in the chromatogram represent different components within the sample. The area under each peak is proportional to the concentration of the corresponding component, allowing for quantification. Comparisons with known standards can further aid in identifying and quantifying the analytes present in the sample.

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Sep 28, 2025 Last updated

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