Pharma Analysis

Discuss various pumping systems, columns & detectors in HPLC

Q.5. (a) Discuss various pumping systems, columns & detectors in HPLC.  Ans.5. (a) pumping system in HPLC Analytical Pumps (Solvent Delivery System) The requirements for HPLC pumps are as follows: They must be able to generate high pressures, have a pulse free output, deliver flow rates ranging from 0.1 to 10 ml/min, have flow reproducibility’s of 0.5% relative or better and they must be resistant to corrosion by a variety of solvents. Various types of pumping systems are

Discuss the theory of Mass spectroscopy

Q.4. (a) Discuss the theory of Mass spectroscopy

Ans.4.(a) Theory: Mass spectrometry is the most accurate method for determining the molecular mass of the compound and its elemental composition. In this technique, molecules are bombarded with a beam of energetic electrons. The molecules are ionized and broken up into many fragments, some of which are positive ions. Each kind of ion has a particular ratio of mass to charge, i.e. m/e ratio (value). For most ions, the charge is one and thus, m/e ratio is simply the molecular mass of the ion.  

Application in study of macromolecules

Q.3. (b)     Explain spin labeling & Hyperfine splitting in ESR spectrometry and its application in study of macromolecules
Ans.3.(b) Spin labeling: Site-directed spin labeling (SDSL) is a technique for investigating the structure and local dynamics of proteins using electron spin resonance. The theory of SDSL is based on the specific reaction of spin labels with amino acids. A spin label's built-in protein structure can be detected by EPR spectroscopy. SDSL is also a useful tool in examinations of the protein folding process. A spin label (SL) is an organic molecule which possesses an unpaired electron, usually on a nitrogen atom, and the ability to bind to another molecule. Spin labels are normally used as tools for probing proteins or biological membrane-local dynamics using EPR spectroscopy. The site-directed spin labeling (SDSL) technique allows one to monitor a specific region within a protein. In protein structure examinations, amino acid-specific SLs can be used. The goal of spin labeling is somewhat similar to that of isotopic substitution in NMR spectroscopy. There one replaces an atom lacking a nuclear spin (and so is NMR-silent) with an isotope having a spin I≠0 (and so is NMR-active).  

Explain the significance of magnitude of “J” constant in structure elucidations.

Q.3. (b) Explain the significance of magnitude of “J” constant in structure elucidations.
Ans.3. (b) Coupling constant: The distance between the centers of the two adjacent peaks in a multiplet is usually constant and is usually called the coupling constant. The value of the coupling constant is independent of the external field. It is measured in Hertz (Hz) or in cps (cycles per second). It is denoted by the letter J.