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Application of Spectroscopy in Isolation, Purification and Identification

  • Application of Spectroscopy play a crucial role in identifying the structure, functional groups, and composition of bioactive compounds in crude extracts.

Common techniques include:

1) UV-Vis Spectroscopy

Principle

  • Measures absorption of ultraviolet or visible light, typically in conjugated systems (e.g., aromatic rings, unsaturated bonds).

Application

  • Rapid screening for characteristic chromophores (e.g., flavonoids, alkaloids).

  • Preliminary quantification of known markers.

2) Infrared (IR) Spectroscopy

Principle

  • Detects molecular vibrations, revealing functional groups (e.g., –OH, –NH, –C=O).

Application

  • Confirmation of functional groups in isolated fractions.

  • Fingerprint region (600–1500 cm⁻¹) aids compound identification.

3) Nuclear Magnetic Resonance (NMR) Spectroscopy

Principle:

  • Uses magnetic fields to analyze hydrogen (¹H NMR), carbon (¹³C NMR), and other nuclei environments.

Application:

  • Structural elucidation of purified natural products.

  • Determination of stereochemistry and molecular connectivity.


4) Mass Spectrometry (MS) (Application of Spectroscopy)

Principle

  • Ionizes molecules and separates ions based on mass-to-charge ratio (m/z).

Application

  • Molecular weight determination.

  • Identification of characteristic fragmentation patterns.

  • Used with chromatographic techniques (GC-MS, LC-MS) for rapid profiling of complex mixtures.

  • These techniques complement each other, providing a comprehensive understanding of compound identity and purity.


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