[2] ASSAY OF VANILLIN USING UV-VIS SPECTROSCOPY

VANILLIN

Primary component of the vanilla (Vanilla planifolia) bean extract

Synthetic vanillin is sometimes used as

a flavoring agent in foods (ethylvanillin), beverages and pharmaceuticals

VANILLIN

appears as

white or very slightly yellow needles with characteristic aromatic odor

VANILLIN

properties

antioxidant & mutagenic

SPECTROSCOPY

Interaction between matter & electromagnetic radiation

ABSORPTION SPECTROPHOTOMETRY

Measurement of an interaction between electromagnetic radiation and the molecules, or atoms, of a chemical substance

UV

(190-380nm

VIS

(380-780nm)

near-IR

(780-3000nm

IR

(2.5-40 mcm)

Absorbance

measure of the amount of light absorbed by a sample as a beam of light passes through it

Transmittance

amount of light transmitted by a sample as a beam of light passes through it

• Reflectance

measures the amount of light that reflects from the surface of a sample

ULTRAVIOLET-VISIBLE (UV-VIS) SPECTROMETER

an optical system capable of producing monochromatic radiation in the range of 200–780 nm and as a device capable of detecting the optical transmittance, usually expressed in absorbance (A), whose primary function is to measure the stated absorbance/transmittance at defined wavelength(s)

ULTRAVIOLET-VISIBLE (UV-VIS) SPECTROMETER

range

monochromatic radiation of 200–780 nm

ULTRAVIOLET-VISIBLE (UV-VIS) SPECTROMETER

capable of detecting

optical transmittance (absorbance (A))

ULTRAVIOLET-VISIBLE (UV-VIS) SPECTROMETER

primary function

measure the stated absorbance/transmittance at defined wavelength(s)

APPLICATION OF UV-VIS SPECTROSCOPY

Quantitative analysis:

Determining the concentration of a drug substance in a sample using Beer's law, which relates absorbance to concentration

APPLICATION OF UV-VIS SPECTROSCOPY

Qualitative analysis:

Identifying unknown compounds based on their characteristic UV-Vis spectra

APPLICATION OF UV-VIS SPECTROSCOPY

Kinetic studies:

Monitoring chemical reactions in real time by measuring changes in absorbance over time

APPLICATION OF UV-VIS SPECTROSCOPY

Protein and nucleic acid analysis:

Studying biomolecules by examining their absorption spectra.

USP Reference Standard

• Used as reference substance in compendial spectrometric tests and assays

• Helps ensure measurement under identical conditions for the test specimen and the reference substance

Peak Spectral Absorbance

max absorption

“Similar preparation” or “Similar solution”

Expressions used in tests and assays involving spectrometry that indicate that the reference comparator (USP RS) should be prepared in an identical manner for all practical purposes to that used for the test specimen

“Concomitantly determine” or “Similar solution”

Expressions used in tests and assays involving spectrometry that indicate that the absorbances of both the solutions for test specimen and reference specimen, relative to the specified test blank, must be measured in immediate succession

Instrumental conditions

mode

uv

Instrumental conditions

analytical wavelength

308 nm

Instrumental conditions

cell

1 cm

formula