Q.1. (a) Draw an optical diagram of spectrofluorometer, Give its theory and pharmaceutical applications.
Ans.1. (a) Optical diagram of spectrofluorometers
Theory of Spectrofluorometer: Several instrument manufacturers offer spectrofluorometers capable of providing both excitation and emission spectra. The optical design of one of these, which employs two grating monochromators, is shown in fig 1.1 radiations from the excitation monochromator is split, part passing to a reference photomultiplier and part to the sample. The resulting fluorescence radiation, after dispersion by the emission monochromator, is detected by a second photo-multiplier. An instrument such as that shown in figure provides perfectly satisfactory spectra for quantitative analysis.
The emission spectra obtained will not, however, necessarily compare well with spectra from other instruments because the output depends not only on the intensity of fluorescence but also on the characteristics of the lamp, transducer and monochromators. A number of methods have been developed for obtaining a corrected spectrum, which is the true fluorescence spectrum free from instrumental effects.
Application of Spectrofluorometers:
Determination of alpha-aminocephalosporins
A selective and highly sensitive fluorometric method was developed for the determination of four alpha-aminocephalosporins, namely cefaclor, cefadroxil, cephalexin and cephradine in biological fluids and pharmaceutical preparations. The produced derivatives exhibit maximum fluorescence intensities at 472–478 nm after excitation at 370–372 nm. The method involves the reaction of the target compounds with fluorescamine at a specific pH, ranging from 7.8 to 8.4. The method is highly specific because other alpha-aminocephalosporins whose alpha amino group was blocked do not react similarly and hence do not interfere.
Spectrofluorometric determination of ibuprofen in pharmaceutical formulations
The spectrofluorometric method is used for determination of ibuprofen in pharmaceutical tablets, creams and syrup. It involves excitation at 263 nm and emission at 288 nm. The linear range is 2-73 mg L-1.
Spectrofluorometric Analysis of New-Generation Antidepressant Drugs in Pharmaceutical Formulations, Human Urine, and Plasma Samples
Five different drugs, sertraline, paroxetine, citalopram, venlafaxine, and fluoxetine, were analyzed for spectrofluorometric detection in pharmaceutical formulations and urine and plasma samples.
Spectrofluorometric determination of nicardipine, nifedipine and isradipine in pharmaceutical preparations and biological fluids
The method is based on the reduction of nicardipine, nifedipine and isradipine with Zn/HCl and measuring the fluorescence intensity obtained (λem/λex) at 460/364, 450/393 and 446/360 nm, respectively.
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