ANALO445: Separation and Identification of Food and Drug Dyes by Thin-Layer Chro

Question

ANALO445: Separation and Identification of Food and Drug Dyes by Thin-Layer Chromatography Thin-layer chromatography (TLC) is an example of solid-liquid chromatography in which a liquid mobile phase ascends, by capillary action, a thin layer of solid adsorbent coated onto a support. A drop of the sample being tested dissolved in the mobile phase is applied to the coated plate before the plate is placed into contact with the mobile phase. As the liquid mobile phase ascends the TLC plate, the components in the mixture being separated partition between the moving liquid and the stationary solid, according to their relative solubility in each phase. This separation process involves a complex series of solubility equilibria related to the polarities of the sample, the mobile phase, and the stationary phase. The components of the mixture will travel different distances, and each component will develop its own spot on the TLC plate. If the sample contains only one component, only one spot will appear on the plate. The plates that result from the TLC process are called chromatograms. Under controlled conditions, such as similar samples exposed to the same solvent, adsorbent, and temperature, a specific mixture component will move a fixed distance relative to the distance traveled by the leading edge of the mobile phase, called the solvent front. We call this ratio the retention factor, or R, of the component under the experimental conditions, as shown in Equation 1. distance traveled by component, mm (Eq. 1): distance traveled by solvent front, mm We express Ry as a decimal. The Rys for several of the FD&C; certified dyes on alumina using a mobile phase of 4:4:1:2 isoamyl alcohol, 95% ethanol, concentrated ammonia (NH,) solution, and water are shown in Table 1. We can qualitatively identify unknown compounds by comparing their Rys with Rp for known compounds determined under identical conditions. Small changes in solvent composition or in temperature can cause changes in the Rt of a compound. Additionally, for good reproducibility, the air surrounding the plate in the chamber must be saturated with mobile phase vapor. To assure this, we cover the chamber in which we prepare the chromatogram. In this experiment you will use food, mouthwash, or cough medicine samples that contain one or more FD&C; certified dyes: Red 3, Red 40, Yellow 5, and Yellow 6 (Figure 1). After you extract the dyes from the sample, you will chromatograph them on an alumina TLC plate using a mobile phase of 4:4:1:2 isoamyl alcohol, 95% ethanol, concentrated NH3 solution, and water. From your experimental measurements, you will calculate Rys for each Table 1 RF Values of Some FD&C; Certified Dyes dye Red 2 0.40 9 1904 Cengage Learning Red 3 Red 40 Yellow 5 Yellow 6 0.77 0.70 0.20 0.75 commen! "de-listed," but may be present in foods packaged before 1976 pinkish-red spot bright red spot bright yellow spot orange spot

Explanation / Answer

The Rf's for FD&C certified dyes with those in Table 1 reflects the main hazard is very close Rf values of Red 3, Red 40, and Yellow 6. These dyes have Rf values 0.77, 0.70, and 0.75 respectively. These closer Rf values can sometimes gives a false positive identification of any one of the dye. Suppose we are trying to identify and confirm Red 3 dye, due to closer Rf values and our inaccurate TLC procedure might lead to identify Red 40 as Red 3 since both have almost red color spot and vice versa. The same situation will also happen with Red 3 and Yellow 6 also, this is due to closer Rf values of red 3 & Yellow 6 and sometimes we are not aware of distinguishing between Organge and Red color spot due to closer wavelengths of these two light frequencies. Overall one should take care of when soleley dependence of Rf based identification of dyes Red 3, Red 40, and Yellow 6. It is advised to couple the final result of identification with other spectrometric methods also while concluding the final result.

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