Data Collection

When collecting data, keep everything well organized and labeled. Think about

the experiments you will be performing. What data will need to be recorded? Set

up tables for yourself to keep the data organized.

Additional Considerations

Think back to last week’s measurements. We measured the absorbance spectrum

and created a calibration curve for a single dye.

One complication when two dyes are present is that the absorbances from the

dyes can overlap. Thus, we need to correct for the overlap since the absorbance

at a wavelength of maximum absorbance can come from two sources (each dye).

Each dye will have its own wavelength of maximum absorbance, but then it may

also have some absorbance at the maximum of the other dye and interfere.

The easiest way to correct for this is

to look at the absorbance spectra of

the pure dyes by themselves to find

the relationships between the two

wavelengths. For example, Yellow

Dye 5 has a maximum absorbance

near 425 nm and Red Dye 40 has a

maximum absorbance near 505 nm.

Unfortunately, Yellow Dye 5 also has

a small absorbance at 505 nm and

Red Dye 40 has an absorbance at

425 nm.

If both dyes are present, then an absorbance at 425 nm (A₄₂₅) comes from two

sources: yellow dye (A_Y5@425)  and red dye(A_R40@425). The absorbance at 505 nm

(A₅₀₅) will also come from two sources: red dye (A_R40@505) and yellow dye (A_Y5@505).

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Figure 9.2: Absorbance spectra of Red Dye No. 40

highlighting important analysis locations. The raw data

is available as a supplementary resource.