I am getting a much better handle on processing spectral data on my software, so I decided to completely redesign my cuvette holder(it is 90 percent inclosed within the holder,) so I could contain as much reflected fluorescent light back into the cuvette, and direct it twoards the slit with more efficiency.

Rhodamine B sample preparation was done as follows:

1) a 100ml flask was placed on a digital scale and tared, 1g of rhodamine b was measured into flask and filled to the 100ml mark with distilled water and then shaken lightly. I then placed the flask on a magnetic stirrer on high for 2hrs until all the rhodamine b was dissolved.

Using a graduated disposable pipette, I then transferred 1ml of rhodamine b to another 100ml flask and filled to the 100ml mark and swirled it several times, this is the 100ppm dilution.

Then I transferred 5ml from the second flask containing the 100ppm dilution to a 3rd 100ml flask and filled to the mark and swirled several times.

This is the 100ppb dilution.

Now I have my 2 samples marked S1-100ppm and S2-100ppb

Reference sample is distilled water.

3 quartz cuvettes were used/ cleaned with Isopropyl alcohol and rinsed with distilled water.

Light source is a 532nm green laser.

S1- 100ppm rhodamine b before data processing

Reference(distilled water in quartz cuvette) and Sample

Subtraction of Ref and Sample 100ppm

Zoomed view of subtraction data

Excitation and emission graph for rhodamine b/100ppm@532nm

Next set is rhodamine b 100ppb:

Rhodamine b 100ppb before data processing

Ref(distilled water in quartz cuvette) and Sample

Subtraction of Ref and Sample rhodamine b 100ppb

Zoomed view subtraction of ref and sample rhodamine b 100ppb

Excitation and emission wavelength of rhodamine b 100ppb@532nm

Rhodamine B sample used

Samples ready for testing

Newly redesigned cuvette holder and laser operation illuminating sample.

Conclusions:

The 2 samples of rhodamine b although differing in solution concentration, appear to both have peak wavelengths of 586nm, predicted peaks vary because of regents used (usually NaOH, predicted peaks are usually 565nm to 573nm) in this case I used distilled water with no Ph adjustment which could account for the higher peak wavelengths.

So experimentally within the parameters that I have set, the peaks for rhodamine b in distilled water with no Ph adjustment appear to be @586nm.