Fluorescein

Exploratory experiments

The goal is to evaluate the concentration required to have an absorbance of around 0.5

We took 14 mg of fluorescein in 100mL of water

14 mg/L

Color	Absorbance	Emission
Red	0.01
Green	0.43
Blue	1.79
UV		-1358

From the previous solution we 0.5mL and bring it to 5mL.

1.4 mg/L

Color	Absorbance	Emission
Red	-0.02
Green	0.14
Blue	0.95
UV		-2878

0.7 mg/L

Color	Absorbance	Emission
Red	0
Green	0.07
Blue	0.56
UV		2953

Exploratory Conclusions

An absorbance in blue of around 0.5 is achieved with a concentration of phenolphathlein of 0.7mg/L.

At this concentration the spectrophotometer has a fluorescence of 2953 internal unit.

CaCl2.2H2O solution

4.64 g/L

Fluorescence in presence of Ca++

Solution A: fluorescein 1.4 mg/L Solution B: CaCl2.H2O 4.64 g/L

A	B	H2O	R	G	B	UV
2	0	2	-0.01	0.10	0.53	2237
2	0.5	1.5	-0.01	0.11	0.55	2160
2	1	1	0	0.10	0.55	2193
2	1.5	0.5	-0.00	0.11	0.55	2265
2	2	0	0.00	0.10	0.55	2250

369mg of CaCl2.2H2O in 2 + 2 mL (2 experiments) (concentration of 92g/L)

A	B	H2O	R	G	B	UV
2	0	2	-0.00	0.09	0.52	2052
2	0	2	-0.00	0.11	0.53	2088

Conclusion

In presence of CaCl2 at high concentration there is no clear evidence of a fluorescence quenching.

Test with KI

Solution A: fluoresceine 1.4 mg/L Solution B: KI 10 g/L

A	B	H2O	R	G	B	UV	UV second experiment
2	0	2	0	0.07	0.56	2953	2942
2	0.5	1.5	0.01	0.07	0.58	2817	2816
2	1	1	0.00	0.08	0.56	2603	2618
2	1.5	0.5	0.00	0.07	0.57	2503	2517
2	2	0	0	0.07	0.57	2283	2243

All data

Conclusions

It seems that we observe a fluorescence quenching but other factors like influence of pH should be checked. Literature shows that this kind of experiment is done in presence of 0.1M of KOH.

Check page 12 of : Fluoroscence - Quenching