Ionic Calcium Determination in Skim Milk with Molecular Probes and Front-Face Fluorescence Spectroscopy: Simple Linear Regression

Fluorescence spectra of Fluo-5N (A) and Rhod-5N (B) at pH 5.8 and 7.0. All spectra were obtained with a probe concentration of 1μM and an ionic calcium concentration of 125μM in distilled water.

Change in pH upon addition of the pH 6.6 and pH 5.5 adjusted skim milk to water containing 2% 4 M KCl.

Background fluorescence spectra (excitation = 470nm, emission = 500 to 560nm) of diluted skim milk (1.9 to 8.9%).

Raw fluorescence spectra (A) and background subtracted fluorescence spectra (B) of Fluo-5N probe (1μM) in pH 7.0 diluted skim milk (1.9 to 8.9%). A one pass 5-point smoothing was performed on the background subtracted spectra. The ionic calcium concentration (μM) of each sample is indicated.

Raw fluorescence spectra (A) and background subtracted fluorescence spectra (B) of Fluo-5N probe (1μM) in pH 6.2 diluted skim milk (1.9 to 8.9%). A one pass 5-point smoothing was performed on the background subtracted spectra. The ionic calcium concentration (μM) of each sample is indicated.

Background fluorescence spectra (excitation = 549nm, and emission = 560 to 600nm) of diluted skim milk (1.9 to 8.9%).

Raw fluorescence spectra (A) and background subtracted fluorescence spectra (B) of Rhod-5N probe (1μM) in pH 7.0 diluted skim milk (1.9 to 8.9%). A one pass 5-point smoothing was performed on the background subtracted spectra. The ionic calcium concentration (μM) of each sample is indicated.

Raw fluorescence spectra (A) and background subtracted fluorescence spectra (B) of Rhod-5N (1μM) in pH 6.2 diluted skim milk (1.9 to 8.9%). A one pass 5-point smoothing was performed on the background subtracted spectra. The ionic calcium concentration (μM) of each sample is indicated.

Linear regression analysis of ionic calcium content (μM) and the Fluo-5N fluorescence intensity (520nm) of background subtracted fluorescence spectra of the pH 7.0 (A) and the pH 6.2 (B) diluted skim milk (1.9 to 8.9%).

Linear regression analysis of ionic calcium content (μM) and the Rhod-5N fluorescence intensity (577nm) of background subtracted fluorescence spectra of the pH 7.0 (A) and the pH 6.2 (B) in diluted skim milk (1.9 to 8.9%).