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Determination of pH, Ca2+ and Mg2+ in Saliva

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1 Determination of pH, Ca2+ and Mg2+ in Saliva
Cody Chalker David Julia Real World Project Fall 2009

2 Electrolytes in Saliva
Na+ K+,Ca2+ Mg2+(mM): Cu2+ Pb2+ (µM), Electrolyte Conc. affected by flow rate and disease Only bound and free Ca2+ can be measured simultaneously 10% bound to proteins, 40% to phosphates, citrates and lactates, 5% to bicarbonate; Thus approx. 45% is in ionic form

3 Bicarbonate and pH Bicarbonate a product of cell metabolism
Bicarbonate does not buffer it mainly neutralizes Bicarbonate concentration in saliva is linked to flow rate

4 Reasons for testing pH of saliva
The ability to monitor health status, disease onset and progression, and treatment outcome through noninvasive means is a highly desirable goal in health care promotion and delivery. Oral fluid is a perfect medium to be explored for health and disease surveillance.

5 Importance A healthy individual should have a saliva pH of 6.5-7.4
Most ailments such as cancer, kidney stones, heart disease, etc. are associated with an acidic pH in the body As you age the pH of your saliva changes Over half of adults are acidic at 6.5, symbolizes signs of aging i.e. calcium deficiency. Most children are around a pH of 7.5 Most cancer patients have a pH of 4.5 while in terminal condition.

6 Why Hasn’t This Been Studied Before?
One of the main reasons this is just becoming a research topic is that the amount of ions in saliva are so small that not until recently were techniques used to be able to make affirmative quantitative assumptions

7 Techniques Used Today Microfluidics and Microelectromechanical systems are MEMS used for salivary diagnostics MEMS are integrated systems composed of mechanical elements, sensors, actuators and electronics on a common silicon substrate developed through micro fabrication technology

8 Other Techniques Used Today
Electrochemical methods have been devised Thiocynate is an example of an ion being studied Process includes high performance liquid chromatography, silica gels, and ion specific electrodes Pros of procedure: that it is quick, precise, and requires few samples

9 Techniques Used pH of saliva:
Saliva was taken from volunteer who were asked to build up approx. 1-2mL of saliva which was then extracted with a disposable pipette. The saliva was then immediately immersed in paraffin oil CO2 + H2O H2CO HCO3- + H+

10 pH cont’d To finish testing the pH of saliva 8mL of water was added to each sample. The water added was boiled for an hour and found to have a pH of 6.75 pH glass-electrode probe was calibrated with different pH buffers to obtain pH values 4,7, and 10 Following the calibration the pH probe was immersed into each sample

11 Determination of Magnesium and Calcium
Determination of the two ions cannot be found exclusively through an EDTA titration since both Magnesium and Calcium complex to EDTA Volunteers were asked to collect as much spit in their mouths(approx. 5mL), sans mucus, and spit only once into the beaker 8mL of buffer-indicator was added. Standard was made of 8mL of water and 1mL of indicator. Standard was titrated with EDTA solution to determine approximate end point. Each sample was titrated to an end point

12 Determination of Magnesium
Thiazole yellow was used as an indicator Ammonium Oxalate was used to precipitate out calcium UV Vis instrument

13 Procedure for Use in UV Vis
Saliva samples were mixed with ammonium oxalate and centrifuged (suspected problem area) 0.75mL of each sample was added to cuvettes. 0.75mL additions were added of water, polyvinyl alcohol, thiazole yellow and of NaOH The UV Vis was calibrated with magnesium concentrations of .1mM, .2mM, and .3mM. Each sample was run against these concentrations

14 Results Using a glass-electrode the pH of 5 samples were found to be 7.25, 7.4, 7.45, 7.3, 7.3. EDTA titration of saliva involves small amounts of either ion concentration. To determine the concentration of the two ions a comparison is made against a standard Sample mM Mg/L .05% EDTA mL added 1 0.11 0.24 2 0.13 0.29 3 0.23 0.35 4 0.34 0.41 5 0.54 0.61 6 0.83 0.95

15 Standard Calibration Graph

16 Results

17 Results Fitted with 4th degree Polynomial

18 Magnesium Results The results for magnesium concentration were obtained via UV Vis. The experimental results proved to be elusive. Quantitative data was unable to be obtained.

19 Conclusion The results show:
An equation that describes the Mg concentration in saliva at low concentrations Can effectively determine pH of Saliva with a glass electrode pH meter What techniques are used today to determine pH, [Mg2+], and [Ca2+]. Real world application and importance.

20 References Starr, Henry. "Studies of Human Mixed Saliva". The Journal of Biological Chemistry. Philadelphia: Robert Hare Laboratory school of medicine, 1922. Wong, David. "Salivary diagnostics powered by". American Dental Association 2006: Ericsson, Yngve. "Simplified Methods for Determination of Calcium and Magnesium in the Saliva". Journal of Dental Research 1955:

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