PH calibration buffers

The pH adjustment step of MPA might also pose potential problems. The pH calibration buffers often contain a preservative that might contaminate MPA if the pH electrode is dipped during pH adjustment (see Figure 5.14). This problem can be eliminated by pouring out small aliquots into separate containers to check pH. 

It is critical that pH electrodes are calibrated often as the slope of their response curve will change with temperature, as defined by the Nernst equation [2], Calibration should be performed with at least two certified pH calibration buffers that bracket the expected pH of the sample. Calibration is performed as described in the manufacturer s instructions provided with the pH meter. Calibration should be checked by measuring the pH of the first calibration buffer. The result should be no more than 0.02 pH units different from the certified pH of the buffer after adjusted for temperature. pH measurements are best performed when calibration buffers and samples are held at a constant temperature. 

The substrates used for lipase assay were tributyrin (C4 0), tricaproin (C6 0), tricaprylin (C8 0), tricaprin (C10 0), tri-laurin (C12 0) from Sigma. Sodium caseinate (Alanate 185) was supplied by the N.Z. Dairy Board, L-a-ledthin was from Sigma, color key pH calibration buffers were from BDH Chemicals and Q-sepharose was from Pharmacia LKB. 

Commercially available buffer solutions can be purchased for virtually any desired pH. A buffer solution commonly used to calibrate pH meters contains 0.025 m Na2HP04(aq) and 0.025 M KH2P04(aq) and has pH = 6.87 at 25°C. However, the method demonstrated in Example 11.1 would give pH = 7.2 for this solution. Because these calculations interpret activities as molarities, not effective molarities, they ignore ion—ion interactions so the values calculated are onl) approximate. 

Figure 3. Calibration curve for serial dilution of gasoline in pH=7 buffered water at an instrument-to-analyte distance of 25 m, using laboratory apparatus.

Measurements were made by first calibrating the pH probe using two buffer solutions at pH s of 7 and 10 supplied by Van Labs of Van Waters and Rogers Equipment Company. These solutions have been certified by the National Bureau of Standards to be accurate to + 0.01 pH unit at 25°C. This pH calibration was made with the probe inserted in buffer solution at the same temperature as the pH measurements were made. 

The use of buffers for the calibration of a pH electrode is a critical step in any pH measurement. Buffer solutions of different pH values can be prepared or are commercially available. They should be traceable to national standards with 0.01... 

E° [equation (15.4)] is also referred to as the offset, the zero potential point, or the isopotential point, since theoretically it is defined as the pH that has no temperature dependence. Most pH electrode manufacturers design their isopotential point to be 0 mV at pH 7 to correspond with the temperature software in most pH meters. The offset potential is often displayed after calibration as an indication of electrode performance. Typical readings should be about 0 30 mV in a pH 7 buffer. In reality, E° is composed of several single potentials, each of which has a slight temperature coefficient. These potentials are sources of error in temperature compensation algorithms. 

The difference in the conductivity of the calibration buffers and sample can cause a very large error on the sample measurement, due to junction potentials in different environments. Solid samples should be dissolved in purified water. It is necessary that the water be carbon dioxide-free. The presence of dissolved carbon dioxide will cause significant bias in the measurement of samples with low buffering capacity. For pH measurements with an accuracy of 0.01 to 0.1 pH unit, the limiting factor is often the electrochemical system (i.e., the characteristics of the electrodes and the solution in which they are immersed). 

When you calibrate an electrode with standard buffers, you measure a voltage with the electrode in each buffer. The pH of buffer SI is pHs, and the measured electrode potential in this buffer is Es>- The pH of buffer S2 is pHS2 and the measured electrode potential is s2-The equation of the line through the two standard points is... 

Littman 19) developed a continuous automatic oxidant recorder based on the use of 20% potassium iodide in neutral (pH = 7) buffered solution, as shown in Figure 3. The calibration is based on the assumption that the iodine is released in stoichiometric proportion to the amount of ozone according to the equation... 

The instrument (see Figure 27-4) is designed so that a manually or microprocessor actuated valve (V) admits calibrator gases, standard buffers, or a sample to a small chamber (C) maintained by a fluid or metal bath (B) to a constant temperature of 37 C 0.1 C, Measuring and reference electrodes (E) protrude into this chamber. In the pH calibration phase of the instrument, high pH standard buffer and low pH standard buffer are alternately admitted into the chamber and electronic responses of the upper and lower limits of a linear pH curve are established. In the gas calibration phase, gas mixtures with high and low fractional concentrations of O2 and CO2 are alternately admitted into the chamber and electronic responses of the upper and lower limits of linear PO2 and PCO2 curves are set. In the measurement phase, an anaerobically collected blood sample is admitted. 

The pH measurement system is calibrated against primary standard buffers admitted either manually or automatically into the sample chamber. The buffers are phosphate solutions that should meet National Institutes of Standards and Technology (NIST) specifications. Calibration buffers meeting NIST specifications are available from the manufacturer of an instrument, usually in containers of appropriate size and shape for mounting as a reservoir on the instrument. The pH values of the low and high calibrator buffers axe set by the manufacturer but always lie close to 6.8 and 7.4 at 37 °C. The tolerance of the specified values should be less than or equal to an SD of 0.003 to achieve SDs of 0.005 to 0.01 in measuring blood pH. Unopened containers should be stored at room temperature. When visual observation or an instrument display warns of low concentrations in the reservoir, recommended practice is to replace the reservoir with a newly opened container rather than to replenish fluid in the current one. Pooling several almost empty containers is not recommended. 

Enzyme activities are expressed generally as international units one unit is the amount of enzyme that will catalyze under optimum conditions the transformation of 1 pmol of substrate per minute. The activities are expressed as lU/L or mLU/ mL. The katal (kat nanokatal = nKat) has been proposed as an alternative unit it is defined as the catalytic activity that, under dehned conditions, converts 1 mol of substrate per second. Thus, the units are moles per second (mol/s). One lU/L corresponds to 16.67 nKat/L. Unfortunately, these dehnitions of enzyme units do not include other major variables that affect their measurement, such as the composition and pH of buffer, identity and concentration of the substrate, temperature, and the presence of activators. This has led to a variety of national and international recommendations for the common enzyme measurements, which confuse comparisons between published data. Calibration of enzyme methodologies— particularly for less common enzymes— remains a laboratory issue (Jansen et al. 2006). 

Values of pH as a function of temperature have been assigned to the same set of pH., seawater buffers,and so alternatively can be used for calibration (Tables 2, 3)... 

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