Basic Questions Regarding All Analytical & Instrumental Methods (p 17-18)

1. Basic Questions Regarding All Analytical & Instrumental Methods (p 17-18) • What accuracy and precision are required? • How much sample do I have available, and how much money do we have available for the analysis?  • What concentration is the analyte present at and can we pre-concentrate or dilute the sample? • What interferences might be present and can we eliminate or mask them? • What are the properties of the sample matrix? • How much time do I have?

2. Table 1-3 p 19 NUMERICAL CRITERIA FOR COMPARING ANALYTICAL METHODS 1) Precision (StdDev, RSD, etc) 2) Bias (systematic error) 3) Sensitivity (DL, LOD, MDL, etc) 4) Detection Limit (blank + 3 x StdDev of blank) 5) Dynamic Range (LOQ  LOL: linearity) 6) Selectivity (coefficient of selectivity)

3. EVERY ANALYSIS YOU PERFORM WITH AN INSTRUMENT WILL BE A COMPETITION: THE COMPETITION BETWEEN SIGNAL AND SELECTIVITY! There is no way to maximize both. You have to choose some happy medium, where you get enough signal to detect the analyte, but can also be selective enough so that you are certain of what you are detecting.

5. CALIBRATION OF INSTRUMENTAL METHODS • Calibration Curve • Standard Addition Method • Internal Standard Method

6. Calibration Curve • Known concentrations of analyte standards are prepared and measured over suspected range of analyte • Blank is subtracted manually or auto by instrument • Graph: Corrected signal vs. concentration of standards • Least-squares line fit to generate equation • e.g., spectrophotometric protein analysis

8. CALIBRATION OF INSTRUMENTAL METHODS • Calibration Curve • Standard Addition Method • Internal Standard Method

9. Standard Addition Method • Appropriate when sample composition is unknown or complex and affects signal • e.g., calibration curves for perchlorate • Known quantities of analyte are added (“spiked”) to the unknown • From increase in signal, quantity of analyte in original unknown is determined.

12. Fig 1-10

13. CALIBRATION OF INSTRUMENTAL METHODS • Calibration Curve • Standard Addition Method • Internal Standard Method

14. Internal Standard Method • A standard different from analyte is added in constant amount to all samples, blanks, and standards • Useful when quantity of samples varies from run to run • Calibration plot: • e.g., Li used as internal standard for determination of Na or K in blood serum by ion chromatography vs.

15. Ion chromatographic separation of unknown (X) and internal standard (S)

16. SIGNALS AND NOISEChap 5 p 110-113 All analytical measurements include: • Analytical Signal (V or I proportional to amount of analyte) • Background (instrument response when no analyte is present; i.e., “baseline”) • Noise (random fluctuations) Signal-to-Noise Ratio (S/N):

17. SOURCES OF NOISE • Chemical Noise: (fluctuations in T, P, humidity, light, etc) • Instrumental Noise: (from all instrumental components) • Thermal (heated components, i.e., resistors and conductors) • Shot (non-homogeneous current flow) • Flicker (1/f dependent; results in “drift”) • Environmental (stray light, 60 Hz, EMI, RF, vibrations, etc)