Biochem exam notes 生化习题.docVIP

Spectrometry (1) Spectrometry: is a qualitative and quantitative technology based on emission spectra, absorption spectra or scattering spectra of the material. Types of spectrometry: Based on the character of emission spectra: Atomic emission spectrometry, Flame photometry, Fluorescence spectrometry Based on absorption spectra: UV Spectrophotometry, Visible Spectrophotometry **Lambert-Beer’s Law: When a monochromatic light passes through an absorbing solution, absorbance of solution is directly proportional to the concentration of the absorbing substance and the depth of the solution through which the light passes. Equation for Lambert-Beers law: A=lg I/T=KCL, where K is absorption coefficient, C is the concentration of the substance absorbing light, L is the length of the path of light. A is dimensionless. L is in centimeters and the concentration of ‘C’=1mol/L. Determination of protein concentration by Lowry Method: EXP. What are the colorimetric methods used to determine the concentration of serum protein? Describe briefly the advantages and disadvantages of each method. There are two methods: standard addition method and calibration curve method. Ploting a standard curve of absorbance vs protein concentration of standards. (2) Using the Lowry Method: Serum protein concentration(g/L)= Asample/AStandard x C standard x Time of dilution Advantages of plotting a standard curve graph: Very easy to obtain concentration of serum protein in a sample, easy to plot, time efficient. Easy to understand graph. No chemical formulas required to be memorized and calculated. Able to determine concentration of other samples because the standard curve has been plotted. Disadvantages: Graph is limited to small samples. Room for the human error as the graph maybe plotted wrongly, the determination of the protein concentration is not very accurate. Advantages of using formula to calculate serum protein concentration: Time efficient without needin