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Biochemistry II - Seminars

Biochemistry II - Seminars. Jiří Dostál Department of Biochemistry, Fac. Med., MU Brno jrdostal @med.muni.cz. Literature. Printed handouts with questions – will be given before seminar Seminar ppt files with answers – uploaded into IS after seminar

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Biochemistry II - Seminars

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  1. Biochemistry II - Seminars Jiří Dostál Department of Biochemistry, Fac. Med., MU Brno jrdostal@med.muni.cz

  2. Literature • Printed handouts with questions – will be given before seminar • Seminar ppt files with answers – uploaded into IS after seminar Spring 2008 VSBC041s Biochemistry II-s Study materials  Work with study materials • Laboratory manual: Biochemistry II – Practicals 2008 • Lectures – ppt files available at is.muni.cz (VSBC04p) • Textbooks: R. K. Murray et al.: Harper´s Illustrated Biochemistry. P. C. Champe, R. A. Harvey: Biochemistry.

  3. Conditions for the credit: see the back side of the syllabus !! • 100% attendance • If you are absent – written elaboration of the chapter must be given to teacher ASAP • at least 70 % from three revision tests

  4. Optical and electrophoretic methods in clinical chemistry Seminar No. 1

  5. Spectrophotometry

  6. Q. 1

  7. A. 1 • 180 – 400 nm UV • 400 – 800 nm VIS

  8. Q. 2

  9. A. 2 • A = εc l • Conditions: • Monochromatic light • Diluted solution (< 0.01 mol/l) • Homogeneous solution • Monomeric substances which do not exhibit fluorescence

  10. Q. 3

  11. A. 3 • Molar absorption coefficient • l/mol. cm

  12. Q. 4

  13. Q. 5

  14. A. 5 • Only coloured compounds • Colourless compounds (e.g. glucose, cholesterol) have to be converted to coloured derivative by chemical reaction

  15. glucoseoxidase glucose + O2 gluconolactone + H2O2 H2O2 + H2A  2 H2O + A colourless colourless colourless colourless peroxidase colourless chromogen coloured product (absorbance is measured) colourless colourless

  16. Q. 6

  17. A. 6 • Solution which contains everything except the compound to be analyzed • Solvent (water) + reagent + (buffer) ......

  18. Q. 7

  19. A. 7 • c = A / εl • c = A × calibration factor • c = A cST / AST obtaned as the slope of calibration curve

  20. Q. 8

  21. A. 8 T = 10 % = 10 × 1/100 = 0.1  A = -log 0.1 = 1 T = 50 % = 0.5  A = 0.3 T = 100 % = 1  A = 0 A = log 1/T = - log T

  22. Q. 9

  23. A. 9 c = A / ε = 0.54 / 13900 = 0.0000388 mol/l = 0.039 mmol/l = 39 μmol/l

  24. Q. 10

  25. A. 10 ε = A / c = 0.805 / 0.0005 = 1610 l/mol.cm

  26. Q. 11

  27. A. 11 Absorbance is additive quantity Absorbances of sample and standard have to be corrected by the absorbance of reagent ASTD = 0.39 - 0.1 = 0.29 ................. 5 mmol/l Asample = 0.54 - 0.1 = 0.44 ................ x mmol/l x = (0.44 × 5) / 0.29 = 7.58 mmol/l

  28. Q. 12

  29. A. 12 0,50 ................. 5 mmol/l 0,75 .................. x X = (0,75 × 5) / 0,50 = 7.5 mmol/l

  30. Q. 13

  31. A. 13 Common feature: Light interacts with a colloidal solution of proteins Light is scattered on particles, absorption is minimal Intensity of scattered ligth (I) is measured Difference: Turbidimetry – I measured in the same direction Nephelometry – I measured in perperdicular direction (90°)

  32. Q. 14

  33. A. 14 • The best scatter of light is when the wavelength is close to the size of dispersed particles • Consider red ligth (500 nm) in fog

  34. Electrophoresis

  35. Q. 15

  36. A. 15 • The pH of solution (buffer) • The nature of protein – the ratio of acidic an basic AA

  37. Q. 16

  38. A. 16

  39. Q. 17

  40. A. 17 • Elevated alfa-2 and beta globulins • Contain proteins of acute phase – indicators of acute inflammation

  41. Q. 19

  42. Q. 20

  43. A. 20 • Casein is the main milk protein • Phosphoprotein – rather acidic – pI = 4.5 • At pH 8.6 it becomes polyanion, goes to + electrode

  44. Q. 21 pH = 9.7, when pH = pKA 50 % dissociation !!!

  45. + ...Glu .........Ile...... Lys ................. - start

  46. Q. 23

  47. A. 23 a thiol is a reducing agent reduces disulfide bonds to separate polypetide chains

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