CHMI 2227 - E.R. Gauthier, Ph.D. 1 CHMI 2227E Biochemistry I Refresher: - acid-base chemistry - spectrophotometry.

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Presentation transcript:

CHMI E.R. Gauthier, Ph.D. 1 CHMI 2227E Biochemistry I Refresher: - acid-base chemistry - spectrophotometry

CHMI E.R. Gauthier, Ph.D.2 Acid base chemistry - strong acids HCl  H + + Cl - pH = - log [H + ] Where [H + ] is in molar (M) concentration units.

CHMI E.R. Gauthier, Ph.D.3 Acid base chemistry - weak acids Ka = [H + ] x [A - ] [HA] pKa = -log Ka Ka = 1 10 pKa HA H + + A -

CHMI E.R. Gauthier, Ph.D.4 Acid base chemistry - weak acids pH = pKa + log [A - ] [HA] Ka = [H + ] x [A - ] [HA] pH = - log [H + ]

CHMI E.R. Gauthier, Ph.D.5 Acid base chemistry - weak acids: titration pH NaOH Mid-equivalence point: - 50% HA - 50% A pH at mid-equivalence point = pKa pKa HA H + + A - 100% HA 100% A -

CHMI E.R. Gauthier, Ph.D.6 Acid base chemistry - weak acids Example 1: What is the pH of a 0.1 M solution of acetic acid (Ka = 1.76 x M).

CHMI E.R. Gauthier, Ph.D.7 Acid base chemistry - weak acids Example 2: What will be the pH of a solution made of 0.3 M acetic acid and 0.1 M acetate (pKa = 4.8).

CHMI E.R. Gauthier, Ph.D.8 Spectrophotometry Incident light Transmitted light Incident light Transmitted light Detector Cuvette Light source

CHMI E.R. Gauthier, Ph.D.9 Spectrophotometry

CHMI E.R. Gauthier, Ph.D.10 Spectrophotometry Absorbance intensity Wavelength (nm)

CHMI E.R. Gauthier, Ph.D.11 Spectrophotometry

CHMI E.R. Gauthier, Ph.D.12 Spectrophotometry - Beer-Lambert equation -logT = log (1/T) =  cl =A Where: T = transmittance A = absorbance l = light path (cuvette size)(cm) c= concentration of analyte (M)  = absorption coefficient (M -1 cm -1 )

CHMI E.R. Gauthier, Ph.D.13 Standard curves Absorbance intensity Concentration(units) Absorbance of the unknown Concentration of the unknown Non-linear part of the curve. BEWARE!!!

CHMI E.R. Gauthier, Ph.D.14 Spectrophotometry