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CHEM 213 Instrumental Analysis. Lab Lecture – Glucose and GC. Sucrose. glucose. Glucose Autoanalyzer. enzyme. Glucose . H 2 O 2. Determination of Sucrose and Glucose. Detect H 2 O 2 electrochemically Does not detect sucrose. slow. Outline of Procedure.
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CHEM 213Instrumental Analysis Lab Lecture – Glucose and GC
Sucrose glucose Glucose Autoanalyzer enzyme Glucose H2O2 Determination of Sucrose and Glucose DetectH2O2 electrochemically Does not detect sucrose
slow Outline of Procedure • Solution prep and hydrolysis of sucrose to glucose • 1.5-2 hours • Done 1-2 weeks in advance 2. Run samples and standards on autoanalyzer Book 35 min time slot
2H+ + O2 + 2e- H2O2 Eapplied = 0.7 V Allows only H2O2 to electrode Controlled rate of diffusion of glucose to electrode Glucose oxidase -D-Glucose + O2 (g) Product + H2O2 2. Principle Current [H2O2] [glucose] Other species can be oxidized
Reactions • In the electrode, have two reactions • At enzyme layer • At electrode surface β-D-glucose + O2 ⇌ Glucono-δ-lactone + H2O2 H2O2 ⇌ 2 H+ + O2 + 2e−
Current due to solution baseline current (PL Current)
Where is this used (A LOT) ? Blood Glucose Monitor(Sec. 17-4, p357-359)
Step F Sucrose glucose -D-glucose Fructose Determination of Sucrose and Glucose
Reactions • During the week before analysis • Hydrolysis of sucrose • Establish equilibrium between α- and β-glucose
Glucose Standards PL current for flasks 1, 3, 5, 7 Step E Pg 39
PLsucrose = PL2 – PL1 Step E (Glucose + Sucrose) Standards Pg 39 For sucrose alone
helium Polar particles Benzene by Gas Chromatography volatility & surface interactions
Internal standard Sample ampoule Standard Prep Table (pg 55) 3. Quantitative Analysis (pg 1 HotTips) Measure weight
You are using internal standard calibration Response used for plotting is: Ianalyte IBenzene IInt. Std IToluene Chromatogram of Sample Label the chromatograms!!! Sample, Run #1
Chart speed = 50 4. Notes on Procedure F (pg 2 HotTips) Chromatogram of Sample
5. Notes on Calculations (pg 2 HotTips) 1. N = number of theoretical plates = 16(tR/W)2 N = 186 (unitless)
5. Notes on Calculations (pg 2 HotTips) 2. Rs = resolution = tR,2 – tR,1. ½(W1+ W2) Rs = 0.95 (unitless)
5. Notes on Calculations (pg 2 HotTips) air peak 3. k′ = capacity factor = (tR-tM)/tM k’ = 8.1 (unitless)