Friday, May 27, 2011

IB Biology IA, IB Chemistry IA, EE Biology, EE Chemistry

IB Biology IA , IB Chemistry IA, EE Biology and EE Chemistry 
IA and EE on Vitamin C quantification 


1. Ammonium Oxalate, prepared by dissolving 0.08g ammonium oxalate in 100ml pH 5 buffer
2. Vit C (0.001M ), prepared by dissolving 0.018g of Vit C in 100ml ammonium oxalate solution
3. Perform 2 fold dilution on 0.001M Vit C.
4. Add 1 ml Vit C into a quartz cuvette and 2ml of ammonium oxalate ( as a stabiliser )
5. Leave for 10 mins
6. Prepare a blank with 1ml water and 2 ml ammonium oxalate
7. Set up UV spectrophotometer at 266nm and quantify.
Result shown below
Latest journal published last month, accurate way to quantify Vit C. 
Click HERE to get in
* Oxalate solution used as  stabiliser and pH around 5 as both affect the stability of Vitamin  C
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IA and EE on Glucose Quantification
1. 1% glucose std was prepared ( 1g in 100ml )
2. Perform 2 fold serial dilution ( 1%, 0.5%, 0.25%, 0.125% )
3. Add 2 ml of 1% glucose + 2 ml of DNS (3,5 dinitrosalicylic acid) into a 50ml test tube.
4. Repeat step 1 to 3 for different glucose concentration
5. Place all tubes in boiling water bath for 5 mins for colour formation. ( red brown)



6. Transfer 1 ml of solution from tube to cuvette and added 2ml of water to dilute it
7. Set visible spectrophotometer at 487nm.
8. Prepare a blank containing 1ml DNS + 2ml water.




9. Measure absorbance at 487nm
10. Plot std calibration curve
11. Beer's Law works only for diluted solution.
12. Recommended range for glucose ( 1% to 0.01%)
13.Expt will not work if too little glucose is used




Video,Glucose Quantification using DNS with visible spec


Click HERE on glucose quantification and preparation of DNS solution.
* Resulting Glucose/DNS solution must be diluted with water as Beer's Law only applies for diluted solution.
*Experimental Kit can be purchased  for EE and IA work. Click HERE to purchase kit
* Glucose solution range must be around 1% to 0.1% for colour determination.
* All item can be purchased from Carolina Biologicals or Kemtecscience

Sample on IA and EE on glucose quantification using DNS and Visible Spec

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IA and EE on Iron Quantification
Preparation
1. 0.01M iron(II), prepared by dissolving 3.92g of Fe(SO4)2(NH4)2.6H2O in 2M HCI to 1liter.
2. Perform 2 fold serial dilution.
4. Add 1 ml of iron (II) std to cuvette
5. Add 0.5ml 5% trisodium citrate/buffer to cuvette( use pH <6)
6. Add 0.5ml 10% hydroxyammonium chloride (reducing agent  to reduced all iron (III ). Ignore this step if you think iron (II) is not oxidised.
7. Add 1ml 0.01M phenanthroline for colour formation.
8. Leave for 24 hours for iron (II) phenanthroline complex to form ( orange red )
9. Set visible spectrophotometer at 508nm
10.Prepare a blank ( 1 ml water, 0.5ml trisodium citrate, 0.5ml hydroxyammonium chloride, 1ml phenanthroline )
10. Measured absorbance for  iron(II) std in cuvette shown above

Result on iron quantification

2 fold dilution from 0.0001M was done, absorbance was measured after 24 hours.




Std Calibration curve for iron(II) standard at 508nm.
Click HERE, sample EE on iron quantification with visible spec
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IA and EE on Quantification for starch iodine solution 
Starch with iodine solution forms blue black colouration.
1. Prepare 0.01% starch solution ( 0.01g in 100ml )
2. Perform a 2 fold dilution ( 0.01, 0.005, 0.00250, 0.00125, 0.000625%)
3. Add 1ml 0.01% starch to cuvette
4. Add 50ul 1% iodine solution ( adjust volume accordingly to obtain appropriate blue black)
5. Set visible spec at 567nm
6. Prepare a blank ( 1ml water + 50ul iodine ) and calibrate/zeroize it
7. Prepare cuvette (1ml 0.01%starch + 50ul iodine) and measure absorbance at 567nm
8. Repeat step 7 with different starch conc and plot std calibration curve.
Hints and tips 
  • Starch iodine must be diluted with water as Beer's Law, works for diluted solution.
  • Starch solution decomposes over time (Prepare fresh)
  • Volume of 1% Iodine must be adjusted accordingly so a blue black is formed.



Video, Starch Iodine quantification using Visible Spec


IA, digestion of starch using amylase measured using visible spec

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EE and IA on DNA Quantification
1. Prepare 5% DNA by adding 5ul of lambda DNA (stock 0.1mg) to 100ul TE ( DNA buffer ) into microcentrifuge 
2. Perform 2 fold dilution. ( 5, 2.5, 1.25, 0.625, 0.3125%)
3. Add 100ul DNA solution into a quartz cuvette
4. Set UV spectrophotometer at 260nm
5. Prepare blank ( 100ul TE buffer )
6. Measure absorbance at 260 and plot DNA std calibration curve at 260nm

Useful for EE in Biology DNA studies


Click HERE, view EE on DNA quantification

DNA std calibration curve at 260nm.
DNA and TE buffer can be purchased from Carolina








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Suggested IA and EE that can be done by IB Bio/IB Chem
For Vitamin C,
  • EE on the effect of UV radiation (A,B,C ) on the decomposition of Vitamin C quantified using UV spectrophotometer.
  • EE/IA on effect of heat, temperature, pH, presence of transition metal ions on the decomposition of Vit C measured using UV spectrophotometer/iodine titration method
For Glucose Quantification,
  • EE/IA on how changing pH, temperature, inhibitors, ionic salt conc affect the amylase activity measured using visible spectrophotometer. ( by either monitoring the decrease in absorbance of starch/iodine solution over time or by measuring the absorbance of starch/iodine solution after a period of time using the std calibration curve)
For Iron Quantification
  • EE/IA on how temperature affect the amount of iron content in spinach, seaweed, brocolli measured using visible spec
  • EE on the relationship between different part of plants with the amount of iron content measured using visible spec
For DNA Quantification
  • EE on how temperature affect the stability of DNA double strand ( breaking of hydrogen bond ) measured using UV spec
  • EE on how different extraction techniques, using SDS, Proteinase K, Protease, Phenol Chloroform affect the amount of DNA/purity of DNA measured using UV spec
For Starch Quantification
  • EE/IA on how changing pH, temperature, inhibitors, ionic salt conc affect the rate of digestion of amylase on starch/iodine solution measured using visible spec ( by either monitoring the decrease in absorbance of starch/iodine over time or by measuring the absorbance of starch/iodine over a period of time using std calibration curve)
List goes on...............endless Research Questions. 
Most importantly we must establish a sound methodology and a reliable way to quantify.

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