Test Menus of Colorimeter

The application of colorimeter in the following substances:

  • Iron in Water
  • Sugar in Blood
  • Phosphates
  • Phosphorus in Plant Material
  • Chromium in Aluminium Alloys

Measurement of Iron in Water

Equipment: (1) Caliber 8mm portable colorimeter; (2) Platinum basin; (3) Volumetric apparatus Reagents

0.4m hydrochloric acidStandard stock iron solution (1mg/ml).
Working standard iron solutions (0- 4ug/ml of iron). Prepare 0.0, 0.8, 1.6, 2.4, 3.2, and 4.0ug/mL solutions in 0.4m HCl.Phenanthroline solution. Dissolve 0.25g of 1,10 phenanthroline hydrate in 100ml of 25% v/v ethanol.
Quinol solution (1%).Tri-sodium citrate solution (8%).
Concentrated HCL1m HCl.

ProcedureTransfer 100ml sample into a platinum basin. Evaporate to dryness. Moisten ash with 5 drops of concentrated HCl. Evaporate to dryness in the water bath. Add 4ml of 1m HCl. Warm. Transfer solution to 10ml centrifuge tube. Dilute to 10ml. If necessary, centrifuge and retain supernatant solution for iron determination. Carry out a blank determination.

Preparation of standard graph

Pipette 5ml of each iron standard into a 25ml flask. Add 1ml quinol solution. Mix. Add 3ml phenanthrene solution. Mix. Add 5ml sodium citrate solution. Dilute to 25ml and allow you to stand for 4 hours. Measure the absorbance in a 40mm cell at 510nm. Construct a graph of absorbance vs. iron concentration.

Sample measurement

Transfer 1ml sample solution into a 25 ml flask. Dilute to 5ml with 0.4M HCI. Add quinoa, phenanthrene, sodium citrate as for standards. Measure in 40mm cell at 510nm.

Calculation of results

Read iron concentration from calibration graph. Allow for any sample dilution factor in calculation.

Sugar in Blood

Equipment: (1) Colorimeter; (2) 10mm test tubes and holder; (3) Volumetric equipment;

Reagent preparation

  • Copper sulfate (dissolve 13g in one liter of water).
  • Mixing reagent: Dissolve 50g sodium bicarbonate in 700ml water. Stir in 50g anhydrous sodium bicarbonate. Add a solution of sodium potassium tartrate in 100ml water. Wash mixture into 1 liter flask. Makeup to the mark with water.
  • Copper reagent (mix equal volumes of A and B daily).
  • Acid Reagent: Dissolve 35g molybdic acid + 5g sodium tungstate in 250ml 2M sodium hydroxide. Boil for 30 minutes. Add 100ml water + 125ml 89% phosphoric acid. Make up to 50ml.
  • Mix 320ml 3% sodium sulphate and 30ml 7% copper sulphate solutions.
  • Sodium tungstate solution (10g/100ml).
  • Glucose stock standard: Dissolve 100mg anhydrous glucose in 0.3% benzoic acid solution. Makeup to 100ml with water.

Sample procedure

Pipette 0.05ml whole blood into 1.85ml isotonic sodium sulfate copper sulfate solution (soln 5) in a test tube. Add 0.1ml sodium tungstate (soln 6). Shake well. Filter. To 0.5ml filtrate add 0.5ml copper reagent (soln 3). Boil for 10 minutes. Cool.

Phosphate Determination

The estimation of orthophosphates is carried out by adding acid ammonium molybdate to a dilute phosphate solution. The yellow phopho-molybdate formed is converted to a blue complex by the addition of a reducing agent such as amino-napthol sulphuric acid.

Equipment: (1) ATO pocket colorimeter; (2) Volumetric equipment;

Reagent Preparation

Ammonium Molybdates, Acid Solution: Add 310ml concentrated sulphuric acid to 400ml water, cool. Add a solution of 50g ammonium molybdate in 200ml water. Makeup to 1 liter. Amino Naphthol Sulphuric Acid Solution: Mix 0.125g of 1, 2,4 amino-naphthol sulphuric acid with 44ml of 15% sodium bisulfite solution in a dark bottle. Add a 20 sodium sulfite solution, dropwise until the solution is clear. Store the stock solution in a dark bottle, well stoppered, and make it freshly every two weeks. About 5 to 10ml will be required for each determination.

Sample procedure

  • The intensity of color produced is in proportion to the standing time and to the temperature of the reagents.
  • Interference will be given by the presence of chromium, iron and arsenate.
  • Estimation of poly-phosphates may be carried out by converting them to orthophosphates.

Standard Preparation

  • Prepare 10, 20, 30, and 40ppm standard phosphate solutions.
  • To 50ml of standards, add 10ml molybdate solution and 4ml amino-naphthol sulphuric acid solution.
  • Make up to 100ml in flask.
  • Allow to stand for exactly ten minutes.

Phosphorous in Plant Material

Equipment: (1) Colorimeter; (2) Volumetric equipment;

Reagents

  • Ammonium molybdate – ammonium metavanadate reagent.

Add 25g ammonium molybdate and 1.25g ammonium metavanadate to 300ml water. Warm to dissolve. Cool. Dilute to 500ml. Filter if necessary.

  • 2 5M HCl.
  • Stock phosphorus solution (1mg/ml). Dissolve 0.879g dry potassium dihydrogen orthophosphate in water. Add 1ml concentrated HCl. Dilute to 200ml. Add 1 drop toluene to this solution.
  • Standard solutions (0 – 50 µg/ml Phosphorus). Prepare daily, solutions containing 0, 10, 20, 30, 40 and 50µg/mL.
  • Distilled water.

Chromium in Aluminium Alloys

The method is based upon the measurement of the violet colour formed between chromate ion and diphenylcarbazide. The sample is dissolved in aqua region, oxidised with ammonium persulphate and the manganese etc. precipitated by making alkaline with sodium hydroxide. The chromium is oxidised to chromate with sodium peroxide. An aliquot is then coloured with sulphuric acid and diphenylcarbazide. The colour density varies with the concentration of sulphuric acid, so this reagent addition should be precise.

Equipment: (1) Colorimeter; (2) Volumetric equipment;

Reagents

Ammonium PersulphateSodium ChlorideSodium Peroxide
Sodium HydroxideSulphuric AcidDiphenylcarbazide

Sample procedure

  • Dissolve 0.5g drillings in 15ml nitric acid, hydrochloric acid, water (1:3:1 ratio) and 2.5mL concentrated sulphuric acid.
  • Boil till fumes of sulphur trioxide are liberated.
  • Add 100ml distilled water. Boil to dissolve the sulphates.
  • Oxidise with a few drops of nitric acid. Boil off any nitrous fumes.
  • Dilute to 150ml, cool, Add 15g ammonium persulphate. Boll for 10 minutes.
  • Add 1g sodium chloride. Boil to ensure complete reduction of manganese.
  • Cool and carefully add 25ml of 30% NaOH. Boil for 5 minutes.
  • Add 1g sodium peroxide. Boil for 10 minutes. Cool in running water.
  • Filter, washing with distilled water.
  • Transfer to 200ml volumetric flask. Dilute to the mark.
  • Pipette 20ml aliquot into 100ml flask. Add 8ml of 25% sulphuric acid. Dilute to 80-90ml. Add 4ml diphenyl-carbazide solution.
  • Dilute to 100ml.
Handheld Color Meter

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