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5.4 Alpha D

How to measure the Alpha D of a product:

1. Turn on the machine approx 30 min before taking first reading.
Lamp should be set to ‘Na’.
2. Decide on solvent to take measurement:
Usually CHCl3 is used as the default solvent (use HPLC grade solvent) or in some cases MeOH (may depend on solubility of substrate). If you are comparing with a known value you should use the same solvent as is reported.
3. Record a blank reading:
(a) Wash the cell out twice with solvent you are going to use to record the rotation to ensure no material remains from the previous user. There is no need to dry the cell.
(b) Fill the cell with solvent ensuring that the full length of the cell is filled. Place stoppers in cell.
(c) Place cell in the machine, close lid, allow the reading to settle and then press zero
(d) Empty the cell and dry gently with heat gun (the central part of the cell will become opaque). Allow to cool to room temperature before placing the sample.

4. Prepare the sample:
Try to use about 10mg/mL of substrate. Note: if you are comparing with a known value use the same quantity of material (if sufficient is available).

(a) If the product can easily be manipulated/transferred place ~20 mg in a special 2 mL vial. (note! Whatever quantity you use you must record exactly how much you have weighed out).

(b) Fill the vial to the mark with the solvent to be used and ensure the product is completely dissolved.

5. Fill the cell:
With a 1 mL syringe transfer the dissolved sample to the cell ensuring that the complete length is filled and no air bubbles are present (place the needle touching the bottom of the cell and very slowly inject the sample – if an air bubble is present withdraw the solvent with syringe and start again). Finally place the plastic stoppers in the cell.

6. Record rotation:
(a) Place the cell in the machine and close the door.
(b) Allow the machine to equilibrate to a steady value (about 15 seconds).
(c) Record this value- remember to record the sign of the reading as well (+ or -)
(d) Continue recording readings every 5 seconds approx until you have  about 20 readings.
(e) Calculate the average reading.
 
7. Calculate the alpha D value of the compound.  


7. How to calculate the alpha D value of the compound: 
 


(a) Calculate the concentration of your sample:

 Mass in grams used in 1 mL × 100


(b) Calculate the alpha D value using the following equation:


(c) To quote the alpha in a publication use the following format:

Example - Following steps 1to 6 above:

20 mg of product in 2 mL of CHCl3 gave an average of  +0.345 over 20 readings.

10 mg in 1 mL = 0.01g


Would be written as:

c = 1.0 (CHCl3)

Then the [α]D is 34.5 (1).

 

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