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Quantitation by Extraction

Paper Type: Free Essay Subject: Chemistry
Wordcount: 1703 words Published: 22nd Jan 2018

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  • Syed Masood Hassan Akbari


A liquid-liquid extraction phase consists of a component (a) which is to be removed from the feed phase. The addition of a second phase (b) a solvent phase which is immiscible with the feed phase but the component (a) is soluble in both the feed phase and the solvent phase. As it happens the solute (a) is transferred from the feed phase to the solvent phase. After extraction the feed and solvent phases are called the raffinate and the extract phases respectively.

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Usually one of the two phases is an organic phase while the other is an aqueous phase. Under equilibrium conditions the distribution of the solute (a) over the two phases is determined by the distribution law. After the extraction the two phases can be segregated because of their immiscibility. Component (a) is then separated from the extract phase by distillation and the solvent is then regenerated. Further extractions may be carried out to remove more of the component (a).

Once the product of interest has been extracted finally an absorbance reading can be found at its corresponding λmax.

Figure 1: Shows the chemical structure of trimethoprim.

Figure 2: Shows the chemical structure of sulfamethoxazole.

This experiment consisted of three main components:

  • The quantification of trimethoprim and sulfamethoxazole in a suspension of Septrin.
  • Interpreting the data gathered and using it in calculating the content of trimethoprim in Septrin.
  • Interpreting the data gathered and using it in calculating the content of sulfamethoxazole in Septrin.


Extraction of trimethoprim and sulfamethoxazole from SEPTRIN.

Figure 3: Illustrates a schematic of the entire extraction procedure consisting of trimethoprim and sulfamethoxazole in SEPTRIN oral solution.

Sulfamethoxazole (Standard, Sample and Blank prep)

Figure 4: Illustrates sulfamethoxazole’s sample preparation.

The standard was prepared in the exact manner in a 250ml volumetric flask with using 0.2g of sulfamethoxazole in 50ml 0.1M NaOH and topping up with water. The procedure to follow was the exact same as the sample prep.

The blank was prepared in the same manner and water was used along with the reagents outlined above in sample prep.

Trimethoprim (Sample and Blank prep)

The already extracted bottom layer with chloroform was extracted again using four 50ml quantities of 1M acetic acid and the top layers were combined and washed with 5ml chloroform and the top layer added to a 250ml volumetric flask and filled with 1M acetic acid to the mark. 10ml of this solution was then added to another 100ml volumetric flask and 10ml of 1M acetic acid was also added and topped up with water.

The blank was made by using 0.2M acetic acid solution.


Determining the quantity of trimethoprim and sulfamethoxazole in an oral suspension of SEPTRIN.

Trimethoprim ƛmax 271nm

Sulfamethoxazole ƛmax 538nm












Table 1: Shows the absorbance values obtained at each respective ƛmax for trimethoprim and sulfamethoxazole.



The sample had a dilution factor of 1:1000 which meant that there were two dilutions carried out where the suspension was diluted to 250ml, which is a 1:100 dilution. Secondly the sample was diluted too 100ml, which is a 1:10 dilution. These two dilutions steps combined gave a 1:1000 dilution factor. The calculations were carried out as below.

2.5ml in 250ml

1:100 (Dilution ratio)

1:10 (10ml in 100ml)

Therefore 1:1000 dilution factor.

Absorbance: 0.332 x 1000 = 332

A1%/1cm=204 so 1% solution has 204 absorbance

1%conc = 204

X = 332

332(1)/204 = x

X= 1.627% w/v It complies with the compliance limit of (1.44-1.76)% w/v.


0.2g in 250ml

200mg in 250ml (1:100 dilution ratio)

0.8mg in 1ml (1:50 dilution)

0.016mg in 1ml (1:10 dilution)

0.0016mg in 1ml

Sample of sulfamethoxazole absorbance = 0.075

So, 0.0016mg in 1ml = 0.293

And, x = 0.025

0.025×0.0016/0.293 = x

X = 0.0001365mg in 1ml

0.0001365mg in 1 ml x 50000 = 6.826mg in 1ml

6.826mg in 1ml  0.06826g in 1ml  6.826g in 100ml

X = 6.826% w/v It does not comply with the compliance limit of (7.4-8.6)% w/v.

Trimethoprim and Sulfamethoxazole

The concentration of trimethoprim and sulfamethoxazole in 5ml of SEPTRIN were calculated as shown below.

Trimethoprim: 1.627%  1.627g in 100ml

So, x in 5ml

100x = 8.135g

X = 0.08135g in 5ml SEPTRIN

Sulfamethoxazole: 6.826%  6.826g in 100ml

So, x in 5ml

100x = 34.13g

X = 0.3413g in 5ml SEPTRIN


The active ingredients in SEPTRIN, trimethoprim 1.627% w/v and sulfamethoxazole 6.826% w/v were segregated by extraction. The concentrations of trimethoprim and sulfamethoxazole in the 2.5ml suspension were calculated to be as stated above respectively. The content of trimethoprim fell within the compliance limit but for the sulfamethoxazole the content fell a bit short of the compliance limit suggesting apparatus malfunction or human error in making up sample. Trimethoprim complies with the limit set out on the BP but for sulfamethoxazole it falls under the limit stated on the BP.


  1. Illustrate the extraction procedure.
  • Support the separating funnel in a ring clamp on a ring stand. Make sure the stopcock of the funnel is closed.
  • Place a stemmed funnel in the neck of the separating funnel. Add the liquid to be extracted, then add the extraction solvent. The total volume in the separating funnel should not be greater than 75% of the funnel volume.
  • Insert a stopper in the neck of the separating funnel.
  • Pick up the separating funnel with the stopper in place and the stopcock closed, and shake it once gently.
  • Point the stem away from your face and slowly open the stopcock to release some build-up of excess pressure, reclose the stopcock and repeat this procedure until only a small amount of pressure is released with it is vented.
  • Shake the funnel vigorously for a minute, release the pressure and then again repeat the step of shaking a few times till there is less pressure and the solutes are at equilibrium between the two solvents.
  • Place the funnel back into the ring and let it rest undisturbed until the layers are clearly segregated. While waiting, remove the stopper and place a beaker under the separating funnel.
  • Carefully open the stopcock and allow the lower layer to drain into the beaker. Drain just to the point that the upper liquid barely reaches the stopcock.
  • The top layer can be left in the separating funnel if further extractions need to be carried out which do need to be carried out. When finished with the separating funnel store the stopper away from the funnel.
  1. Explain why sulfamethoxazole is soluble in the alkaline aqueous solution and trimethoprim is not?

It is already known that trimethoprim is classified as a benzylpyrimidine and it protentiates the antimicrobial activity of sulphonamides eg., sulfamethoxazole. So trimethoprim has a tendency to form insoluble complexes when combined with sulfamethoxazole in alkaline aqueous media.

  1. Write out the chemical reaction which leads to the coloured product.

The reaction mechanism above shows the coloured dyeing agent N-(naphthalene-1-yl)ethane-1,2-diamine reacting with the diazonium to form the coloured product.

  1. Why is ammonium sulphanate added? What would happen if it was forgotten?

AMS is used as a reagent to speed up the chemical reaction if it is not included in the reaction before adding the dying agent then the process will either delay in reaching to its completion or not reach it at all.

  1. After extraction of sulfamethoxazole, how is the organic solvent processed?

The organic solvent was extracted in four 50ml quantities of 1 M acetic acid. The upper layers were all combined and kept and washed with 5ml of chloroform. The top layer was then dispensed into a 250ml volumetric flask and topped up with 1 M acetic acid. 10ml of this solution was transferred to a 100ml volumetric flask, 10ml of 1 M acetic acid was also added and the flask was topped up with water. A blank was made and the sample from the organic layer was read at 271nm.


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