The strength of rhGH measured by RP-HPLC

The strength of blue tops from China

First I'd like to explain how the strength of Growth Hormone is measured.

First, the activity of Growth hormone was tested by using small animals, mostly rats. After much protest of organisations for animal welfare, the WHO asked some labs to seek the relationship between HPLC and the activity.  After a lot of experimenting, scientists concluded that the activity result of the RP-HPLC  (reversed phase) corresponded to the activity resulting by administration of growth hormone to small animals later these animals where killed to measure growth/strength. Then, in pharmacopoeia, the testing of activity in animals, changed to testing by RP-HPLC.

On picture 1 you'll see an analysis report how the analytical laboratory sends it. The graphic from the sample is compared with the graphic of a pharmaceutical reference standard mostly called standard on picture 3.

A pharmaceutical reference standard is small amount of very pure API (active pharmaceutical ingredient) that on the street is named "raw powder". This standard is exactly dosed en therefore used to measure the standard strength that can be used to calculate the strength of the sample. On picture 3 you'll see both the sample and the standard compared.  I cut out the data and pasted it into the graphic, for your convenience.

Important for calculation is the area of the compound in the graphic. To make that visible I colored the area of the related peptides from the blue-tops red. The RT is the retention time, in this case the peak was measured at the 29 st minute.

The area, as said colored red,  is over 13%.( 17.51%)13% is the standard of EP (European Pharmacopeia).

That means that a legitimate manufacturer couldn't sell such product.

I diluted your sample 1.6* and standard 3.2*. The result shows that the hgh of your sample is only 2.7 iu   (174442/258360 *4=2.69iu).

The purity of the product is in this case measured in three peaks. That is an old method.

In fact, the main peak of somatropine should include peak 3. The third peak  was done by the laboratory technician’s analysis costume. Long time ago, when we were developing the product, there was a very small peak at the position of peak 3, and it was considered  as related proteins or containment. Now the third peak no longer exist , but the technician still calculate the third peak  to lower the main peak percentage in order to make the purification technicians to realise they must work hard to make the product quality better. In this case the related proteïnes are 17,51 % and the purity of the somatropine is 80,12% plus 2,37 % makes a total of 82,50%.

Today the protein chromatograms just comprise two parts, related proteins and somatropine.

The electrophoresis shows some containments. Also vagely visible on the graphic of the sample at the 16th and 18th minute.

It is very difficult to translate this result to other bluetops because these vial with blue fliptops come in different versions and can be bought by anyone in China. Lets realise some manufacturers sell highly purified peptides other sell cheap crap to make more profit. We also analysed some blue-tops that where of excellent quality.