Using working standard in analytical method validation

Dear Sir,

I would like to ask you for the question:

Can I using working standard (previously determined the potency against USP reference standard) in analytical method validation for dissolution and assay. I am perform validation for dissolution and assay of Danazol capsule. But in testing accuracy I used spiking sample by weigh Danazol standard in range of 60-130% level. If I used USP reference standard in this test, I need a large of reference standard and it is problem with cost.

Is there legal requirement for using working standard or reference standard in method validation. I can not find this information in USP

Thank you very much!

Use of working standard is generally accepted as long as its properly qualified. Only testing potency against USP RS may not be enough although this will be the most critical test.

Thank you for you reply.

We are prepare working from Danazol raw materials with full test as in USP and the potency was determined against USP refoerence standard. My manager told me that in method validation, it not acceptable when using working standard (prepared from the raw material). But I can not find any requirements in legal documents about this.

I have also heard from some colleagues in industry that validation requires USP RS while some others say there is no such requirement. I guess the reason is somewhere some inspector(s) requested this and it goes around. In my opinion this is not required and most people don’t do it. A well characterised and qualified working standard should be acceptable in validation work. Full tests in USP is for routine testing of API. Working standard qualification requires more tests than the tests used for routine release testing.

We don’t use USP-purchased RS here for our topical OTC products, would be way too costly and inefficient as USP supplies only tiny amounts. We use USP certified material from its original container and store the certificates of analysis. Our QA department has given its blessing, for both R&D stability testing and for QC assays at production.

We do our cGMP validations using similar USP-grade standards.

Thanks KM for sharing your information. Could you explain little more about “USP certified material from its original container”. Is this meaning USP grade API come from the API supplier as API for manufacturing? Or a small sample of API come as material for reference standard? Original container meaning a small container or could it be a large drum in some cases? You are using supplier COA for standard purity information? Really appreciate if you could explain little more and also tell us whether your practice had any comments from a FDA inspector.

I’m in R&D and our products are topical OTC formulations, and we haven’t been blessed by an FDA inspector here, but our manufacturing facilities have been audited by FDA (multiple times), so this practice is not “a secret”. If we can purchase USP-grade material independently, we do that instead of relying on the API supplier for a standard. If material cannot be purchased cannot be purchased independently, we request a separate sample of USP-certified material of different lot number than we would be using in manufacturing (most times this is supllied for free). We get this in like a 100 gram bottle with manufacturer’s label on it, and have their certificate of analysis on file.

One cannot use the same lot of API as standard as was used in its manufacturing. We don’t approve the use a sample of the API from a different drum to use as a reference standard, as then our company becomes responsible for the acquisition of the sample and the labeling/tracking of that material (would be “our” label).

Thanks KM for sharing your information. I think they make sense and fairly defendable. I can see the logic and agree with you. Just one thing if you could explain. Why you say “One cannot use the same lot of API as standard as was used in its manufacturing”. I can understand selecting the best batch or highest purity batch. But it could be the same batch you happened to use in your development or later work. Hope you can clarify.

We’ve been told by our QA (not by FDA) that the reference standard for the analytical testing must be a different lot than what was used in the production of that batch of finished product.

It makes sense to me. For example: if that reference standard was pulled from a drum of API and was really only 75% active, and that drum was used to make product, then the assay results of the product would show 100% as then the 75% would cancel out in the equations.

Of course, each incoming drum of API is supposed to be assayed for purity before using, agree? But things can happen…

I think you have to go back to your bench and recoin your message.

Do you mean that you get reference standards in Drums?

Reference Standards are highly-characterized physical specimens used in testing by pharmaceutical and related industries to help ensure the identity, strength, quality, and purity of medicines (drugs, biologics, and excipients), dietary supplements, and food ingredients

All the drums are also USP grade, and certified.

You don’t think that the USP actually purifies those “official” reference standards further before they sell them, do you ???

Understand that you are in Denmark so English may be an issue, but what do you mean by “go back to your bench and recoin” the message ?

I mis understood you due to my poor english.

I have a small question from your sentence in above post : “if that reference standard was pulled from a drum of API.”

Do USP sells these reference materials in drums? What i understand is that they sell in small vials or containers with all quality lables and materials.

Please enlighten me on this issue. Your comments are valuable.

Further I apologise reagrding my english too.


My company purchases pure USP grade API by the drum (for example: triclosan, octyl methoxycinnamate) and manufactures finsihed OTC pharmaceutical products such as antibacterial soaps, sunscreens…

We do NOT use a sample pulled from such drum as our analytical standard. We either buy, or request USP grade standard from the supplier.

USP grade standard from supplier is a working standard and cannot be a surrogate to USP RS. we can use a sample as working standard as long as it is characterized properly using USP RS or other primary standard.


In Validation have to use reference standard or qualified working standard with proper documentation.
but for Recovery test you can use the drug substance or working standard since more quantity of std is required.


Dear phucto,

Your question was :“Can I using working standard (previously determined the potency against USP reference standard) in analytical method validation for dissolution and assay.” so the answer is “Yes” U can use WS if that has been qualified against a primary ref std supplied by USP.All u have to do is also attach the COA of WS u will be using with details of primary USP ref. std against which is has been qualified.

Further, If you are performing a accuracy test like for disso/ assay where you have to spike a more amount of API in placebo, u have got two options:

  1. Use your WS where u know its purity for spiking in placebo.
  2. you can use any other API(not necessarily WS)which is analysed & within all acceptance criteria of your RM release specification & for which a COA is available. But take into consideration the purity of API used for spiking while doing accuracy calculation bcause in this case the purity of API used for spiking & purity of WS used may be different.