Dr. Martina Kotthaus, describing the different types of reference standards used by the pharmaceutical industry.

<strong>Types of pharmaceutical reference standard (RS)</strong>

In the pharmaceutical area, there are four major types of reference standards:
<ul>
 	<li>Primary RS</li>
 	<li>Pharmacopoeial RS (for monograph use)</li>
 	<li>Secondary RS</li>
 	<li>Impurity RS</li>
</ul>
Apart from the pharmacopoeial standards, all of the above come with certificates of analysis (CoAs), or – in the case of a secondary RS – with a comparison statement. The information provided on the CoA should be suitable with regard to the specific use planned for the corresponding RS.

There is a fifth type – research materials – which is often used at the very beginning of analytical research and development, but not normally for method development, validation, transfer or quality control.

<strong>Primary reference standards</strong>

In the pharmaceutical context, a primary reference standard is a standard for which the properties (usually identity, very often also purity/assay values) have been characterized by certain analytical techniques, without, however, being compared to any other standard of the same kind.

The definition for a primary standard in the GMP guideline Q7 of the ICH states:

<em>Reference Standard, Primary</em>

<em>A substance that has been shown by an extensive set of analytical tests to be authentic material that should be of high purity. This standard can be: (1) obtained from an officially recognized source, or (2) prepared by independent synthesis, or (3) obtained from existing production material of high purity, or (4) prepared by further purification of existing production material. </em>

<strong>When to use a primary standard?</strong>

In pharmaceutical QC, the use of reference standards to calibrate the analytical procedure is mandatory when measurements are performed with relative methods such as HPLC in combination with a UV or MS detector. These measurements need to be traceable to a primary standard. This requirement is realized either by using the primary standard directly for the calibration purposes, or by using a secondary standard which is compared to the primary one.

<strong>ICH guideline Q7 states:</strong>

<em>11.17 Primary reference standards should be obtained as appropriate for the manufacture of APIs. The source of each primary reference standard should be documented. Records should be maintained of each primary reference standard’s storage and use in accordance with the supplier’s recommendations. Primary reference standards obtained from an officially recognized source are normally used without testing if stored under conditions consistent with the supplier’s recommendations.</em>

Officially recognized sources, however, are not specified in Q7, but the FDA does mention sources in their ‘Guidance for Industry on Analytical Procedures and Methods Validation for Drugs and Biologics’ but, interestingly, does not refer to these institutions as an official or definitive list:

<em>Reference standards can often be obtained from the USP and may also be available through the European Pharmacopoeia, Japanese Pharmacopoeia, World Health Organization, or National Institute of Standards and Technology. </em>

Instead, the FDA states that <em>“reference materials from other sources should be characterized by procedures including routine and beyond routine release testing” and that producers “should consider orthogonal methods for reference material characterization”. </em>

For primary RSs, both the ICH guideline and FDA guidance allow other sources than the “officially recognized sources”. Independent manufacturers can provide such primary standards, ideally characterized by processes like those outlined in the general text 5.12. of the European Pharmacopoeia (Ph.Eur).

<strong>Correct use of primary reference standards: what to keep in mind?</strong>

In essence, a primary RS needs to be fit for its intended purpose. A pharmacopoeial RS has been shown to be fit for its compendial purpose, but has not been demonstrated to be fit for any other purpose; this needs to be proven by the user. Consequently challenges of compendial standard use for non-compendial purposes have been reported in regulatory inspections. Other primary standards with fully documented CoAs can be used for most applications, providing they have been characterized appropriately. If a primary RS is used to establish a secondary standard then the secondary RS can only be used for the same purpose as the primary one.

<strong>Pharmacopoeial reference standards</strong>

Reference standards from the pharmacopoeias – also often referred to as compendial RSs – are, in principle, primary standards that have a special status for use in connection with the monograph methods for which they have been designed.

This position is specified in the USP (United States Pharmacopeia) general chapter &lt;11&gt;:

<em>When approved as suitable for use as comparison standards … in the United States Pharmacopeia (USP) or National Formulary (NF), USP RS also assume official status and legal recognition in the United States. Assessment of the suitability for use in other applications rests with the user.</em>

It is also described in the European Pharmacopoeia (Ph.Eur.) in general text 5.12.6:

<em>European Pharmacopoeia reference standards are shown to be suitable for their intended purpose; they are not necessarily suitable for other purposes. Any value assigned to a reference standard is valid for the intended use and not necessarily for other uses.</em>

So, strictly speaking, the special status of a pharmacopoeial RS can only be assumed when the standards are used as required by the monographs. For anything other than these monograph purposes, compendial RSs stand on the same level as RSs from other sources. This means that you need to prove suitability of a compendial RS for the desired non-compendial purpose in the same way you would need to for any other RSs. This is clearly mentioned in the Ph.Eur. chapter 5.12., and can be seen as a blueprint for other compendial RS use as well:

<em>If a European Pharmacopoeia reference standard is to be used for any purpose other than that for which it has been established, its suitability for the new use has to be fully demonstrated and when applicable, to be described in the marketing authorization application.</em>

The use of a compendial RS for non-monograph applications is also made difficult by the fact that certificates of analysis, which might otherwise provide a rich data set of characterization testing results, are normally not provided by the compendial institutions.

<strong>Impurity reference standard (IRS)</strong>

Although the term ‘primary impurity standard’ does not really exist, any impurity standard that is not compared to another material of the same chemical structure and which is fit for its designated purpose can be considered such a primary material.

IRSs are designed to detect, identify, quantify and qualify impurities in a drug substance in accordance with the ICH guidelines (Q3A to Q3D and Q7). ICH Guidelines 3A to 3D regulate the approach to impurities in the pharmaceutical industry. ICH does not prioritize at any point any given range of standards over another, whether officially recognized or not, for use within the pharmaceutical industry.  According to these guidelines, impurities can be primarily classified into either drug substance or drug product impurities, referring in the first case to any component in the drug substance which is not the chemical entity itself. A drug product impurity, meanwhile, is any chemical entity present that is not the drug substance or an excipient used to manufacture the drug product. Based on the cause triggering their appearance, impurities can be classified as starting materials, by-products, intermediates, degradation products or reagents/ligands and catalysts. Finally, by their chemical nature, impurities can be classified as organic, inorganic or solvents.

Pharmacopoeial IRSs, like all compendial materials, are designed for the specific monograph purposes. For other purposes, an IRS from other sources suitable for the intended purpose should be preferred, for two major reasons:
<ul>
 	<li>The use of a pharmacopoeial IRS outside the monograph can be challenged by regulatory authorities. Customers have reported to us that authorities have insisted on a CoA for a pharmacopoeial IRS used for non-compendial purposes. A CoA is not provided by pharmacopoeial sources, rendering the material unsuitable for use without separate detailed internal characterization.</li>
 	<li>Furthermore, supply of pharmacopoeial IRSs can be insecure. For example, there have been occasions where a change in a monograph has resulted in a neat impurity standard being replaced by a system suitability or peak identification mixture, usually consisting of a mix of the API and traces of one or more impurities. Such a replacement is not a problem if the user works according to the monograph methods. However, if the pharmacopoeial material is used for non-compendial purposes, the laboratory might face a challenge replacing the neat IRS if it is not available anymore. If the pharmaceutical company has stated the use of that specific material in their dossier, they may even need to report the change to the authorities. Cessation of supply is also a risk with any catalogue reference standard provided by a third party, but switching between suppliers is significantly easier when the materials are “primary” and accompanied by CoAs.</li>
</ul>
In addition, the current regulation situation – especially for the finished dosage forms dealt with in ICH guideline Q3B – often requires additional impurity testing, on top of the requirements of the pharmacopoeial monographs. Then it can often be the case that there are no suitable impurity standards available from compendial sources, and the support of experienced commercial IRS manufacturers thus becomes necessary.

<strong>Secondary standards</strong>

These compounds are second-line materials. They consist in each instance of a material that is compared against the primary material, and used in its place.

It does not matter whether the secondary standard is compared against a pharmacopoeial primary standard, or against a primary standard obtained in-house or from a third source. A secondary standard can only be used for the same purposes as the primary standard. Thus if a primary standard was designed solely for a qualitative purpose (i.e. identification via IR, system suitability test or peak identification), then to use the corresponding secondary standard for quantitative purposes is not valid. For example, a large number of Ph.Eur. reference standards for APIs have been set up for IR comparisons only, and should not be used as a basis for quantitative secondary standards.

In addition, the EDQM (European Directorate for the Quality of Medicines) does not recommend measuring secondary standards against even their quantitative materials. This is because:
<ul>
 	<li>While pharmacopoeial primary RSs have a determined measurement uncertainty (MU) in line with the requirements of the relevant pharmacopoeial monograph, the value of this uncertainty is not currently specified.</li>
 	<li>The comparison of a candidate material for a secondary RS to the pharmacopoeial RS will result – by the principle of uncertainty propagation – in a larger but non quantifiable MU for the secondary standard, by virtue of the fact that the MU for the pharmacopoeial material is not published</li>
 	<li>This unknown but enlarged MU for the secondary RS might no longer be appropriate for the acceptance interval specified in the relevant monograph.</li>
</ul>
For the same reason, the use of a pharmacopoeial RS with a non-compendial method can be difficult to justify on the basis that the uncertainty of its value assignment is unknown.

<strong>Research materials</strong>

Often used at the very beginning of analytical research and development, these should not be employed when the methods are fully developed. During validation and implementation in the QC or stability testing lab, they are usually replaced with better-characterized materials, especially when any quantitative purpose is connected with their usages.

Research materials come with only basic characterization data that makes them useful for initial development and identification work. They cannot be considered reference standards and can be distinguished from reference standards by their relatively simple CoA.

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For further details on Mikromol reference standards,

<strong><u>Please contact: </u></strong>

Jane Firth at LGC UK on <a href="mailto:jane.firth@lgcgroup.com">jane.firth@lgcgroup.com</a> or

Suryakanth.G at LGC India – <a href="mailto:Suryakanth.gudgunti@lgcgroup.com">Suryakanth.gudgunti@lgcgroup.com</a>