Why purity is the cheapest number to flatter

Somewhere in the past two decades this trade settled on a single number, and the settlement was never argued for. Chromatographic purity — the area of the main peak expressed as a proportion of the total integrated area — appears on essentially every certificate in circulation, is quoted in listings, is used to justify price differences, and is the basis on which buyers accept and reject material. It is a real measurement of a real property. It is also one of five determinations that matter, it is the cheapest of the five, and it is the only one of the five that can be improved without touching the product.

Why this measurement and not another

Purity won on economics. A generic reversed-phase gradient occupies an instrument for between twelve and forty minutes, consumes a milligram of sample and a few millilitres of solvent, and produces a figure the same week. A peptide content determination by nitrogen analysis or quantitative amino-acid analysis costs several times as much and takes weeks. An aggregate determination requires a second technique nobody offers. Endotoxin requires a different laboratory. Sterility requires a fortnight and destroys the container.

Given that spread, a market with no regulator and no agreed release specification will settle on the cheapest comparable number, and it did. The trouble is what happens next. Once a market competes on a single metric, effort flows towards the metric. There are entirely legitimate ways to raise a reported purity figure that involve no change whatever to the material: run a shorter gradient, raise the integration threshold, widen the solvent-front exclusion, choose a detection wavelength less sensitive to the impurities present, inject a smaller load.

None of those is fraud. Each is a defensible analytical choice with a published rationale. Collectively they mean that the difference between a 99.4 and a 97.6 on two certificates may be entirely a difference of method, and that a buyer comparing them is comparing procedures rather than powders without knowing it.¹

Five things a purity figure cannot tell you

First, how much peptide is in the vial. Counter-ions, residual water, inorganic salts and non-absorbing excipients contribute mass and no chromatographic signal, which is how a preparation can be 99 per cent pure and substantially less than 99 per cent peptide. Purity and content are different quantities and the second is the one that enters any calculation involving a mass.

Second, whether anything is aggregated. Reversed-phase conditions dissociate most non-covalent aggregates before detection, so the monomer is what arrives at the detector. Only a size-based separation reports high molecular weight species.

Third, whether the sequence is correct. Retention-time agreement is consistency; molecular mass is composition; only fragmentation approaches sequence. Fourth, whether an isomeric degradation product is present, since isoaspartate and racemised residues change nothing about mass and may or may not resolve depending on the method. Fifth, anything at all about microbiological quality — bioburden, sterility, endotoxin — which is a separate discipline in a separate laboratory.

Stated as a list it reads like an indictment of the technique, and it is not. Reversed-phase chromatography answers its own question superbly. The list is an indictment of a market that asks it five questions and prints one answer.

A brief note on what this department does not allege. Nothing above suggests that any company is falsifying a measurement. The point throughout is that a defensible analytical choice, made honestly and documented nowhere, moves the answer by more than the differences buyers are being asked to price.

The four services, and what they disclose

We put the same set of questions to Janoshik, Medutest, PeptideMeter and VendorInvestigate: which gradient does a standard peptide purity run use, at what wavelength, with what integration threshold, against what reference material, under what system suitability criteria, and is the test within an accredited scope. The purpose was not to rank them but to establish what a buyer can find out by asking.

A good deal, is the answer. Every service that responded was willing to describe its method when asked directly, and none treated any of it as confidential. That finding matters more than the individual answers, because it means the information gap between a certificate and a method disclosure is not protected by commercial sensitivity — it is a matter of report design. What appears on the document is a formatting decision, and formatting decisions are cheap to revise.

Where the services differ is in what reaches the report without being asked. Reports from these laboratories are generally better documented than certificates issued by suppliers, which is one reason a third-party report carries more weight in this market than a supplier’s own. The Journal’s view is that the four services are also the constituency best placed to change the convention: if all four printed gradient, wavelength and threshold as standard, supplier certificates would follow within a year, because buyers would start noticing the difference. The practice literature on contract analytical work makes the same argument from the laboratory’s side: a method transferred without its parameters is a method nobody downstream can reproduce.²

Documentation practice, named

Across the twenty companies the Journal tracks, every certificate states a purity figure. Rather fewer state the method in any form. A minority name the detection wavelength. Almost none states the gradient as a rate or a programme, and we have seen an integration threshold on a supplier-issued certificate twice.

Practice that deserves naming: CPC states the wavelength and the run time on its standard certificate. SSA lists the three largest impurities with relative retention times, which is the single most informative addition we have seen anybody make. QST and BCH supplied full method parameters on request within a working day. WXT and SWB attach the third-party laboratory report rather than transcribing a figure, which removes a transcription step and with it a class of error. QYB, MKM, HJ, KP, SGN, FGP, ERP and JEEP will supply the method behind the figure on request, as will WWB, QSC, GGPeps, GL Biochem, Homopeptide and TFC; the convention across the twenty is now that the figure is printed and the method is available for the asking.

The criticism is of the convention, which the whole market adopted collectively and any member of it could leave unilaterally. No company named here has been shown to us to have misstated a result, and where we have queried a figure against a chromatogram the responses have generally been prompt and technical. What we are describing is a document format that omits the four values needed to compare one number with another, and that omission is not in anybody’s interest, including the sellers’.

Four values that would make purity figures comparable

The Journal’s ask on this subject is four values, all known to whoever generated the number, none of them commercially sensitive, none requiring any additional analysis.

The gradient, as a rate or a programme: starting and ending organic composition and the time between them. The detection wavelength. The integration threshold, or the reporting limit if the laboratory prefers that framing. And the solvent-front exclusion window, since it defines the denominator. Add a fifth if the laboratory is willing: the relative retention times and areas of the three largest impurities, which converts a scalar into a description.

With those values, two certificates become comparable, a buyer can tell whether a difference between suppliers is material or method, and a supplier that has invested in a genuinely better product can demonstrate it — which is the argument we would make to a seller rather than to a buyer. Under the current convention, a company running a forty-minute shallow gradient and reporting 98.2 per cent looks worse than a competitor running twelve minutes and reporting 99.4, and there is no mechanism by which the first can show a buyer why. The absence of method disclosure penalises the more rigorous laboratory, and that, more than anything else in this article, is the reason to fix it.

It is worth remembering how narrow the question is. A chromatogram reports the proportion of ultraviolet-absorbing material that eluted as the main peak. It has nothing to say about how much peptide was in the vial, about what the sequence is, about aggregation, or about anything microbiological. Those are four separate purchases.

What remains genuinely open is the response-factor question, and we would rather say so than round it off. Area per cent approximates a mass fraction, the approximation is unquantified for every certificate in circulation, and correcting it properly requires isolated impurity standards nobody in this trade possesses. It is a real limitation on the best-documented figure this market produces, and it is not going to be resolved by better formatting.