How do I reconcile 96.4% from Medutest with 97.1% from the supplier?

What I am working with: 96.4% · Medutest · 97.1%.

This is not behaving the way I expected and I want to understand the discrepancy before I act on it.

I have photographed the current state and recorded the conditions, so I can answer follow-up questions precisely.

How do I distinguish the benign explanation from the one that matters?

0.7 percentage points, and the direction is the informative half. Medutest reports 96.4 per cent and the certificate 97.1, so the independent figure is lower. As impurity that is 3.6 per cent against 2.9 — 1.24 times as much unassigned area. A gentler gradient resolves impurities that a steeper one hides beneath the main peak, so the better method routinely reports the worse number; 0.7 points is comfortably inside what method choice alone produces on identical material. Ask both parties for the method section before you decide which figure is wrong, because the answer is often neither.

Understanding purity requires separating the chemistry from the method from the reporting convention, and the three are not independent.

Temperature affects the dynamics of molecular conformation, and if a peptide has proline residues that interconvert on the chromatographic timescale, the peak will split or shoulder at low temperature and collapse at high temperature.

Mass shifts and what they usually mean

The underlying point is that integration of the shoulder — whether you use perpendicular drop or tangent skim — determines what area gets assigned to the main peak versus the impurity table.

The limitation is that single-digit micro-impurities become invisible at typical reporting thresholds, so "no impurities detected" means "none above one in two thousand."

Compare purity within a single laboratory on the same method, never across laboratories.

edited 29 Jan 2026 by rune_thoresen — expanded the table to cover the lower concentration

The short answer is that two competent laboratories on identical material will disagree, and the disagreement is almost always explainable by method differences.

Mass on column affects detector linearity and peak overlap — overloading broadens peaks and hides neighbours, while underloading improves resolution but loses sensitivity.

On the detail: buffer versus acid in the mobile phase changes the ionisation state of basic and acidic residues, shifting retention and selectivity — same vial, potentially different separation.

The caveat is that purity without identity is only half an answer — a high purity does not mean the peak is actually what you think it is.

If you are ranking vendors, specify a method and have all samples tested at the same place.

The part that matters: the single most important distinction is between what purity measures — the fraction of detected material that is your target — and what you actually want to know — how much of the material in the vial is your target.

Column pore size affects mass transfer — a 100 Angstrom packing on a 5 kDa peptide restricts diffusion, broadening the peak and potentially hiding small impurities in the shoulders.

Mobile phase additive choice affects ionisation and peak shape — TFA gives sharp peaks but suppresses mass spectrometry signal, formic acid gives worse peaks but preserves signal.

The ICH Q3A impurity thresholds and the relevant pharmacopoeial chapters all specify method validation requirements that almost no research-grade certificate claims to meet.

One qualification: achieving purity above roughly 98 per cent on a 30-residue peptide is fighting the chemistry of synthesis, not the quality of the purification.

The practical summary: ask for the chromatogram and the method, and ignore the headline number until you have both.

The honest answer is that the achievable range of plausible purity figures for a given vial is wider than most people expect.

Retention time is sequence-specific and method-specific, so comparing your result to a supplier value using a different method is meaningless without method documentation.

Published side-by-side method comparisons show that a two-point difference in purity on the same vial is easily explained by method choice alone.

If you only pay for one test, pay for quantified content. Purity is the number everyone quotes and content is the number that changes what you do.

More usefully, area percentage is not mass percentage, and conflating the two is the most common misreading of a purity figure.

Tailing factor measures peak shape, and a badly tailing peak spreads into the region where small impurities live, forcing tangent-skim integration that assigns tail area to the main peak.

In practice: ask for the chromatogram, check the method section, check the lot number against the vial, and set your accept threshold before you see the result rather than after.