Glossary category

Analytical chemistry

Terms describing the determination of identity, purity, content and quality.

Terms

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Table 1. Terms in the analytical chemistry category.

Term	Definition
Adsorption	Loss of peptide to container and syringe surfaces, material at low concentration in the absence of a surfactant. A source of content loss that no certificate reports.
AggregationFibrillation	Association of peptide molecules into higher-order species, driven by agitation, freeze–thaw cycling and proximity to the isoelectric point. Not detected by a reverse-phase purity method and requires size-exclusion analysis.
Average mass	The mass calculated using isotope-averaged atomic weights, and the value returned by deconvolution of an electrospray charge envelope at moderate resolution.
Chromatographic purityArea percentage purity · HPLC purity	The area of the main peak as a percentage of total integrated area under stated conditions. A relative measure that assumes comparable molar absorptivity across species and that does not detect anything without a chromophore.
Counter-ionSalt form	The ion paired with a charged peptide, most often trifluoroacetate or acetate from purification. Can constitute a substantial fraction of the mass of a basic peptide and is not detected by a purity method.
Deamidation	Conversion of asparagine or glutamine to the corresponding acid, adding one dalton and altering charge. A principal storage-related degradation route for peptides in solution.
Deletion sequence	An impurity lacking one or more residues, arising from incomplete coupling during synthesis. Often close-eluting and detectable only by mass spectrometry.
Enantiomeric purityChiral purity	The proportion of residues in the intended stereochemical configuration. Racemisation during synthesis produces species of identical mass and identical reverse-phase retention, detectable only by a chiral method.
EndotoxinBacterial endotoxin · LPS	Lipopolysaccharide from Gram-negative bacterial cell walls, pyrogenic on parenteral administration. Determined by a lysate-based or recombinant assay against a dose- and route-dependent limit, and reported essentially never in the research-supply channel.
Forced degradationStress testing	Deliberate exposure of a sample to heat, light, acid, base and oxidant to generate degradation products against which a method’s specificity is demonstrated.
Freeze–thaw	Repeated freezing and thawing of a solution. Concentrates solutes in the shrinking liquid fraction, drives pH excursion as buffer components crystallise differentially, and promotes aggregation.
Gradient	The programmed change in mobile-phase composition over a chromatographic run. A shallow gradient resolves close-eluting species that a steep gradient co-elutes, and the two report different purities for the same material.
Identity	Confirmation that the material is the substance claimed. Retention-time correspondence alone is weak evidence of identity; mass-spectrometric confirmation of the expected mass is stronger; sequence confirmation is stronger still.
In-use periodBeyond-use date	The time a reconstituted preparation may be stored and used, supported by stability-indicating data on that presentation. The Institute publishes no in-use claim for any research-supply presentation because no such data exist for one.
Integration window	The retention-time range over which peak areas are summed. Changing the window changes the reported purity without changing the material, which is why the Institute requires the window to be reported.
Isomerisation	Conversion of aspartate to isoaspartate through a succinimide intermediate, producing a species of identical mass and different retention. Undetectable by mass alone.
Karl Fischer titrationKF	The reference method for water content, based on the stoichiometric reaction of water with iodine and sulfur dioxide. The only water determination the Institute accepts for a hygroscopic lyophilisate.
Limit of detectionLOD	The lowest concentration at which the presence of a species can be distinguished from noise. Lower than the limit of quantitation and not a basis for a numerical result.
Limit of quantitationLOQ	The lowest concentration at which a determination can be made with stated precision and accuracy. An impurity below the limit is not absent; it is unquantified.
Mass balance	The sum of peptide content, water content and counter-ion content, which should approach one hundred per cent. A certificate that does not permit a mass balance to be closed is incomplete under the Institute’s minimum data standard.
Mass spectrometryMS · ESI-MS	Determination of mass-to-charge ratio, used to confirm identity. For a peptide, deconvolution of the multiply charged envelope should return an average mass within a stated tolerance of the theoretical value.
Monoisotopic mass	The mass calculated using the most abundant isotope of each element. Requires high resolving power to determine for a peptide of several kilodaltons, and is not interchangeable with average mass.
Orthogonal method	A second analytical method resting on a different physical principle from the first. Identity confirmed by two orthogonal methods is materially stronger than identity confirmed by one, and is required by the Institute’s minimum data standard.
Oxidation	Addition of oxygen to methionine, tryptophan or cysteine, adding sixteen daltons. Accelerated by peroxide, light and metal ions.
Peptide contentPeptide assay	The mass fraction of a lyophilisate that is peptide, the remainder being water, counter-ion and residual solvent. Determined by amino-acid analysis or nitrogen determination. Distinct from purity, and the two are frequently conflated.
Peptide mapping	Enzymatic digestion followed by chromatographic and mass-spectrometric analysis of the resulting fragments, used to confirm sequence. Rarely performed on research-supply material.
Reference standard	A characterised material with an assigned content value against which a determination is made. A result obtained without a named standard reports an area percentage rather than a content.
Related substancesImpurities	Impurities structurally related to the target peptide, arising from synthesis or degradation. Reported by name and retention time on a complete certificate and as a single aggregate figure on most.
Reverse-phase HPLCRP-HPLC	Separation on a hydrophobic stationary phase under an acidified aqueous–organic gradient, with ultraviolet detection at 214 nanometres. The Institute’s reference method for the chromatographic purity of synthetic peptides.
Size-exclusion chromatographySEC	Separation by hydrodynamic radius, the standard method for aggregate determination. Reported on no certificate in the Institute’s sampled set.
Stability-indicating	A property of a method that separates the intact compound from its degradation products, demonstrated by forced-degradation study. A purity method that is not stability-indicating cannot support a shelf-life or in-use claim.
Sterility	The absence of viable microorganisms, demonstrated by a compendial test on the finished presentation. Distinct from endotoxin content, which persists after sterilisation.
System suitability	A set of criteria confirmed at the time of analysis establishing that the chromatographic system is performing adequately. A result obtained on a system failing suitability is not reportable.
TrifluoroacetateTFA	The counter-ion most commonly introduced by reverse-phase purification with trifluoroacetic acid. Quantified by ion chromatography or fluorine nuclear magnetic resonance, and reported on few certificates.
Water contentResidual moisture	The mass fraction of a lyophilisate that is water. Substantial for a hygroscopic peptide and a direct subtraction from delivered peptide mass.