Kisspeptin-10 — analytical characterisation
Chromatographic conditions, identity, related substances, presentation, reconstitution and in-use stability.
§6Analytical characterisation
§6.1Chromatographic conditions
- Column
- C18, 4.6 × 250 mm, 5 µm
- Mobile phase and gradient
- A: 0.1 % trifluoroacetic acid in water; B: acetonitrile. Gradient 15–45 % B over 25 min
- Detection
- UV 214 nm; 280 nm strong (Trp3, Tyr1) — reliable secondary wavelength
- Retention
- Intermediate
§6.2Identity by mass spectrometry
[M+H]⁺ at m/z 1303.5; [M+2H]²⁺ at m/z 652.3. Average mass 1302.5 ± 1 Da.[3]
§6.3Related substances and degradation
Table 7. Related substances recorded for Kisspeptin-10, with the process or storage route that generates each and its analytical signature.
| Related substance | Origin | Analytical signature |
|---|---|---|
| Kisspeptin-54 | Substitution | A markedly different mass; trivially distinguishable, yet conflated in labelling |
| C-terminal free acid | Incomplete amidation | +0.98 Da; the C-terminal amide is essential to receptor binding and its loss abolishes activity, so this near-isobaric impurity is functionally critical |
| Trp3 oxidation | Oxidation and light | +16 and +32 Da |
| Deamidated Asn2 and Asn4 | Storage | +1 Da each; two adjacent-region asparagines make this sequence notably deamidation-prone, and the deamidated species are near-isobaric with the parent |
| Trifluoroacetate counter-ion | Purification | Arg9 and the N-terminus are basic |
Degradation routes
- Tryptophan oxidation
- Deamidation at two asparagine sites
- Hydrolysis of the C-terminal amide
- Aspartimide formation from the deamidated intermediates
§7Presentation, reconstitution and storage
§7.1Presentation and reconstitution
- Presentation
- Lyophilised powder in vial
- Reconstitution
- A 5 mg vial with 2.5 mL gives 2 mg/mL; a 1 mg quantity is 0.50 mL, that is 50 units on a U-100 syringe.
- Storage, lyophilised
- −20 °C, desiccated, protected from light
- Storage, reconstituted
- 2–8 °C protected from light; the combination of an oxidation-labile tryptophan and two deamidation-prone asparagines makes this a short-lived peptide in solution
- In-use period
- No supported claim
The +1 Da species arising from deamidation and the +0.98 Da species arising from amide hydrolysis are both near-isobaric with each other and with the parent. Distinguishing them requires high resolution and, ideally, peptide mapping. For this sequence a purity figure from a low-resolution method is close to meaningless.
§7.2In-use stability
Applicable standards: CEI-MS-01 · CEI-MS-02 · CEI-MS-03 · CEI-MS-04 · CEI-MS-05 · CEI-MS-06. The full series is at methodological standards.
Working calculators: reconstitution and insulin-unit conversion · purity against peptide content · certificate minimum-data checker.
References cited on this page
References are numbered in order of first citation in this document. Each superscript in the text links to its entry below.
- United States Pharmacopeial Convention. General Chapter ⟨1225⟩ Validation of Compendial Procedures. United States Pharmacopeia — National Formulary (USP–NF) 2024;USP 2024 Issue 1. identifier not held by the Institute
- International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use. ICH Q2(R2) Validation of Analytical Procedures. ICH Harmonised Guideline 2023;Step 4 version, 1 November 2023. identifier not held by the Institute
- International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use. ICH Q6B Specifications: Test Procedures and Acceptance Criteria for Biotechnological/Biological Products. ICH Harmonised Tripartite Guideline 1999;Step 4 version. identifier not held by the Institute
- International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use. ICH Q1A(R2) Stability Testing of New Drug Substances and Products. ICH Harmonised Tripartite Guideline 2003;Step 4 version. identifier not held by the Institute
- Manning MC, Chou DK, Murphy BM, Payne RW, Katayama DS. Stability of protein pharmaceuticals: an update. Pharmaceutical Research 2010;27(4):544–575. doi:10.1007/s11095-009-0045-6 · PMID 20143256
Identifiers are reproduced only where the Institute holds them. Where a digital object identifier or PubMed identifier is not shown, the Institute has recorded the journal and year and has not constructed an identifier.