Short answer: GHK-Cu and AHK-Cu are copper(II) complexes of two tripeptides that differ by a single residue. GHK is Gly-His-Lys; AHK is Ala-His-Lys. Swapping the N-terminal glycine for alanine adds one methyl group (CH2, +14.016 Da) and one extra stereocentre, and leaves the copper coordination site intact, because both peptides carry histidine in position 2. Both complexes are blue to blue-violet solids. On a certificate of analysis they are told apart by mass: the free peptides sit at 340.19 and 354.20 Da (monoisotopic), and the copper-bearing ions at m/z 402.11 and 416.12.
This guide compares the two materials on structure and analytical specification only: sequence, composition, how copper is held, how the mass is quoted, and what a lot certificate for each should report. It does not describe what either compound does in any biological system.
GHK-Cu vs AHK-Cu at a glance
| GHK-Cu | AHK-Cu | |
|---|---|---|
| Peptide sequence | H-Gly-His-Lys-OH (GHK) | H-Ala-His-Lys-OH (AHK) |
| Length | 3 residues | 3 residues |
| Free peptide formula | C14H24N6O4 | C15H26N6O4 |
| Free peptide mass (monoisotopic / average) | 340.1859 / 340.38 Da | 354.2016 / 354.41 Da |
| 1:1 complex, two protons displaced (formula) | C14H22CuN6O4 | C15H24CuN6O4 |
| 1:1 complex mass (monoisotopic 63Cu / average) | 401.0999 / 401.91 Da | 415.1155 / 415.94 Da |
| Expected ESI+ ion of the complex | m/z 402.107 (63Cu), 404.105 (65Cu) | m/z 416.123 (63Cu), 418.121 (65Cu) |
| Theoretical copper content (1:1, no counter-ion) | 15.8% w/w | 15.3% w/w |
| Stereocentres | 2 (L-His, L-Lys) | 3 (L-Ala, L-His, L-Lys) |
| Modifications | None on the peptide; Cu(II) coordinated | None on the peptide; Cu(II) coordinated |
| Salt form typically supplied | Copper complex, often with acetate or chloride as counter-ion; check the lot COA | Copper complex; the registry record is a monohydrochloride |
| CAS (peptide / copper complex) | 49557-75-7 / 89030-95-5 | 126828-32-8 / 682809-81-0 |
| Appearance | Blue to blue-violet powder | Blue to blue-violet powder |
Masses in the table are calculated from the formulas with standard isotopic masses and checked against public chemical registries. The ion values assume the common positive-mode species [Cu(peptide − H)]+, which is the neutral 1:1 complex plus one proton.
What is the structural difference between GHK and AHK?
Only the first residue changes. Glycine has a hydrogen as its side chain; alanine has a methyl group. Everything else (the histidine imidazole, the lysine ε-amine, the free C-terminal carboxylate) is the same. Three practical consequences follow:
- Mass shift. Each AHK species is 14.0157 Da heavier than its GHK counterpart. That is large enough for any mass spectrometer to resolve, so a certificate that reports an observed mass settles which peptide is in the vial.
- One more chiral centre. Glycine is achiral; alanine is not. AHK can therefore carry a D-Ala diastereomer as a synthesis impurity, a related substance GHK cannot have. Diastereomers have identical masses, so only chromatography or chiral amino acid analysis would reveal one.
- Slightly more retention. The extra methyl usually makes AHK elute a little later than GHK on reversed phase, though both are small, polar and early-eluting.
How is copper held in GHK-Cu and AHK-Cu?
Both peptides belong to the N-terminal Xxx-His family of Cu(II) ligands. In this arrangement the metal is coordinated by three nitrogens from the peptide: the free N-terminal amine, the deprotonated amide nitrogen between residues 1 and 2, and a nitrogen of the histidine imidazole. A fourth equatorial position is filled by an outside ligand. Hureau and colleagues solved the structure of [Cu(II)(GHK)] by X-ray crystallography and multiple spectroscopies. The complex is a dimer in the solid state, with the fourth site taken by a carboxylate oxygen from a neighbouring molecule. It is monomeric in solution, with the same three nitrogen donors1. Calorimetric titration found that GHK and Cu(II) combine predominantly in a 1:1 ratio2. The motif is defined well enough that GHK fused to the end of a protein has been used as a copper tag for crystallographic phasing3.
Alanine leaves the amine, the first amide and the histidine untouched, so AHK presents the same donor set. Two points matter for analysis. First, forming the complex displaces protons from the peptide, and how many are subtracted when the formula is written sets the quoted mass. Second, the complex can partly or fully dissociate under strongly acidic conditions such as a 0.1% TFA mobile phase, so an HPLC trace may show free peptide, complex, or both, depending on the method.
Why do suppliers quote different masses for GHK-Cu?
Four values circulate for “GHK-Cu”, and none of them is a typo. They describe different species:
| Species written | Formula | Average mass |
|---|---|---|
| GHK plus Cu, no protons removed | C14H24CuN6O4 | 403.93 |
| Cationic complex, one proton removed | [C14H23CuN6O4]+ | 402.92 |
| Neutral complex, two protons removed | C14H22CuN6O4 | 401.91 |
| Bis complex, Cu(GHK)2 | C28H46CuN12O8 | 742.3 |
The same spread applies to AHK-Cu, shifted by 14.03. When comparing lots or suppliers, check which convention a certificate uses before reading a 1–2 Da difference as a discrepancy. The free peptide mass is unambiguous, and it is the most reliable anchor for identity.
Why are copper peptides blue?
The free tripeptides are white. Cu(II) in a nitrogen-rich, roughly square-planar site absorbs weakly in the red, so the complexes look blue to violet. Colour is a fair appearance check, but it cannot tell GHK-Cu from AHK-Cu or show that copper is present in the stated ratio.
What each COA should show
GHK-Cu
- Identity by mass spectrometry. The free GHK ion at [M+H]+ m/z 341.19, the copper complex ion at m/z 402.11, or both. A copper-containing ion should show its isotope partner 2 Da higher at roughly 45% of the main peak’s height, because natural copper is about 69% 63Cu and 31% 65Cu. That pattern confirms copper is in the ion.
- HPLC purity as area percent, with the method stated, because acidic conditions can strip the copper and change what the chromatogram shows.
- Copper content as % w/w, ideally by ICP-OES or atomic absorption, or the peptide:copper ratio. About 15.8% is the theoretical value for a 1:1 complex without counter-ions; counter-ions and water lower it.
- Which mass convention the expected value uses (see the table above).
AHK-Cu
- The same set, with the free AHK ion at [M+H]+ m/z 355.21 and the complex at m/z 416.12 (with its 65Cu partner at 418.12).
- A theoretical copper content of about 15.3% for a 1:1 complex without counter-ions. If the material is the monohydrochloride, the chloride lowers the copper fraction further.
- Optionally, a note on chiral purity of the alanine residue, which is the one related substance AHK can have that GHK cannot.
An observed mass of 341 or 402 on an “AHK-Cu” certificate, or 355 or 416 on a “GHK-Cu” one, means the vial holds the other peptide. For the general layout of a peptide certificate see how to read a peptide COA. Counter-ions and net peptide content are covered in salt forms and net peptide content.
Product pages: GHK-Cu and AHK-Cu. Research profile: GHK-Cu. Lot certificates are in the certificate library.
Frequently asked questions
Is AHK-Cu the same as GHK-Cu?
No. They are different tripeptides. AHK has alanine where GHK has glycine, so every AHK species is 14.016 Da heavier. The copper coordination site, built from the N-terminal amine, the first amide nitrogen and the histidine imidazole, is the same in both.
What is the molecular formula of GHK-Cu?
It depends on how the complex is written. The free peptide GHK is C14H24N6O4 (340.38). The neutral 1:1 copper complex with two protons displaced is C14H22CuN6O4 (401.91). Catalogues also quote 402.92 and 403.93 for other ways of writing the same complex.
Why is GHK-Cu powder blue?
Copper(II) in a nitrogen-rich coordination environment absorbs red light weakly, so the complex appears blue to violet. The free peptide is white. AHK-Cu is the same colour for the same reason.
How can a mass spectrum confirm copper is present?
By the isotope pattern. A copper-containing ion appears as a pair 2 Da apart, with the heavier peak at about 45% of the lighter, matching the natural 63Cu:65Cu ratio. A peptide ion without copper does not show that pair.
Does HPLC purity measure the copper?
No. HPLC purity is the share of UV-absorbing peptide material that is the target peptide. Copper content is a separate measurement, usually by ICP-OES or atomic absorption, reported as % w/w.
References
- Hureau C, Eury H, Guillot R, Bijani C, Sayen S, Solari PL, et al. X-ray and solution structures of Cu(II) GHK and Cu(II) DAHK complexes: influence on their redox properties. Chemistry. 2011;17(36):10151–10160. PMID 21780203
- Trapaidze A, Hureau C, Bal W, Winterhalter M, Faller P. Thermodynamic study of Cu2+ binding to the DAHK and GHK peptides by isothermal titration calorimetry (ITC) with the weaker competitor glycine. J Biol Inorg Chem. 2012;17(1):37–47. PMID 21898044
- Mehr A, Henneberg F, Chari A, Görlich D, Huyton T. The copper(II)-binding tripeptide GHK, a valuable crystallization and phasing tag for macromolecular crystallography. Acta Crystallogr D Struct Biol. 2020;76(Pt 12):1222–1232. PMID 33263328
Research context: This guide explains the composition and analytical characterization of research materials for laboratory work. It is not guidance for any use outside qualified research.