AHK-Cu vs GHK-Cu: Two Copper Tripeptides Compared

AHK-Cu vs GHK-Cu comes down to one residue: AHK-Cu is the tripeptide Ala-His-Lys bound to copper(II), and GHK-Cu is Gly-His-Lys bound to copper(II). The only difference is the N-terminal residue, alanine in place of glycine, which adds one methyl group (CH2 to the formula) and 14.016 Da to the monoisotopic mass. GHK was first reported from human plasma in 1973 and has about 315 PubMed records; AHK is a synthetic analogue with two.

AHK-Cu vs GHK-Cu at a glance

Source: PubChem records 7408502, 168431292, 73587 and 71587328; AHK CAS also in the Global Substance Registration System (UNII 1AVY5QO8WR); masses and m/z values recomputed from the formulas; references as numbered.

Research materials: Vortex supplies GHK-Cu as a lyophilised research reagent: GHK-Cu 50mg and GHK-Cu 100mg. Vortex does not carry AHK-Cu. Lab reports: /coa/.

The one structural difference: glycine vs alanine

Glycine is the only standard amino acid without a side chain; its alpha carbon carries two hydrogens. Alanine replaces one of those hydrogens with a methyl group. Swap the first residue of GHK for alanine and every other atom stays where it was: the histidine in position 2, the lysine in position 3, the free N-terminal amine and the free C-terminal carboxylate. Backbone basics are in peptide structure.

That methyl group has three consequences that matter for identity work:

• Mass. CH2 adds 14.0157 Da to the monoisotopic mass and 14.03 g/mol to the average molar mass, in both the free peptide and the copper complex.
• Chirality. Glycine has no stereocentre, so GHK has two (His and Lys). Alanine has one, so AHK has three. A synthesis that racemises the first residue can give a D-Ala diastereomer of AHK. GHK has no equivalent impurity at that position.
• Hydrophobicity. On reversed-phase HPLC scales built from model peptides, alanine is more hydrophobic than glycine [7], so AHK should elute somewhat later than GHK under the same method.

The N-terminus of human serum albumin begins Asp-Ala-His-Lys (DAHK), so Ala-His-Lys is residues 2 to 4 of albumin [1]. In DAHK the histidine sits in position 3; in free AHK, with the aspartate removed, it sits in position 2. That shift changes its copper-binding class.

AHK-Cu vs GHK-Cu copper coordination

AHK and GHK differ in where histidine sits, and that position sets how each binds copper(II).

Two classes of N-terminal copper site

Peptides with a free N-terminus bind copper(II) in two well-defined ways, set by where histidine sits [3, 4]. With histidine in position 2 (the NH2-Xxx-His motif), copper is held by three peptide nitrogens and an external ligand in the equatorial plane, called the 3N form. With histidine in position 3 (NH2-Xxx-Zzz-His, the ATCUN motif), copper is held by four peptide nitrogens, the 4N form [3]. GHK is the canonical Xxx-His peptide and DAHK the canonical ATCUN peptide [3]; AHK, with histidine in position 2, belongs by sequence to the Xxx-His class. Both families are compared in copper peptides and the ATCUN motif.

What is measured for GHK-Cu

At neutral pH, GHK forms a mononuclear 1:1 copper(II) complex. Freedman and colleagues found an EPR spectrum consistent with two or three equatorial nitrogen donors, one of them on the histidine imidazole, with optical transitions at apparent pK values of 3.6, 9.2 and 11.4 as pH changed [2]. Hureau and colleagues confirmed by X-ray, EPR, HYSCORE, X-ray absorption and NMR spectroscopy that in solution copper is held by the terminal amine, a deprotonated amide nitrogen and the imidazole nitrogen, with a labile oxygen in the fourth equatorial position. In the crystal the complex is a dimer [1]. Conato and colleagues showed that the lysine side-chain amine joins the copper site only at alkaline pH [5].

What is known about AHK-Cu

No structural or thermodynamic study of Cu(II)-AHK was found in PubMed: there is no measured binding constant, pK set, crystal structure or EPR spectrum. The closest measured system is Ala-His-His (AHH), which starts with the same Ala-His pair. Gonzalez and colleagues found that Cu(II)-AHH forms the 3N, Xxx-His-type complex predominantly at pH 4 to 6.5, switching to a 4N form at pH 6.5 to 10 because its third residue is also a histidine [3]. A 2019 follow-up showed that external ligands can switch Cu(II)-AHH between the two modes [4]. AHK has lysine, not histidine, in position 3, so the 4N mode through a third-position imidazole is not available to it.

On that basis, AHK-Cu is expected to use the same 3N donor set as GHK-Cu: terminal amine, amide nitrogen and histidine imidazole. That is an inference from sequence and from related peptides, not a measurement. Whether the methyl group shifts binding strength, copper exchange speed or redox behaviour has not been reported, and no stability data exist for lyophilised AHK-Cu.

Source: references 1, 2, 3 and 4.

Where each molecule came from

GHK has a documented natural origin. Pickart and Thaler reported the tripeptide from human plasma in 1973, and later chemistry papers describe GHK and DAHK as naturally occurring high-affinity copper(II) chelators in blood plasma [1]. Older papers abbreviate it GHL [2]. Its discovery and solution chemistry are covered in the GHK-Cu peptide reference.

AHK has no comparable history. No isolation of free Ala-His-Lys from a natural source was found in PubMed, and the earliest indexed paper naming AHK-Cu dates from 2007. The free peptide is registered under CAS 126828-32-8, with the systematic name alanylhistidyllysine. The only PubChem copper record, CID 168431292, is a monohydrochloride (CAS 682809-81-0) whose sources are all chemical vendors. AHK is a synthetic analogue of GHK.

How big is each literature?

A PubMed title-and-abstract search on 1 November 2026 for GHK, glycyl-histidyl-lysine and glycyl-L-histidyl-L-lysine returned 315 records, a few of them unrelated uses of the abbreviation. They span coordination chemistry, structural work, cell culture, rodent pharmacology and reviews.

The same search for AHK-Cu, Ala-His-Lys and alanyl-histidyl-lysine returned 31 records, but nearly all of them match the four-residue albumin sequence DAHK or longer peptides that contain Ala-His-Lys. Only two describe the free tripeptide or its copper complex: a 2007 culture study of AHK-Cu and a 2018 cell-culture paper on a vitamin C-linked AHK conjugate. No chemistry, structure or stability paper on AHK-Cu was found, and no controlled human studies of either compound have been published.

Telling AHK-Cu and GHK-Cu apart by analysis

One methyl group is enough for routine methods to separate the two. What matters is which method a report used.

Intact mass by LC-MS

Electrospray ionisation brings small peptides into the mass spectrometer intact [8]. Under acidic LC mobile phases the copper complex often dissociates, so the free peptide ion is usually the main signal. Free GHK gives [M+H]⁺ at m/z 341.19 and free AHK at m/z 355.21, a gap of 14.016. When a copper-containing ion survives, [Cu(L−H)]⁺ appears as a ⁶³Cu/⁶⁵Cu pair, roughly 2.2 to 1 in height: m/z 402.11 and 404.11 for GHK-Cu, 416.12 and 418.12 for AHK-Cu. Each of these ions can be checked against its expected value; LC-MS peptide analysis explains how.

Why +14 Da alone is not proof of AHK

A +14.016 Da shift from GHK means “one more CH2”, not “alanine at the N-terminus”. The GHK methyl ester (methylated C-terminal carboxyl) has exactly the same formula as AHK, C15H26N6O4, and so does an N-methylglycine (sarcosine) variant of GHK. All three give an [M+H]⁺ of m/z 355.21. Intact mass can confirm that a material is not GHK; it cannot by itself confirm that it is AHK.

Locating the methyl group by MS/MS

Tandem mass spectrometry breaks the backbone and reports fragment ions named by a standard convention: b ions keep the N-terminus, y ions keep the C-terminus [9]. For these two peptides:

Source: our calculation from monoisotopic residue masses, singly charged ions. Fragment naming follows reference 9.

A +14 b2 ion with unchanged y ions places the extra methyl on the first two residues, which is what AHK predicts. A +14 shift in the y ions with an unchanged b2 points to a C-terminal modification of GHK instead. The b2 ion cannot separate alanine from N-methylglycine, which are isomers with the same residue mass, so the last step is a retention match against an AHK reference standard.

Retention by HPLC

Reversed-phase selectivity for peptides depends on column chemistry and organic solvent [6], and both tripeptides elute early, near the solvent front. The alanine methyl group makes AHK somewhat more retained than GHK [7], so a method can resolve them. Retention alone does not identify a peak, though: an identity claim based on HPLC means the main peak matched a reference standard run under the same conditions. A D-Ala diastereomer of AHK has the same mass and shows up only as a separate HPLC peak. What a purity figure does and does not measure is covered in HPLC peptide purity.

Copper

Neither HPLC-UV nor the free-peptide MS ion measures copper. A copper-to-peptide ratio needs a separate method such as ICP-MS or ICP-OES alongside a peptide assay. GHK-Cu is blue to violet-blue as a solid, and colour is not a test that separates two copper complexes with the same expected donor set.

Checking a lot of GHK-Cu

Vortex publishes two GHK-Cu reports at /coa/, checked on 1 November 2026:

• GHK-Cu 50 mg, lot VR01-GHK. Tested for Vortex Research by Freedom Diagnostics, reported 06/20/2026. Purity 99.85% by HPLC-UV, identity by LC-MS, net content 54.92 mg in a vial labelled 50 mg.
• GHK-Cu 100 mg, batch PS07-GHK100. Partner-facility report: issued by ILS Laboratories to Vortex’s wholesale partner facility for the batch Vortex stocks, tested 08/12/2026. Purity 99.34% by HPLC, identity by HPLC, net content 106.36 mg in a vial labelled 100 mg. The report also lists heavy metals (ICP-MS), sterility (PCR), endotoxin (USP <85>) and a fentanyl screen.

The two identity methods answer the AHK question differently. An LC-MS identity result measures mass, so an AHK substitution would appear as a 355.21 ion where 341.19 is expected. An HPLC identity result is a retention match against a reference standard. Neither report lists a separate copper-to-peptide assay, so this article draws no conclusion about copper content for either lot. Each report links to the testing lab; how to read a peptide COA explains each field.

Frequently asked questions

What is the difference between AHK-Cu and GHK-Cu?

AHK-Cu is Ala-His-Lys bound to copper(II); GHK-Cu is Gly-His-Lys bound to copper(II). The only difference is the first residue, alanine instead of glycine, which adds one methyl group and 14.016 Da. Histidine and lysine are in the same positions in both.

What is the AHK-Cu peptide?

AHK-Cu is the copper(II) complex of the synthetic tripeptide L-alanyl-L-histidyl-L-lysine. The free peptide is C15H26N6O4, 354.41 g/mol, CAS 126828-32-8, PubChem CID 7408502. Its PubMed record is two papers.

Does AHK bind copper the same way as GHK?

By sequence it should. Both have histidine in position 2, the Xxx-His motif, which binds copper through the terminal amine, an amide nitrogen and the imidazole. GHK-Cu’s site has been measured by EPR, X-ray and NMR; no equivalent study of Cu(II)-AHK has been published.

How can a lab tell AHK-Cu from GHK-Cu?

By LC-MS, the free peptides give [M+H]⁺ ions at m/z 355.21 (AHK) and 341.19 (GHK). Because other GHK variants share AHK’s mass, MS/MS confirms the position: the b2 ion is m/z 209.10 for AHK and 195.09 for GHK.

Does Vortex sell AHK-Cu?

No. Vortex does not carry AHK-Cu. It supplies GHK-Cu in 50 mg and 100 mg vials, with lab reports published on the /coa/ page.

Research materials: GHK-Cu 50mg and GHK-Cu 100mg are supplied as lyophilised research reagents. Vortex does not carry AHK-Cu. Published lab reports are at /coa/.

About this article: Written by the Vortex Research Editorial Team. Sources are peer-reviewed papers indexed in PubMed, each checked for title, journal and year and matched to its abstract. Formulas and masses are from PubChem; m/z values are calculated from the formulas. Lab report figures are from the published reports at /coa/.

References

1. Hureau C, Eury H, Guillot R, 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. doi:10.1002/chem.201100751
2. Freedman JH, Pickart L, Weinstein B, Mims WB, Peisach J. Structure of the glycyl-L-histidyl-L-lysine-copper(II) complex in solution. Biochemistry. 1982;21(19):4540-4544. PMID: 6291585. doi:10.1021/bi00262a004
3. Gonzalez P, Vileno B, Bossak K, et al. Cu(II) binding to the peptide Ala-His-His, a chimera of the canonical Cu(II)-binding motifs Xxx-His and Xxx-Zzz-His. Inorg Chem. 2017;56(24):14870-14879. PMID 29190078. doi:10.1021/acs.inorgchem.7b01996
4. Gonzalez P, Bossak-Ahmad K, Vileno B, et al. Triggering Cu-coordination change in Cu(II)-Ala-His-His by external ligands. Chem Commun (Camb). 2019;55(56):8110-8113. PMID: 31233054. doi:10.1039/c9cc03174j
5. Conato C, Gavioli R, Guerrini R, et al. Copper complexes of glycyl-histidyl-lysine and two of its synthetic analogues: chemical behaviour and biological activity. Biochim Biophys Acta. 2001;1526(2):199-210. PMID: 11325542. doi:10.1016/s0304-4165(01)00127-1
6. Mant CT, Cepeniene D, Hodges RS. Reversed-phase HPLC of peptides: assessing column and solvent selectivity on standard, polar-embedded and polar endcapped columns. J Sep Sci. 2010;33(19):3005-3021. PMID: 21038458. doi:10.1002/jssc.201000518
7. Kovacs JM, Mant CT, Hodges RS. Determination of intrinsic hydrophilicity/hydrophobicity of amino acid side chains in peptides in the absence of nearest-neighbor or conformational effects. Biopolymers. 2006;84(3):283-297. PMID: 16315143. doi:10.1002/bip.20417
8. Fenn JB, Mann M, Meng CK, Wong SF, Whitehouse CM. Electrospray ionization for mass spectrometry of large biomolecules. Science. 1989;246(4926):64-71. PMID: 2675315. doi:10.1126/science.2675315
9. Roepstorff P, Fohlman J. Proposal for a common nomenclature for sequence ions in mass spectra of peptides. Biomed Mass Spectrom. 1984;11(11):601. PMID: 6525415. doi:10.1002/bms.1200111109

Vortex Research products are sold for in-vitro laboratory research by qualified researchers. Not for human or veterinary use. Not a drug, dietary supplement or cosmetic. Statements have not been evaluated by the FDA.