In vitro
This computational study introduces the concept of "carbonless" biomolecular design, in which all carbon atoms in amino acids and peptides are systematically replaced by boron and nitrogen atoms under an isoelectronicity constraint. Using glycine, histidine, lysine, and the copper-binding tripeptide Gly-His-Lys (GHK) as model systems, the researchers applied density functional theory (DFT) calculations with aqueous solvation modeling and conformer sampling to identify the most stable carbonless analogues (cGly, cHis, cLys, and cGHK) among all possible boron-nitrogen constitutional isomers. The study predicted that cGHK displays a broader conformational landscape than native GHK under physiological aqueous conditions, suggesting enhanced structural flexibility. Copper(II) binding was modeled using an experimentally informed coordination motif, and thermodynamic calculations indicated that cGHK binds Cu(II) more favorably than GHK by approximately 6.24 kcal/mol. The work is entirely theoretical; no synthesis, cell-based, animal, or human experiments were conducted. Limitations include the absence of experimental validation of the proposed carbonless structures, reliance on computational approximations for solvation and conformational sampling, and uncertainty about whether these novel BN-substituted molecules could be synthesized or would exhibit biological stability.
Physical chemistry chemical physics : PCCP · Apr 2026DOI ↗ In vitro
This study investigated the catalytic properties of GHK-Cu (a copper-bound tripeptide composed of glycine, histidine, and lysine) as a mimic of the enzyme laccase, which is naturally used in the detection and degradation of phenolic compounds. Researchers characterized GHK-Cu's enzyme-like kinetics, finding a maximum reaction velocity (Vmax) of 1.735 × 10⁻⁴ mM·s⁻¹ and a Michaelis constant (Km) of 0.061 mM, suggesting strong substrate affinity and catalytic efficiency compared to natural laccase. Building on this, the team developed colorimetric assays to detect two phenolic compounds—epinephrine (EP) and 2-aminophenol (2-AP)—across defined concentration ranges in ultrapure water and seawater. Additionally, a portable cotton-based sensor paired with a smartphone platform was constructed to enable field-ready detection of 2-AP in seawater. The study is conducted entirely in vitro and focuses on analytical chemistry applications rather than biological or therapeutic effects of GHK-Cu. Key limitations include the absence of any cell-based, animal, or human testing; findings are restricted to a controlled laboratory sensing context. The work proposes GHK-Cu's copper-coordination structure as a template for designing novel synthetic laccase mimetics.
Biosensors · Apr 2026DOI ↗ In vitro
This review/research paper examines two naturally occurring peptides — carnosine (β-alanyl-L-histidine) and GHK (glycyl-L-histidyl-L-lysine) — and explores strategies to improve their stability and bioavailability through glycoconjugation. A key challenge addressed is the rapid degradation of carnosine by the enzyme carnosinase and the inherent low stability of GHK. The authors report that conjugating these peptides to either trehalose (a disaccharide) or hyaluronan (a polysaccharide) inhibits carnosinase activity and increases tripeptide stability, while also protecting the saccharide components from degradation. The study further investigates copper-binding properties of these glycoconjugates, finding that the saccharide components potentiate the Cu,Zn-superoxide dismutase-like (antioxidant) activity of the resulting copper(II) complexes. The glycoconjugates are reported to act as copper ionophores in cell culture, increasing intracellular copper levels and stimulating copper-driven signaling pathways, leading to enhanced expression of trophic and angiogenic proteins including BDNF, BMP-2, and VEGF. Copper chaperones CCS and Atox-1 are implicated as transcription factors in these pathways. Limitations include reliance on in vitro cell culture models and biochemical assays, with no human clinical data presented.
Antioxidants (Basel, Switzerland) · Dec 2025DOI ↗ In vitro
This study investigated the synthesis and biological activity of novel conjugates combining hyaluronic acid (HA) with the copper-binding tripeptide glycyl-l-histidyl-l-lysine (GHK) at varying loading ratios, forming GHK-HA conjugates. The conjugates were designed to address the individual limitations of HA and GHK, including susceptibility to hydrolytic degradation and oxidative stress. In vitro assays demonstrated that GHK-HA bound copper(II) ions effectively and showed enhanced antioxidant properties compared to the individual components alone. When complexed with copper, the conjugates promoted the expression and release of several growth and trophic factors, including brain-derived neurotrophic factor (BDNF), vascular endothelial growth factor (VEGF), and bone morphogenetic protein-2 (BMP-2), suggesting synergistic osteogenic and angiogenic potential. The study also explored a proposed mechanistic pathway, finding that copper's intracellular chaperones — CCS and Atox1 — translocated to the nucleus and appeared to function as transcription factors. Limitations include the exclusively in vitro nature of the experiments, meaning results cannot yet be extrapolated to animal models or human clinical outcomes. No in vivo validation was performed.
Bioconjugate chemistry · Mar 2025DOI ↗ In vitro
This study presents a novel analytical application of capillary electrophoresis coupled to inductively coupled plasma tandem mass spectrometry (CE-ICP-MS/MS) to investigate the encapsulation of the copper tripeptide complex GHK-Cu within liposomes — a delivery system commonly used in cosmetic formulations. Using an ethanol injection method to form liposomes, the researchers monitored copper (Cu) and phosphorus (P) signals to confirm liposome formation and quantify the concentration of GHK-Cu encapsulated within them. The study demonstrates that CE-ICP-MS/MS, valued for its high sensitivity and ability to preserve analytes in their intact chemical form under mild, physiologically compatible conditions, can be successfully applied in cosmetic science contexts. The authors propose this methodology could support the development and optimization of diverse liposomal cosmetic formulations. Importantly, this paper is a methodological/analytical chemistry study — it does not test GHK-Cu efficacy in humans or animals, make clinical claims, or evaluate biological outcomes. Its findings are limited to demonstrating an analytical technique for characterizing a cosmetic ingredient's encapsulation efficiency in a laboratory setting.
Electrophoresis · Oct 2024DOI ↗