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Resposta da Adria Science

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Dear Ivan,

Thank you for your review and for raising these technical questions. We have reviewed the certificate and supporting analytical documentation ourselves, alongside the technical clarification provided by LiquiLabs. Below is a detailed explanation of the testing documented in report LYO-0399 for batch 2441235.

1. Why the mass spectrum shows a signal at 341

The LC/MS method documented in this report uses a mobile phase containing 0.1% trifluoroacetic acid (TFA), followed by positive-ion electrospray mass spectrometry.

LiquiLabs explains that these acidic analytical conditions can disrupt copper binding to GHK. Consequently, the spectrum can predominantly show the peptide after copper has dissociated during analysis.

The signal at approximately m/z 341 is consistent with protonated GHK, [GHK+H]⁺. The approximately 402 Da value refers to the intact GHK-Cu complex. The observed peptide ion and the intact complex therefore represent different chemical species.

This explains how the 341 signal can arise when analysing GHK-Cu. Its presence alone does not establish that copper was absent from the original sample; copper coordination is assessed through additional analytical evidence.

2. How copper coordination is assessed

LiquiLabs explains that it uses a separate Fourier-transform infrared spectroscopy (FTIR) measurement to assess copper binding. This includes evaluating features in the carboxylate stretching region around 1350–1600 cm⁻¹ and comparing the spectrum against certified reference-material spectra.

For batch 2441235, the certificate reports an FTIR identification score of 987 ± 5. LiquiLabs describes its spectral similarity scale as ranging from 0 to 1000, with higher scores indicating closer agreement with the reference spectrum. This is an identity comparison score, rather than a purity percentage.

The purpose of this separate measurement is to provide evidence about the complex alongside the peptide information obtained through LC/MS.

3. How content and purity are measured

LiquiLabs states that its quantitative assay uses HPLC with UV/VIS detection, calibrated against certified GHK-Cu reference material, with the result expressed as GHK-Cu complex content.

The results reported for this sample are:

- GHK-Cu assay: 89.4 ± 0.4 mg
- Chromatographic purity: >99.8%
- Retention-time identification result: 0.991 ± 0.005

These measurements serve different purposes. The assay reports the measured content, chromatographic purity describes the relative chromatographic peak areas under the stated method, and retention-time comparison provides additional identity information. They should be interpreted together with the FTIR and MS results.

4. Supporting spectra and further technical clarification

The complete nine-page report includes the analytical method, calibration information, chromatograms and MS spectral plots. These provide more context than the summary mass entry alone.

For questions about individual MS peak assignments, the supporting FTIR comparison or access to the original instrument data, you are welcome to contact Ján Galbavý at LiquiLabs directly at jan.galbavy@liquilabs.cz. The laboratory can explain the acquisition conditions and interpretation of its results in further detail.

You can also contact us at support@adriascience.com, and we will help coordinate any additional documentation or clarification you require.

We appreciate the opportunity to explain the methodology. Clear analytical documentation matters, and we want customers to understand both what each measurement demonstrates and how the results fit together.

Kind regards,
Adria Science Team