Tripeptide-29 is the cosmetic-ingredient (INCI) name for glycyl-L-prolyl-4-hydroxy-L-proline, the Gly-Pro-Hyp triplet that repeats through collagen. It is a 285.3 Da synthetic tripeptide, confirmed in the laboratory by LC-MS and quantified by HPLC. The name itself is a registry number and encodes nothing about the structure.
- Tripeptide-29 is the INCI registry name for glycyl-L-prolyl-trans-4-hydroxy-L-proline (H-Gly-Pro-Hyp-OH), the most common triplet in type I collagen.
- The molecular formula is C12H19N3O5: average mass 285.30 g/mol, monoisotopic 285.13 Da, seen at m/z 286.1 as the protonated molecule in electrospray MS.
- Both backbone bonds are tertiary amides because proline and hydroxyproline are imino acids, so the molecule populates cis and trans conformers in solution.
- With no aromatic residues the compound absorbs only in the low-UV backbone region near 214 nm; a 280 nm trace reads zero.
- The N-terminal Gly-Pro motif predisposes solutions to diketopiperazine formation, releasing cyclo(Gly-Pro) at m/z 155.1 and free hydroxyproline.
- No USP or European Pharmacopoeia monograph exists for the compound; its regulatory identity is a cosmetic-ingredient listing, as of August 2026.
The name is a registry entry
Tripeptide-29 is not a chemist's name. It is an INCI designation, an entry in the International Nomenclature of Cosmetic Ingredients maintained by the Personal Care Products Council, and the number is a registration index. Tripeptide-1 is glycyl-histidyl-lysine. Tripeptide-29 is a different molecule entirely. Nothing in the numbering encodes composition, residue order, or anything else a chemist would want to know; consecutive numbers were assigned as peptide ingredients were registered, in the order they arrived.
What the entry denotes is the tripeptide of glycine, proline and hydroxyproline. The compound sold under the name is glycyl-L-prolyl-trans-4-hydroxy-L-proline, written in peptide shorthand as H-Gly-Pro-Hyp-OH. The EU's CosIng ingredient database carries the same composition. Collagen researchers write the same triplet as GPO, using the single-letter code O that the collagen field assigned to hydroxyproline long ago.
The practical consequence for a buyer is blunt. A listing that names only Tripeptide-29 has identified a registry entry, and a registry entry is a pointer rather than a proof. Documentation for a specific lot should state the sequence in full, name the stereochemistry of the hydroxyproline, and back both with a measured mass. The certificate is the thing the trade name should point to.
Structure and bonding
Three residues, two peptide bonds, and one detail that shapes everything else: two of the three residues are imino acids. Proline and hydroxyproline carry their backbone nitrogen inside a five-membered pyrrolidine ring, so that nitrogen is a secondary amine. When either residue accepts a peptide bond the resulting amide is tertiary, with no N-H at all. Both backbone bonds in this molecule are of that kind.
A laboratory will actually see the consequences. Tertiary amides lack the hydrogen-bond donor that ordinary peptide bonds carry, which is part of why proline disrupts regular protein structure. And amide bonds to proline-type nitrogens interconvert between cis and trans geometries slowly, on timescales of seconds to minutes near room temperature. A short peptide with two such bonds exists in solution as a small population of conformers, and chromatography run cold enough can resolve them into a broadened or doubled peak that looks alarmingly like an impurity and is not one.
| Property | Value |
|---|---|
| Sequence | H-Gly-Pro-Hyp-OH (glycyl-L-prolyl-trans-4-hydroxy-L-proline) |
| Molecular formula | C12H19N3O5 |
| Average mass | 285.30 g/mol |
| Monoisotopic mass | 285.13 Da |
| Expected ESI-MS ions | [M+H]+ at m/z 286.1; [M+Na]+ at m/z 308.1 |
| Ionizable groups | N-terminal amine and C-terminal carboxyl only; zwitterionic near neutral pH |
| UV behaviour | Backbone amide absorbance near 214 nm; nothing at 280 nm |
The molecule is otherwise unremarkable in the best way. No aromatic rings, no thiols, no amide side chains, no basic side chains. It is small, very hydrophilic and freely water-soluble, which simplifies handling and complicates chromatography, as covered below.
Why this triplet: the collagen repeat
Collagen's defining sequence pattern is (Gly-X-Y)n. Glycine occupies every third position because the three chains of the triple helix pack too tightly at their shared axis to admit any side chain, and glycine is the only residue without one. The X position is frequently proline and the Y position is frequently 4-hydroxyproline. Sequence surveys of type I collagen put Gly-Pro-Hyp at roughly one triplet in ten, the most common single triplet in the protein. Tripeptide-29 is that repeat unit, isolated.
In the body, hydroxyproline enters collagen after translation. Ribosomes install proline, and the enzyme prolyl 4-hydroxylase then oxidizes Y-position prolines on the assembled chain, using iron, 2-oxoglutarate, molecular oxygen and ascorbate as its supporting cast. Scurvy is what collagen chemistry looks like when the ascorbate is missing. A synthetic tripeptide takes a shorter road: hydroxyproline is a stock protected building block in peptide synthesis and is coupled directly, no enzyme involved.
Why the hydroxyl matters was settled in the late 1990s. Work from the Raines laboratory replaced 4-hydroxyproline with 4-fluoroproline in collagen model peptides and found the fluorinated version formed an even more stable triple helix, which ruled out the older water-bridge explanation and established a stereoelectronic effect: the electron-withdrawing 4R substituent biases the ring pucker, and with it the backbone, toward the conformation the helix needs.
One honest caveat belongs here. A single Gly-Pro-Hyp unit does not form a triple helix. Helix formation requires long repeats and three associated strands folding cooperatively. In water, the isolated tripeptide is a flexible small molecule, whatever a marketing illustration implies.
Confirming identity by mass spectrometry
Electrospray LC-MS is the workhorse. The protonated molecule appears at m/z 286.1, usually with a sodium adduct at 308.1 alongside. Two questions are settled almost instantly. Whether the peptide is the right size: any deletion impurity from synthesis is short by a whole residue, 57 Da for glycine, 97 for proline, 113 for hydroxyproline, which are enormous gaps at this scale. And whether the hydroxylation is present at all: hydroxyproline outweighs proline by 16 Da, so an unhydroxylated Gly-Pro-Pro impurity announces itself at 270.1.
What an intact mass cannot settle is order. Gly-Pro-Hyp, Gly-Hyp-Pro and Pro-Gly-Hyp share one formula and one mass. Distinguishing them takes tandem MS, where the b- and y-fragment series place each residue, or chromatographic comparison against an authentic standard. The same limit applies one level down: 4-hydroxyproline and 3-hydroxyproline are isomers, and a mass spectrometer alone cannot say which of the two a vial contains. A retention match against a known standard, or NMR, closes that gap.
Amino acid analysis is the older cross-check and still a good one. Acid hydrolysis followed by quantitation should return glycine, proline and hydroxyproline in a 1:1:1 ratio. Hydroxyproline assays are among the most practiced measurements in protein chemistry, since Hyp content is how collagen itself has been quantified for decades.
Reading purity claims for a very small peptide
The standard purity instrument for peptides is reversed-phase HPLC on a C18 column, and this compound is close to a worst case for it. With no hydrophobic side chains beyond its two rings, the molecule barely retains. Run on a routine gradient it can elute at or near the void volume, in the same region as salts and the injection front. A purity percentage computed from a peak sitting there is soft evidence, whatever the number says.
A method that means something looks different: an ion-pairing acid such as TFA in the mobile phase, a gradient starting almost fully aqueous, and a chromatogram showing the analyte retained and resolved well clear of the void. HILIC, which retains by hydrophilicity, is the natural alternative for a molecule like this and increasingly common in short-peptide QC. Detection runs at low wavelength, around 214 nm, because the backbone amide is the only chromophore present. A 280 nm trace of this compound is flat, and a concentration estimated from 280 nm absorbance would read zero.
Chromatographic purity is also not mass. The powder in a vial is a salt, usually trifluoroacetate or acetate left over from purification, plus bound water. With a single basic site, a stoichiometric TFA salt is about 28 percent TFA by mass; an acetate salt is about 17 percent. A vial that is 98 percent pure by HPLC can be 70-something percent peptide by weight, and both numbers are correct at the same time. A certificate should state them separately, and the quality standard page sets out the documentation expected here.
Handling, storage and solution behaviour
As a dry solid, Gly-Pro-Hyp is an easy peptide to keep. It carries none of the residues that dominate peptide degradation stories: no methionine, cysteine or tryptophan to oxidize, no asparagine or glutamine to deamidate. The general rules in the storage and stability guide still apply, sealed, cold, dark and desiccated, because moisture uptake is generic to lyophilized solids rather than specific to fragile sequences. Let a cold vial reach room temperature before opening it. The condensation argument does not care how sturdy the molecule is.
Solutions are where this compound has one specific vulnerability worth knowing by name. Peptides with proline in the second position are the textbook case for diketopiperazine formation: the free N-terminal amine curls back and attacks the second backbone bond, clipping off the first two residues as a stable cyclic dipeptide. For this sequence the products are cyclo(Gly-Pro), mass 154.2, and free hydroxyproline. In an aged solution the signature is a new early-eluting peak and an ion at m/z 155.1 that was absent at preparation. The reaction is slow at refrigerator temperature and near-neutral pH, faster warm and at mildly basic pH, and it is one more reason prepared solutions get dated and used within a defined window rather than kept around indefinitely.
Preparation arithmetic is ordinary lab documentation. Bringing 5 mg into 1 mL of diluent gives:
5 mg ÷ 1 mL = 5 mg/mL, which at 285.3 g/mol is 17.5 mM
a concentrated stock by any measure for a molecule this small. The vial concentration calculator handles other volumes, and the bacteriostatic water guide covers diluent choice for a vial that will be entered more than once.
FOR LABORATORY AND IN-VITRO RESEARCH USE ONLY. NOT FOR HUMAN OR ANIMAL CONSUMPTION. NOT FOR PERSONAL, MEDICAL, DIAGNOSTIC, THERAPEUTIC, OR RECREATIONAL USE.
Regulatory status and documentation
Tripeptide-29 is a cosmetic-ingredient designation, and that is the extent of its regulatory identity. It appears in the INCI dictionary and in the EU's CosIng database as a cosmetic ingredient. It is not an approved drug in any jurisdiction this article is aware of, and no USP or European Pharmacopoeia monograph exists for it, so no compendial specification defines what a compliant lot looks like. That is the picture as of August 2026; anyone who needs the current status should check the databases directly rather than trust a page with a date on it.
The absence of a monograph moves the weight onto the certificate of analysis. For this compound, a certificate worth the name states the sequence in full with stereochemistry, reports an observed mass alongside the theoretical 285.3, shows a chromatogram in which the analyte is retained rather than parked at the void, names the counterion, and reports water content or net peptide content. Those five lines are what make a small, cheap, easily synthesized tripeptide verifiable. A certificate missing them leaves the material asserted rather than measured.
Common questions
Is Tripeptide-29 the same as collagen tripeptide?
How does Tripeptide-29 differ from Tripeptide-1?
Why does the mass spectrometer show 286 when the listed mass is 285.3?
Can HPLC alone confirm what the compound is?
Does the isolated tripeptide have collagen's triple-helix structure?
What should an aged solution of this peptide look like analytically?
Sources
- INCI Dictionary (Personal Care Products Council) and the EU CosIng database entry for Tripeptide-29. Establish the registry name and its assigned composition of glycine, proline and hydroxyproline.
- PubChem entry for glycyl-prolyl-hydroxyproline. Molecular formula, average and monoisotopic masses used throughout this article.
- Holmgren et al., Nature, 1998. 4-fluoroproline substitution in collagen model peptides, establishing the stereoelectronic basis of hydroxyproline's stabilization of the triple helix.
- Pharmaceutical peptide degradation literature on diketopiperazine formation. Documented susceptibility of peptides with proline in the second position; described generically because no single paper is load-bearing for the claim.