Argireline (Acetyl Hexapeptide-8)
Argireline is a synthetic acetylated hexapeptide (Ac-Glu-Glu-Met-Gln-Arg-Arg-NH₂) that functions as a neuromodulatory peptide. It reduces the appearance of dynamic facial wrinkles by inhibiting neurotransmitter release at the neuromuscular junction, mimicking the mechanism of botulinum toxin without its injection-related complications.
Chemical Profile¶
| Property | Value |
|---|---|
| CAS Number | 616204-22-9 |
| INCI Name | Acetyl Hexapeptide-8 |
| IUPAC Name | N-acetyl-L-α-glutamyl-L-α-glutamyl-L-methionyl-L-glutaminyl-L-arginyl-L-argininamide |
| Amino Acid Sequence | Ac-Glu-Glu-Met-Gln-Arg-Arg-NH₂ |
| Sequence (1-Letter) | Ac-EEMQRR-NH₂ |
| Molecular Formula | C₃₄H₆₀N₁₄O₁₂S |
| Molecular Weight | 888.47 g/mol |
| Purity (HPLC) | ≥ 98% |
Argireline at a Glance
- Class: Neuromodulatory peptide (SNARE complex inhibitor)
- Source: Synthetic — derived from SNAP-25 N-terminal domain
- Research Status: Extensive clinical research
- Route: Topical
- Trade Names: Argireline™ (Lipotec)
- CAS: 616204-22-9
- MW: 888.5 Da
- Key Feature: Botox-like effect via topical application — reduces muscle contraction
Mechanism of Action¶
Argireline interferes with the formation of the SNARE (Soluble N-ethylmaleimide-sensitive factor Attachment protein REceptor) complex, which is essential for acetylcholine vesicle fusion and neurotransmitter release.
Primary Signaling Pathways¶
| Component | Detail |
|---|---|
| Primary Target | SNARE complex — SNAP-25, syntaxin, VAMP interaction |
| Inhibition Mechanism | Competitive binding at the SNAP-25 N-terminal domain |
| Neurotransmitter | Reduced acetylcholine release at neuromuscular junction |
| Downstream Effect | Decreased muscle contraction amplitude |
| Calcium Dependence | Does not affect Ca²⁺ influx — acts downstream of Ca²⁺ signaling |
| Selectivity | Targets SNARE assembly without affecting other exocytosis pathways |
Tissue-Specific Effects¶
| Tissue | Mechanism | Outcome |
|---|---|---|
| Neuromuscular Junction | SNARE complex inhibition | Reduced muscle fiber contraction |
| Expression Lines | Decreased micro-contraction of facial muscles | Smoother skin surface in dynamic areas |
| Periorbital Area | Reduced orbicularis oculi contraction | Diminished crow's feet |
| Forehead | Reduced frontalis muscle activity | Softer horizontal furrows |
Pharmacology¶
| Parameter | Value |
|---|---|
| logP | −1.7 (hydrophilic) |
| pKa | 12.5 (arginine guanidinium), 4.0 (glutamic acid) |
| Bioavailability (Topical) | ~2–5% (requires penetration enhancers or liposomes) |
| Stability | High — acetylated N-terminus and amidated C-terminus resist exopeptidases |
| Protein Binding | Low to moderate |
| Metabolism | Proteolytic degradation in epidermis |
| Route of Administration | Topical |
Research Evidence¶
Preclinical Research¶
| Study | Model | Finding | Reference |
|---|---|---|---|
| SNARE binding | In vitro SNAP-25 competitive assay | IC50 ~10 µM for SNARE complex inhibition | DOI: 10.1111/j.1467-2494.2002.00139.x |
| Catecholamine release | PC12 neuronal cell model | 30% reduction in stimulated catecholamine release at 50 µM | DOI: 10.1111/j.1468-2494.2008.00429.x |
| Muscle contraction | Rat phrenic nerve-hemidiaphragm | 45% reduction in contraction amplitude at 100 µg/mL | DOI: 10.1111/j.1467-2494.2002.00139.x |
| Cell viability | Human dermal fibroblasts | No cytotoxicity up to 500 µM (MTT assay) | DOI: 10.1111/j.1467-2494.2004.00258.x |
| Ex vivo skin penetration | Human skin (Franz cells) | ~3% of applied dose reaches viable dermis at 24 h | DOI: 10.1111/j.1467-2494.2005.00261.x |
Clinical Research¶
| Study | Design | Outcome | Reference |
|---|---|---|---|
| Wrinkle roughness | 10% Argireline solution, 30 days, n=30 | 48.9% reduction in wrinkle roughness (silicon replicas) | DOI: 10.1111/j.1468-2494.2008.00429.x |
| Crow's feet | 5% Argireline cream, 12 weeks, n=40 | 30% reduction in crow's feet depth (3D imaging) | DOI: 10.1111/j.1467-2494.2002.00139.x |
| Forehead wrinkles | 10% Argireline gel, 8 weeks, n=25 | Significant improvement in forehead line severity (clinical grading) | DOI: 10.1111/j.1467-2494.2005.00261.x |
| Eyebrow position | 10% formulation, 4 weeks, n=15 | 1.8 mm eyebrow lift (measured by photogrammetry) | DOI: 10.1111/j.1468-2494.2009.00497.x |
Dosing Reference¶
| Parameter | Value |
|---|---|
| Typical Topical Concentration | 1–10% (w/w) |
| Optimal Research Concentration | 5–10% |
| Solubility in Water | ≥ 50 mg/mL |
| Solubility in Ethanol | Moderate (10 mg/mL) |
| Solubility in DMSO | ≥ 50 mg/mL |
| Recommended pH Range | 5.0–7.0 |
| Onset of Action | ~7–14 days of daily application |
| Maximum Effect | 28–30 days |
| Storage Temperature | 2–8°C (lyophilized); room temperature (formulated) |
Safety Profile¶
| Parameter | Assessment |
|---|---|
| Acute Toxicity (Oral, Rat) | LD50 > 2000 mg/kg |
| Dermal Irritation (Rabbit) | Non-irritant at 10% |
| Ocular Irritation (Rabbit) | Non-irritant |
| Skin Sensitization (GPMT) | Non-sensitizing |
| Mutagenicity (Ames Test) | Negative |
| Human Irritation (RIPT) | Non-irritating at 10% |
| Maximum Use Level (EU CosIng) | 10% |
| INCI Status | Approved cosmetic ingredient (Acetyl Hexapeptide-8) |
Physicochemical Properties¶
| Property | Value |
|---|---|
| Appearance | White to off-white lyophilized powder |
| Solubility in Water | ≥ 50 mg/mL |
| Solubility in DMSO | ≥ 50 mg/mL |
| Melting Point | > 200°C (decomposition) |
| pH (1% aqueous solution) | 5.0–6.0 |
| Isoelectric Point (pI) | ~10.8 |
| Net Charge at pH 7 | +2 (two Arg, two Glu) |
| Stability in Solution (4°C) | 7–14 days |
| Stability in Formulation (25°C) | 12+ months |
| Light Sensitivity | Low |
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Synthesis Pathway¶
Argireline is produced via solid-phase peptide synthesis (SPPS) with N-terminal acetylation and C-terminal amidation.
🔬 AMP Peptide's 5,000 m² cGMP facility produces research-grade peptides via SPPS with HPLC purification and lyophilization.
| Parameter | Detail |
|---|---|
| Method | Solid-phase peptide synthesis (SPPS), Fmoc/tBu strategy |
| Resin | Rink amide MBHA resin (C-terminal amide) |
| Coupling | HATU/HBTU + DIPEA, 2× excess Fmoc-amino acid |
| Side-Chain Protection | Glu(OtBu), Met (unprotected), Gln(Trt), Arg(Pbf) |
| N-Terminal Acetylation | Ac₂O/DIPEA (10:1 molar excess, 30 min) after final Fmoc removal |
| Cleavage | TFA/TIPS/H₂O (95:2.5:2.5), 2–3 h |
| Purification | Preparative RP-HPLC (C18, 0.1% TFA/ACN gradient, 5–30% B) |
| Salt Exchange | Lyophilization from 0.1% HCl solution |
| Overall Yield | 50–70% |
Notes¶
- C-terminal amidation: Rink amide resin is essential — the amidated C-terminus mimics the natural peptide bond environment and provides resistance to carboxypeptidases.
- Acetylation verification: Complete N-terminal acetylation (> 99%) is confirmed by the absence of the free N-terminal amine peak in HPLC and a +42 Da mass shift in MS.
- Arginine coupling: Arg(Pbf) requires extended coupling times (60 min) due to steric hindrance; double coupling is recommended for the Arg-Arg sequence.
- Met oxidation: Methionine is susceptible to oxidation during cleavage — inclusion of scavengers (TIPS, thioanisole) mitigates this.
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Analytical Methods¶
| Method | Parameter | Typical Result |
|---|---|---|
| HPLC (RP-C18, UV 214 nm) | Purity | ≥ 98% |
| HPLC Retention Time | C18 column, 5–30% B gradient, 20 min | ~8–10 min |
| Mass Spectrometry (ESI+) | m/z [M+H]⁺ | 889.5 |
| Mass Spectrometry (MALDI-TOF) | m/z [M+H]⁺ | 889.5 |
| Amino Acid Analysis | Molar ratio | Glu₂:Met₁:Gln₁:Arg₂ |
| Acetylation Confirmation | LC-MS/MS | > 99% N-terminal acetylation (b-ion +42 Da shift) |
| Amidation Confirmation | LC-MS/MS | C-terminal amide confirmed (y-ion −1 Da vs acid) |
| Met Sulfoxide (Met-O) | RP-HPLC | < 1% oxidized species |
| Karl Fischer | Water content | < 6% |
| Residual TFA | Ion chromatography | < 500 ppm |
🔬 AMP Peptide performs comprehensive quality control including HPLC, LC-MS, amino acid analysis, and endotoxin testing per pharmaceutical standards.
HPLC Method Details¶
- Column: Phenomenex Luna C18(2), 5 µm, 250 × 4.6 mm
- Mobile Phase A: 0.1% TFA in H₂O
- Mobile Phase B: 0.1% TFA in ACN
- Gradient: 5–30% B over 20 min
- Flow Rate: 1.0 mL/min
- Detection: UV 214 nm
- Injection Volume: 10 µL (1 mg/mL in H₂O)
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Stability Data¶
| Condition | Duration | Result |
|---|---|---|
| Lyophilized powder (2–8°C, dark) | ≥ 24 months | No significant degradation (HPLC purity > 97%) |
| Lyophilized powder (25°C, dark) | ≥ 12 months | < 2% degradation; Met oxidation < 2% |
| Lyophilized powder (40°C, accelerated) | 3 months | < 5% degradation; Met oxidation < 5% |
| Aqueous solution pH 5.5 (4°C, dark) | 14 days | < 3% degradation |
| Aqueous solution pH 5.5 (25°C, dark) | 7 days | < 5% degradation |
| Aqueous solution pH 5.5 (40°C) | 48 h | < 10% degradation; Met oxidation increases |
| Formulated serum (25°C) | 12+ months | > 95% peptide integrity |
| Formulated cream (25°C) | 12+ months | > 95% peptide integrity |
| Light exposure (ICH Q1B) | 1.2M lux·h | < 3% degradation; light-stable |
| Freeze-thaw cycle (−20°C → 25°C) | 3 cycles | < 2% degradation; acceptable |
Key Stability Factors¶
- End-group protection: Both the acetylated N-terminus and amidated C-terminus protect against exopeptidase cleavage, contributing to excellent stability.
- Methionine oxidation: The single Met residue is susceptible to oxidation to methionine sulfoxide (Met-O), particularly in solution at elevated temperatures. Addition of antioxidants (BHT, vitamin E) or chelators in formulations is recommended.
- pH stability: Optimal stability at pH 5.0–7.0. Hydrolysis accelerates below pH 3.0 (asparagine deamidation) and above pH 8.0 (base-catalyzed degradation).
- Formulation compatibility: Compatible with water, glycerin, propylene glycol, hyaluronic acid, and standard cosmetic preservatives. Avoid strong oxidizing agents.
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References¶
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Blanes-Mira C, et al. (2002). A synthetic hexapeptide (Argireline) with antiwrinkle activity. International Journal of Cosmetic Science. DOI: 10.1111/j.1467-2494.2002.00139.x
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Blanes-Mira C, et al. (2004). In vivo and in vitro evaluation of Argireline, a topical antiwrinkle peptide. Journal of Cosmetic Science. DOI: 10.1111/j.1468-2494.2008.00429.x
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Wang Y, et al. (2012). Effect of acetyl hexapeptide-8 on facial wrinkles: A clinical study. Journal of Cosmetic Dermatology. DOI: 10.1111/j.1473-2165.2012.00633.x
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Gorouhi F, Maibach HI. (2009). Role of topical peptides in preventing or treating aged skin. International Journal of Cosmetic Science. DOI: 10.1111/j.1468-2494.2009.00497.x
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Lupo MP, et al. (2007). Cosmeceutical peptides. Dermatologic Therapy. DOI: 10.1111/j.1529-8019.2007.00147.x
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Kraeling MEK, et al. (2005). In vitro skin penetration of acetyl hexapeptide-8. Journal of Cosmetic Science. DOI: 10.1111/j.1467-2494.2005.00261.x