Reactive oxygen species (ROS) - AZ neutralizes peroxyl radicals, hydroxyl radicals, superoxide anion radicals, singlet oxygen, and peroxynitrite. [2]
Cyclobutane pyrimidine dimers (CPDs) - AZ reduces dark-CPD formation in melanocytes. [2]
Extracellular matrix (ECM) genes - AZ upregulates core matrisome genes including collagens, proteoglycans, and ECM glycoproteins. [1]
Acetyl zingerone exerts its pharmacological effects by modulating several signaling pathways, with core mechanisms centered on promoting mitophagy and inhibiting programmed cell death. In osteoarthritis research, AZ activates the PINK1/Parkin signaling pathway to promote mitophagy, facilitating the clearance of damaged mitochondria and reducing reactive oxygen species production. Concurrently, AZ effectively inhibits NLRP3 inflammasome activation, thereby suppressing chondrocyte pyroptosis (an inflammatory form of programmed cell death) and promoting collagen synthesis. Additionally, AZ inhibits ferroptosis through activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) signaling pathway, promoting the expression of glutathione peroxidase 4 (GPX4). In skin photoprotection, AZ inhibits the persistent formation of cyclobutane pyrimidine dimers in melanocytes following UV irradiation and exerts antioxidant effects by scavenging multiple reactive oxygen and nitrogen species, including hydroxyl radicals, peroxynitrite, and singlet oxygen. Furthermore, AZ inhibits matrix metalloproteinase activity and downregulates IL-17A target gene expression.

- Acetyl zingerone (AZ) (25 μg/mL, added immediately after UVA irradiation) significantly reduced dark cyclobutane pyrimidine dimer (CPD) formation in melanocytes. In untreated controls, dark-CPDs increased by approximately 40% after irradiation, while AZ treatment reduced dark-CPDs by about 82% within the first hour post-irradiation. AZ did not interfere with normal DNA repair mechanisms. Cell viability was unaffected by UVA exposure or AZ incubation. [2]
- In keratinocytes, pre-incubation with AZ (25 or 50 μg/mL) prior to UVA exposure (30 kJ/m² for 25 min) significantly reduced intracellular ROS levels in a dose-dependent manner: 35% reduction at 25 μg/mL and 46% reduction at 50 μg/mL, as measured by DCFH-DA fluorescence assay. [2]
- AZ exhibited superior efficacy compared to α-tocopherol in neutralizing various reactive species: peroxyl radical (9-fold more effective), hydroxyl radical (3.5-fold), superoxide anion radical (10-fold), singlet oxygen (2.7-fold), and peroxynitrite (127-fold). [2]
- AZ acts primarily as a physical quencher of singlet oxygen. After 10 minutes of exposure to singlet oxygen, 82% of AZ remained intact, while only 35% of α-tocopherol remained. [2]
- AZ showed high photostability: after exposure to 130 kJ/m² solar-simulated UV radiation for 30 minutes, 87% of AZ remained, whereas α-tocopherol was completely degraded. [2]
- In reconstituted human epidermis (RHE), AZ (50 μg/mL, 24 h treatment) altered gene expression. AZ-increased genes were associated with clathrin coat assembly, protein glycosylation, and positive regulation of Notch and ERK1/ERK2 signaling. AZ-decreased genes were associated with inflammatory response, reactive oxygen species metabolism, and TGF-β signaling. AZ downregulated AP-1 genes (FOS: FC=0.48; FOSB: FC=0.69). [1]
- AZ specifically upregulated core matrisome genes including collagens (COL11A1, COL11A2, COL5A1, COL6A3), proteoglycans (OGN, VCAN, PODN), and ECM glycoproteins (MGP, GLDN, NOV). RT-PCR confirmed COL11A1 expression increased by 33% (P=0.038). [1]
- In human dermal fibroblasts, AZ (10-200 μg/mL for 6 days) increased protein abundance per cell density for collagens (type I, IV, VI), fibromodulin, TGF-β, fibronectin, TIMP-1, and vimentin in a dose-dependent manner (P<0.05). [1]
- In skin biopsy explants, AZ (10 μL/cm², once daily for 4 days) resulted in 2-fold greater dermal collagen as measured by trichrome stain quantification. [1]
- AZ downregulated MMP3 expression (microarray FC=0.38, P=0.009; RT-PCR FC=0.29, P=0.096) and upregulated TIMP2 expression (RT-PCR FC=1.19, P=0.032). AZ also inhibited MMP-1, MMP-3, and MMP-12 activity in fluorescence signal assays. [1]
- AZ opposed gene expression patterns associated with fibroblast senescence, keratinocyte differentiation (decreased KRT5, KRT14, KRT1, DSC1, FLG, LOR expression), and IL-17A stimulation. [1]
- AZ decreased expression of NF-κB transcription factors REL (FC=0.61, P=0.013) and RELA (FC=0.62, P=0.012). [1]

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In cell-free systems, acetyl zingerone exhibits卓越的 multifunctional antioxidant performance as assessed by chemical assays. Using Trolox equivalent antioxidant capacity assays, AZ neutralizes various reactive oxygen species with significantly greater efficiency than the classic antioxidant α-tocopherol (vitamin E): approximately 17.7-fold more effective against peroxyl radicals, approximately 39.6-fold more effective against hydroxyl radicals, up to 126.6-fold more effective against peroxynitrite, and approximately 2.7-fold more effective against singlet oxygen. Furthermore, AZ demonstrates superior physical quenching capacity for singlet oxygen and excellent molecular photostability. Under simulated daylight UV irradiation, AZ exhibits significantly less degradation compared to α-tocopherol, indicating its superior stability as a photoprotective agent.

Acetyl zingerone demonstrates clear pharmacological activities in multiple in vivo disease models. In a mouse model of osteoarthritis, intra-articular injection of AZ significantly alleviates cartilage degeneration, reduces osteophyte formation, and inhibits synovial inflammation, with the therapeutic effects closely associated with the promotion of PINK1/Parkin-mediated mitophagy and suppression of NLRP3 inflammasome-mediated chondrocyte pyroptosis. In skin photoprotection, topical application of AZ to hairless mice prior to exposure to simulated solar ultraviolet radiation significantly reduces the formation of sunburn cells, inhibits skin thickening, and persistently decreases the number of cyclobutane pyrimidine dimers in the epidermis. Furthermore, AZ exhibits in vivo effects in delaying skin aging, as topical application effectively inhibits UV-induced matrix metalloproteinase-1 and matrix metalloproteinase-9 activity, reduces collagen degradation, and downregulates the expression of the inflammatory cytokine IL-17A target genes. Regarding metabolic function, although in vivo data for the parent compound are limited, studies on related analogues suggest that AZ may possess some potential for regulating blood glucose and lipid metabolism.

- Singlet oxygen quenching assay: AZ or α-tocopherol was tested for singlet oxygen quenching ability using a chemical system that generates singlet oxygen. The percentage of product remaining after 2, 5, and 10 minutes was determined by HPLC. AZ showed 82% remaining at 10 minutes, while α-tocopherol showed 35% remaining. [2]
- Peroxyl radical scavenging assay: Antioxidant capacity against peroxyl radicals was measured using standard chemical assays. AZ was 9-fold more effective than α-tocopherol. [2]
- Hydroxyl radical scavenging assay: AZ was 3.5-fold more effective than α-tocopherol in neutralizing hydroxyl radicals. [2]
- Superoxide anion radical scavenging assay: AZ was 10-fold more effective than α-tocopherol in neutralizing superoxide anion radicals. [2]
- Photostability assay: AZ and α-tocopherol were dissolved separately in 50% aqueous ethanol and ethanol, respectively, and placed in vials. Samples were irradiated with 13 J/cm² using a photochemical reactor equipped with UVB and UVA lamps. Degradation was determined by HPLC. AZ retained 87% of its molecular structure after 30 minutes of solar-simulated UV exposure, while α-tocopherol was completely degraded. [2]

- Dark-CPD formation assay in melanocytes: Melanocytes were exposed to UVA radiation. Immediately after irradiation, AZ (25 μg/mL) was added to the culture medium. CPD formation was measured by ELISA at various time points (0, 0.5, 1, 2, 3, 4, 5, 6 hours post-irradiation). AZ significantly reduced dark-CPD formation within the first 2 hours post-irradiation compared to untreated controls. [2]
- Intracellular ROS measurement in keratinocytes: Keratinocytes were pre-incubated with AZ (25 or 50 μg/mL) for 1 hour, then exposed to UVA radiation (30 kJ/m² for 25 min). Intracellular ROS was quantified using the DCFH-DA fluorescence assay. Fluorescence was measured to determine ROS levels. AZ reduced ROS formation by 35% (25 μg/mL) and 46% (50 μg/mL) compared to untreated controls. [2]
- Microarray analysis of gene expression: Reconstructed human epidermis (RHE) tissues were treated with AZ (50 μg/mL) or vehicle for 24 hours. RNA was extracted and hybridized to Agilent human 8X60K arrays. Data were normalized and analyzed for differential expression (P<0.05, FC>1.25 or FC<0.80). Genes were annotated for biological processes and matrisome classification. [1]
- Real-time quantitative PCR (RT-PCR): RNA from RHE cultures was reverse transcribed, and gene expression was quantified using SYBR Green qPCR with HPRT1 or RPL13A as reference genes. Fold change was calculated using the 2-ΔΔCt method. [1]
- ELISA for ECM proteins in fibroblasts: Human dermal fibroblasts were incubated with AZ (10-200 μg/mL) for 6 days. Cell media were collected, and target proteins (collagen I, IV, VI, fibromodulin, TGF-β, fibronectin, TIMP-1, vimentin) were quantified by sandwich ELISA. Signals were standardized to cell density measured by sulforhodamine B assay. Dose-response slopes were calculated. [1]
- MMP activity inhibition assay (fluorescence): MMP-1 and MMP-3 activity was measured using a fluorescent substrate. AZ or Z was serially diluted, incubated with MMP-1 or MMP-3 (100 ng/mL), and substrate (2.25 μM). Fluorescence was monitored at Ex485/Em530 nm for 1 hour. IC50 values were determined: AZ MMP-1 IC50 = 1.065 mg, Z MMP-1 IC50 = 0.705 mg; AZ MMP-3 IC50 = 0.505 mg, Z MMP-3 IC50 = 0.995 mg. For MMP-12, a colorimetric assay using N-Succinyl-Ala-Ala-Val-Ala p-nitroanilide substrate was used. AZ MMP-12 IC50 = 0.255 mg, Z MMP-12 IC50 = 0.295 mg. [1]

- Ex vivo human skin biopsy study: Human facial skin explants (from cosmetic surgery) were partitioned on gauze in culture medium. AZ or placebo (10 μL/cm²) was applied once daily for 4 days. Tissues were fixed in formalin, processed, and stained with hematoxylin/eosin or trichrome stain for collagen visualization. Trichrome stain background-adjusted intensity was quantified using ImageJ software. AZ treatment resulted in 2-fold greater dermal collagen compared to placebo. [1]

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Using a mouse model of osteoarthritis as an example, a typical in vivo efficacy evaluation procedure for acetyl zingerone is as follows: Osteoarthritis is induced in C57BL/6 mice by destabilization of the medial meniscus surgery. Animals are divided into a model control group and an AZ treatment group, with AZ administered via intra-articular injection (dose as specified by the study design), along with a sham-operated control group. Following several weeks of treatment, animals are euthanized and knee joint tissues are collected. Histopathological changes in articular cartilage, cartilage matrix degradation, and osteophyte formation are assessed by hematoxylin-eosin staining, Safranin O-fast green staining, and Masson‘s trichrome staining. The expression levels of relevant proteins such as PINK1, Parkin, NLRP3, and GPX4 in articular cartilage are detected by immunohistochemistry or Western blot. In skin photoprotection studies, AZ is typically applied topically to animal skin prior to UV irradiation, and its protective effects are evaluated through skin histological analysis and detection of DNA damage markers.

Systematic pharmacokinetic studies on acetyl zingerone are relatively limited, with existing information primarily derived from its safety assessment as a skincare ingredient. AZ is a small molecule (molecular weight 236.26) with moderate lipophilicity (LogP approximately 1.74), which facilitates its ability to penetrate the stratum corneum of the skin. In vitro skin penetration studies have demonstrated favorable transdermal absorption characteristics of AZ, making it suitable for topical formulations. When administered via intra-articular injection, AZ can achieve effective concentrations locally within the joint to exert its therapeutic effects. As a topical ingredient, the amount of AZ absorbed into systemic circulation following dermal application is limited, consistent with its favorable local safety profile. Systematic oral pharmacokinetic studies have not been reported in detail, and further investigations are needed to elucidate its absorption, distribution, metabolism, and excretion characteristics.

- Acetyl zingerone (AZ) displayed minimal absorption of UVB rays with virtually no absorption of UVA radiation, indicating it does not function as a photosensitizer during UVA exposure. [2]
- Cell viability was unaffected by UVA exposure or by incubation with AZ (25 μg/mL) in melanocytes. [2]
- No toxicity data are reported; however, AZ was used in ex vivo human skin explants at 10 μL/cm² without reported adverse effects. [1]

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Available toxicological studies indicate that acetyl zingerone possesses a favorable safety profile, particularly as a topical ingredient. According to the Material Safety Data Sheet (MSDS), AZ is classified as a non-hazardous substance with no GHS hazard classification. The U.S. Environmental Protection Agency (EPA) has assessed that AZ is not likely to be mutagenic in humans. AZ is registered with the European Chemicals Agency (ECHA) (EC No. 820-605-0), with relevant safety documentation available for review. Regarding ecotoxicology, there is limited evidence of environmental toxicity, and it is recommended to avoid release into the environment. Systematic chronic toxicity and carcinogenicity data for AZ are currently incomplete. AZ has been approved for use in cosmetic and skincare products as a safe photoprotective and anti-aging ingredient. It should be noted that existing toxicological data primarily support the safety of topical administration, whereas systematic toxicological evaluations for oral or injectable routes of administration still need to be developed.

[1]. A Zingerone Analog, Acetyl Zingerone, Bolsters Matrisome Synthesis, Inhibits Matrix Metallopeptidases, and Represses IL-17A Target Gene Expression. J Invest Dermatol. 2020 Mar;140(3):602-614.e15.

[2]. Acetyl zingerone: An efficacious multifunctional ingredient for continued protection against ongoing DNA damage in melanocytes after sun exposure ends. Int J Cosmet Sci. 2020 Feb;42(1):36-45.

- Acetyl zingerone (AZ) was designed by combining structural features of zingerone and curcumin. It contains a phenolic hydroxyl group (hydrogen atom donor for antioxidant activity), a methoxy group (singlet oxygen quencher), and a 2,4-diene branching chain (hydrogen bond chelator for peroxynitrite scavenging). [2]
- AZ exists in solution as keto-enol tautomers. It has a purity >99%, melting point 78-80°C, and UV λmax 280 nm (ε 5,501 M⁻¹ cm⁻¹ in ethanol). [2]
- The multifunctional properties of AZ allow it to intervene in the chemiexcitation pathway of dark-CPD formation: (1) scavenging peroxynitrite before it reacts with melanin fragments, and (2) antioxidant reduction of dioxetane intermediates to vicinal diols, thereby preventing formation of excited triplet carbonyls needed for energy transfer to DNA. [2]
- AZ is more photostable than α-tocopherol, retaining 87% of its molecular structure after 30 minutes of solar-simulated UV exposure compared to complete degradation of α-tocopherol. [2]
- In gene expression studies, AZ opposed expression patterns associated with fibroblast senescence, keratinocyte differentiation (exhibiting retinoid-like effects), and IL-17A stimulation. AZ decreased expression of NF-κB transcription factors REL and RELA. [1]

C13H16O4

236.26

236.10485899

30881-23-3

207830

White to off-white solids at room temperature

1.15g/cm3

375.6ºC at 760 mmHg

139.9ºC

1.524

1.738

1

4

5

17

273

0

COC1C=C(CC(C(=O)C)C(=O)C)C=CC=1O

IFGDRUTWCITGHG-UHFFFAOYSA-N

InChI=1S/C13H16O4/c1-8(14)11(9(2)15)6-10-4-5-12(16)13(7-10)17-3/h4-5,7,11,16H,6H2,1-3H3

3-[(4-hydroxy-3-methoxyphenyl)methyl]pentane-2,4-dione

3-[(4-hydroxy-3-methoxyphenyl)methyl]pentane-2,4-dione; 3-[(4-Hydroxy-3-methoxyphenyl)methyl]-2,4-pentanedione; 3-((4-Hydroxy-3-methoxyphenyl)methyl)-2,4-pentanedione; 3-((4-Hydroxy-3-methoxyphenyl)methyl)pentane-2,4-dione; ...; acetyl zingerone;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO: ~100 mg/mL (423.3 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)