Microbial Metabolite

Trans - Caffeic acid shows growth - stimulating activity on 2 - mm sections of avena coleoptiles. Dilute solutions of trans - caffeic acid can significantly promote the straight growth of pea epicotyls [1]

Metabolism / Metabolites
The enzymes involved in caffeic acid metabolism have not yet been identified. In the following experiments, caffeic acid (CA), chlorogenic acid (CGA), and dihydrocaffeic acid (DHCA) were incubated with hepatocytes, and the results showed that they can be metabolized by cytochrome P450, catechol-O-methyltransferase (COMT), and β-oxidase. Ferulic acid (FA) or dihydroferulic acid (DHFA), generated by COMT O-methylation of CA or DHCA, can also be O-demethylated by CYP1A1/2, but not by CYP2E1. DHCA or DHFA also undergoes side-chain dehydrogenation reactions to generate CA and FA, respectively, which can be inhibited by thioglycolic acid (an inhibitor of β-oxidase acyl-CoA dehydrogenase). The rate of glutathione conjugate formation catalyzed by NADPH/microsomes (CYP2E1) follows a decreasing order of DHCA > CA > CGA, which is the reverse of the rate of COMT O-methylation. CA and DHCA-o-quinones generated by NADPH/P450 may inhibit COMT, but they can readily form glutathione conjugates. CA, DHCA, and DHFA intermetabolize in isolated rat hepatocytes and can be metabolized to FA, while FA is only metabolized to CA and not to DHCA or DHFA. CA, DHCA, FA, DHFA, and CGA all exhibit dose-dependent hepatotoxicity, with the measured LD50 (2 hours) arranged in decreasing order of toxicity: DHCA > CA > DHFA > CGA > FA. In summary, evidence suggests that O-methylation, GSH conjugation, hydrogenation, and dehydrogenation reactions are all involved in the hepatic metabolism of CA and DHCA. The O-methylation pathway of CA and DHCA is a detoxification pathway, while the P450-catalyzed o-quinone formation pathway is a toxic pathway. In rats, chlorogenic acid is hydrolyzed in the stomach and intestines to caffeic acid and quinic acid. Several metabolites have been identified. Glucuronide of m-coumaric acid and m-hydroxyhippuric acid appears to be the major metabolites in humans. Following oral administration of caffeic acid to human volunteers, O-methylated derivatives (ferulic acid, dihydroferulic acid, and vanillic acid) are rapidly excreted in the urine, while m-hydroxyphenyl derivatives appear later. The dehydroxylation reaction is attributed to the action of intestinal bacteria. Known metabolites of caffeic acid include (2S,3S,4S,5R)-6-[5-[(E)-2-carboxyvinyl]-2-hydroxyphenoxy]-3,4,5-trihydroxyoxacyclohexane-2-carboxylic acid and (2S,3S,4S,5R)-6-[4-[(E)-2-carboxyvinyl]-2-hydroxyphenoxy]-3,4,5-trihydroxyoxacyclohexane-2-carboxylic acid.

Interactions
Caffeic acid enhanced the uptake of radioglucose by C2C12 cells in a concentration-dependent manner. Phenylephrine had a similar effect on the uptake of radioglucose by C2C12 cells. Prazosin attenuated the effect of caffeic acid, acting in a manner similar to blocking the effect of phenylephrine. Nine-week-old female ICR/Ha mice were fed a diet containing 0.06 mmol/g (10 g/kg diet) of caffeic acid (99% purity). Starting from day 8 of the experiment, mice were administered 1 mg of benzo[a]pyrene twice weekly by gavage for 4 weeks. Three days after the final benzo[a]pyrene treatment, the feeding of caffeic acid-containing diets was discontinued. Mice were sacrificed at 211 days of age. In 17 effective mice, caffeic acid significantly reduced the number of forestomach tumors (≥ 1 mm) per mouse (histological details not specified) (p < 0.05) (3.1 tumors per mouse, compared to 5.0 tumors per mouse in 38 mice treated with benzo(a)pyrene alone).

[1]. Vendrig J C, Buffel K. Growth-stimulating activity of trans-caffeic acid isolated from Coleus rhenaltianus. Nature, 1961, 192(4799): 276-277.

According to the International Agency for Research on Cancer (IARC) of the World Health Organization, caffeic acid is potentially carcinogenic. 3,4-Dihydroxycinnamic acid appears as yellow prismatic or flaky forms, or pale yellow granules, in chloroform or petroleum ether solutions. In alkaline solutions, the color changes from yellow to orange. (NTP, 1992) Caffeic acid is a hydroxycinnamic acid formed by replacing the 3 and 4 positions of the benzene ring of cinnamic acid with hydroxyl groups. It exists in both cis and trans forms, with the trans form being more common. Caffeic acid is a plant metabolite and is also an inhibitor of EC 1.13.11.33 (arachidonic acid 15-lipoxygenase), EC 2.5.1.18 (glutathione transferase), EC 1.13.11.34 (arachidonic acid 5-lipoxygenase), an antioxidant, and EC 3.5.1.98 (histone deacetylase). It is a hydroxycinnamic acid belonging to the catechol group of compounds.
It has been reported that caffeic acid is found in Salvia miltiorrhiza, Salvia miltiorrhiza var. albopictus, and other organisms with relevant data.
Caffeic acid is a hydroxycinnamic acid derivative and polyphenol with high oral bioavailability and potential antioxidant, anti-inflammatory, and antitumor activities. After administration, caffeic acid acts as an antioxidant, preventing oxidative stress and thus preventing DNA damage caused by free radicals. Caffeic acid targets and inhibits the histone demethylase (HDM) oncoprotein gene 1 (GASC1; JMJD2C; KDM4C) that amplifies squamous cell carcinoma, thereby inhibiting cancer cell proliferation. GASC1 is a member of the KDM4 subgroup of proteins containing the Jumonji (Jmj) domain. It demethylates lysine 9 and lysine 36 (H3K9 and H3K36) on histone H3 and plays a key role in tumor cell development.
Caffeic acid is a metabolite found or produced in Saccharomyces cerevisiae.
See also: Black cohosh (partial); Lithospermum erythrorhizon root (partial). Burdock root (partial)...View more...

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Mechanism of Action
Caffeic acid phenethyl ester (CAPE) is synthesized from caffeic acid and phenylethanol (ratio 1:5) at room temperature using dicyclohexylcarbodiimide (DCC) as a condensing agent, with a yield of approximately 38%. CAPE inhibits the growth of human leukemia HL-60 cells. It also inhibits the synthesis of DNA, RNA, and proteins in HL-60 cells, with IC50 values of 1.0 M, 5.0 M, and 1.5 M, respectively.
To understand the hypoglycemic effect of caffeic acid, this study used myoblast C2C12 cells to investigate their glucose uptake. Caffeic acid enhanced the uptake of radioactive glucose by C2C12 cells in a concentration-dependent manner. A similar effect of phenylephrine on the uptake of radioactive glucose was also observed in C2C12 cells. Prazosin attenuates the effect of caffeic acid in a similar way to blocking the effect of phenylephrine. The effect of caffeic acid on α1-adrenergic receptors was further confirmed by the binding substitution of [3H]prazosin in C2C12 cells. Furthermore, the glucose uptake-enhancing effect of phenylephrine in C2C12 cells was inhibited by the α1A-adrenergic receptor antagonists tamsulosin and WB 4101, but not by the α1B-adrenergic receptor antagonist chloroethyl clonidine (CEC). Therefore, the presence of α1A-adrenergic receptors in C2C12 cells can be considered. Similar inhibition of caffeic acid effects was also observed in C2C12 cells co-incubated with these antagonists. Activation of α1A-adrenergic receptors appears to be the reason for the action of caffeic acid in C2C12 cells. In the presence of the phospholipase C-specific inhibitor U73312, caffeic acid-stimulated uptake of radioactive glucose into C2C12 cells decreased in a concentration-dependent manner, while U73343 (a negative control of U73312) did not have this effect. Furthermore, chelerythrine and GF 109203X attenuated the effects of caffeic acid at concentrations sufficient to inhibit protein kinase C. Therefore, the data suggest that caffeic acid activation of α1A-adrenergic receptors in C2C12 cells may increase glucose uptake via the phospholipase C-protein kinase C pathway. Studies have shown that 2% dietary caffeic acid (CA, 3,4-dihydroxycinnamic acid) can lead to cancer in the forestomach and kidneys of F344 rats and B6C3F1 mice. Given that caffeic acid is present in coffee and many other foods, and considering the extrapolation of cancer incidence using linear interpolation within a 0% to 2% dose range, the risk of cancer in humans is considerably high. In both target organs, tumor formation precedes proliferation, which may be the primary mechanism of its carcinogenic effect. This study investigated the dose-response relationship of CA in male F344 rats after feeding them with different dietary concentrations (0, 0.05%, 0.14%, 0.40%, and 1.64%) for 4 weeks. Two hours after intraperitoneal injection, immunohistochemical analysis was performed to observe cells in the S phase of DNA replication. In the forestomach, at concentrations of 0.40% and 1.64%, both the total number of epithelial cells per millimeter of slice length and the unit length labeling index (ULLI) of BrdU-positive cells increased by approximately 2.5-fold. No effect was observed at the lowest concentration (0.05%). At a concentration of 0.14%, both indices decreased by approximately one-third. In the kidneys, the labeling index of proximal renal tubular cells also showed a J-shaped (or U-shaped) dose-response, increasing 1.8-fold at 1.64%. No dose-related effects were observed in non-target organs—the proventriculus and liver. Data showed that there was a good correlation between cancer-induced organ specificity and cell division stimulation. Linear extrapolation does not seem appropriate in terms of dose-response relationship and extrapolating animal tumor data to human cancer risk. Transcaffeic acid may be a very important natural growth regulator with growth-promoting effects similar to indole-3-acetic acid. It is one of the active growth substances in the ether extract of Coleus leaf[1].

C9H8O4

180.16

180.042

C, 60.00; H, 4.48; O, 35.52

501-16-6

Caffeic acid; 331-39-5

689043

White to off-white solid powder

1.5±0.1 g/cm3

416.8±35.0 °C at 760 mmHg

211-213ºC (dec.)(lit.)

220.0±22.4 °C

0.0±1.0 mmHg at 25°C

1.707

1.42

3

4

2

13

212

0

C1=CC(=C(C=C1/C=C/C(=O)O)O)O

QAIPRVGONGVQAS-DUXPYHPUSA-N

InChI=1S/C9H8O4/c10-7-3-1-6(5-8(7)11)2-4-9(12)13/h1-5,10-11H,(H,12,13)/b4-2+

(E)-3-(3,4-dihydroxyphenyl)prop-2-enoic acid

Caffeic acid; AI3-63211; caffeic acid; 3,4-Dihydroxycinnamic acid; 331-39-5; 3,4-Dihydroxybenzeneacrylic acid; (E)-3-(3,4-dihydroxyphenyl)prop-2-enoic acid; Cinnamic acid, 3,4-dihydroxy-; 3-(3,4-Dihydroxyphenyl)-2-propenoic acid; ...; 501-16-6;

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

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.)