Safrole Research Articles

Formation of RNA adducts resulting from metabolic activation of spice ingredient safrole mediated by P450 enzymes and sulfotransferases.

Safrole (SFL) is an IARC class 2B carcinogen. To better understand the mechanism involved in SFL toxicity, we explored the potential interactions between SFL metabolites and RNA. Three guanosine adducts (G1-G3), two adenosine adducts (A1-A2), and two cytosine adducts (C1-C2) were detected by LC-MS/MS in mouse liver S9 incubations, cultured mouse primary hepatocytes, and liver tissues of mice after exposure to SFL. These adducts were chemically synthesized, and one of the guanosine adducts was structurally characterized by 1H-NMR. Studies in vitro and in vivo showed that SFL was oxidized by cytochrome P450 enzymes to the corresponding 1'-hydroxyl metabolite which was further metabolized by sulfotransferases to form allylic sulfate esters. The formed reactive intermediate(s) subsequently reacted with bases of RNA, leading to RNA adduction, which could play a partial role in the toxicities of SFL through the alteration of RNA biochemical properties and interruption of RNA functions.

Safranal inhibits estrogen-deficiency osteoporosis by targeting Sirt1 to interfere with NF-κB acetylation

BackgroundOsteoporosis is a prevalent bone metabolic disease in menopause, and long-term medication is accompanied by serious side effects. Estrogen deficiency-mediated hyperactivated osteoclasts is the initiating factor for bone loss, which is regulated by nuclear factor-κB (NF-κB) signaling. Safranal (Saf) is a monoterpene aldehyde produced from Saffron (Crocus sativus L.) and possesses multiple biological properties, particularly the anti-inflammatory property. However, Saf's role in osteoporosis remains unknown. PurposeThis study aims to validate the role of Saf in osteoporosis and explore the potential mechanism. Study DesignThe RANKL-exposed mouse BMM (bone marrow monocytes) and the castration-mediated osteoporosis model were applied to explore the effect and mechanism of Saf in vitro and in vivo. MethodThe effect of Saf on osteoclast formation and function were assessed by TRAcP staining, bone-resorptive experiment, qPCR, immunoblotting and immunofluorescence, etc. Micro-CT, HE, TRAcP and immunohistochemical staining were performed to estimate the effects of Saf administration on OVX-mediated osteoporosis in mice at imaging and histological levels. ResultsSaf concentration-dependently inhibited RANKL-mediated osteoclast differentiation without affecting cellular viability. Meanwhile, Saf-mediated anti-osteolytic capacity and Sirt1 upregulation were also found in ovariectomized mice. Mechanistically, Saf interfered with NF-κB signaling by activating Sirt1 to increase p65 deacetylation and inactivating IKK to decrease IκBα degradation. ConclusionOur results support the potential application of Saf as a therapeutic agent for osteoporosis.

DFT study of the antiradical properties of some aromatic compounds derived from antioxidant essential oils: C–H bond vs. O–H bond

This paper describes a theoretical comparative study of the antiradical properties of six aromatic compounds namely eugenol (EUG), safrole (SAF), myristicin (MYR), carvacrol (CAR), cinnamaldehyde (CIN), and isoeugenol (ISO) found in antioxidant essential oils. Using density functional theory (DFT) calculations, some structural characteristics such as molecular descriptors, frontier molecular orbitals, and molecular electrostatic potential have been studied. Three main antiradical mechanisms, hydrogen atom transfer (HAT), single electron transfer proton transfer (SETPT, and sequential proton loss electron transfer (SPLET) have been also investigated. In addition, the Gibbs free energies related to the reactions of the studied compounds with two reactive oxidant species (HO• and HOO•) have been computed. Throughout the study, the implicitly of polar and nonpolar solvents (water and benzene) has been taken into account. It was found that EUG, SAF, CIN, and ISO scavenge free radicals by means of a CH bond, while CAR and ISO by means of an OH bond. In the gas and benzene phases, all the studied compounds prefer to undergo HAT mechanism, while in water, SPLET is more favoured for CIN, CAR, and ISO and both HAT and SPLET are possible for EUG, SAF, and MYR. In all the studied mediums, the sequence of the antiradical potential is: MYR ≈ SAF ≈ ISO ≈ EUG > CAR > CIN. This order is in line with the available experimental results.

Comparative analysis of phenolic profile of Monodora myristica and Monodora tenuifolia

This study sought to identify phenolics present in the seeds of Monodora myristica and Monodora tenuifolia by Gas chromatography (GC) coupled to Flame ionization detector (FID). GC-FID analysis identified fifty-three different types of phenolics in both M. myristica and M. tenuifolia seeds. Predominant phenolics are Myristicin (RT: 13.78), Caffeic acid (RT: 14.16), Safrole (RT: 11.37), Methyl Eugenol (RT: 12.32), Catechin (RT: 3.63), Elemicin (RT: 13.69), Quercetin (RT: 25.05), Kaempherol (RT: 21.52), Methyl Isoeugenol (RT: 12.69) and Eugenol (RT: 11.70). It was observed that M. myristica is rich in Myristicin (42.60%), Caffeic acid (23.39%), Elemicin (3.82%) and Eugenol (1.02%), while M. tenuifolia is rich in Safrole (11.86%), Methyl Eugenol (6.28%), Catechin (5.27%), Quercetin (2.97%), Kaempherol (2.27%) and Methyl Isoeugenol (1.45%). However, the two species of Monodora contained virtually the same kinds of phenolics but in varying quantities with M. myristica having a higher total phenolic content (1478.32 mg/100 g) than M. tenuifolia (1171.52 mg/100 g). Both species are promising sources of phenolics. Key words: Monodora myristica, Monodora tenuifolia, phenolics.

Electrochemical Oxidation of Safrole Using Ti/Ru X Ti(1 − X)O2 System: Preparation, Characterization, and Role of Electrode Composition

Oxidation processes introduce structural changes in organic substrates. Electrooxidation is an alternative synthetic route that can help scientists conduct desired changes in organic compounds. In this study, we investigated the Ti/Ru X Ti(1 − X)O2 system and used safrole (SF) as model substrate, to understand the role played by electrode composition in organic substances oxidation. We characterized the surface of the prepared electrodes and conducted controlled potential electrolysis with potential pulse, which oxidized 98 % SF. Electrode efficiency depended on the electrode composition—higher RuO2 concentration led to a more efficient process. Our results showed that all the electrodes generated the same intermediates regardless of the composition, indicating that the same reaction mechanism occurred in all cases, as corroborated by mechanistic parameters. Electrolysis under controlled current generated the same intermediates as electrolysis under controlled potential. The main difference between these two types of electrolyses was that controlled current promoted at least five times faster reaction, which resulted in substrate mineralization, an outcome that is not interesting for the synthetic process.

Development of a Medium-term Animal Model Using <i>gpt</i> Delta Rats to Evaluate Chemical Carcinogenicity and Genotoxicity

In this study, the potential for development of an animal model (GPG46) capable of rapidly detecting chemical carcinogenicity and the underlying mechanisms of action were examined in gpt delta rats using a reporter gene assay to detect mutations and a medium-term rat liver bioassay to detect tumor promotion. The tentative protocol for the GPG46 model was developed based on the results of dose-response exposure to diethylnitrosamine (DEN) and treatment with phenobarbital over time following DEN administration. Briefly, gpt delta rats were exposed to various chemicals for 4 weeks, followed by a partial hepatectomy (PH) to collect samples for an in vivo mutation assay. The mutant frequencies (MFs) of the reporter genes were examined as an indication of tumor initiation. A single intraperitoneal (ip) injection of 10 mg/kg DEN was administered to rats 18 h after the PH to initiate hepatocytes. Tumor-promoting activity was evaluated based on the development of glutathione S-transferase placental form (GST-P)-positive foci at week 10. The genotoxic carcinogens 2-acetylaminofluorene (2-AAF), 2-amino-3-methylimidazo [4,5-f] quinolone (IQ) and safrole (SF), the non-genotoxic carcinogens piperonyl butoxide (PBO) and phenytoin (PHE), the non-carcinogen acetaminophen (APAP) and the genotoxic non-hepatocarcinogen aristolochic acid (AA) were tested to validate the GPG46 model. The validation results indicate that the GPG46 model could be a powerful tool in understanding chemical carcinogenesis and provide valuable information regarding human risk hazards.

In vitro responses and production of phytochemicals of potential medicinal value in nutmeg, Myristica fragrans Houtt

The in vitro production of phytochemicals by the embryogenic cultures of nutmeg- Myristica fragrans Houtt., a tropical tree of considerable commercial and medicinal value, was investigated. The regeneration potential of the somatic embryos and the morphogenetic responses of foliar explants were also studied. The metabolite profiling of the long term embryogenic cultures established from zygotic embryos in media with activated charcoal, revealed the presence of several monoterpenes and essential oils including α&β-pinene, myristicin, safrole, methyl eugenol and betasitosterol. The essential oil profile varied with the age of the cultures and the 3 week old cultures had a rich variety of monoterpenes. The spent charcoal medium of these embryogenic cultures exhibited strong anti-microbial activity against the pathogens Salmonella typhi and Staphylococcus aureus . This is considered to be the first report of in vitro production of phytochemicals by the embryogenic mass. The results obtained in this study suggest that both the embryogenic cultures and the spent charcoal medium can act as sources of products of pharmacological interest and provide a basis for further biotechnological investigations of this rare, medicinally important species and for conservation of its germplasm by cryopreservation. Germination of somatic embryos derived from zygotic embryos was achieved in media with NAA. Direct formation of somatic embryos was achieved in leaf explants from juvenile plants in MS media with kinetin, 2,4-D and NAA.

Abuse of Nutmeg (Myristica fragrans Houtt.): Studies on the Metabolism and the Toxicologic Detection of its Ingredients Elemicin, Myristicin, and Safrole in Rat and Human Urine Using Gas Chromatography/Mass Spectrometry

Seeds of nutmeg are used as spice, but they are also abused because of psychotropic effects described after ingestion of large doses. It was postulated that these effects could be attributable to metabolic formation of amphetamine derivatives from the main nutmeg ingredients elemicin (EL), myristicin (MY), and safrole (SA). In a case of a suspected nutmeg abuse, neither such amphetamine derivatives nor the main nutmeg ingredients could be detected in urine. The metabolites of EL, MY, and SA were identified using gas chromatography-mass spectrometry in rat urine and their presence in human urine of the nutmeg abuser was confirmed. The identified metabolites indicated that EL, MY, and SA were once and twice hydroxylated at the side chain. In addition, EL was O-demethylated at 2 positions followed by side chain hydroxylation. MY and SA were demethylenated and subsequently methylated. In the human urine sample, the following metabolites could be identified: O-demethyl elemicin, O-demethyl dihydroxy elemicin, demethylenyl myristicin, dihydroxy myristicin, and demethylenyl safrole. As in the human urine sample, neither amphetamine derivatives nor the main nutmeg ingredients could be detected in the rat urine samples. Finally, toxicologic detection of nutmeg abuse was possible by identification of the described metabolites of the EL, MY, and SA in urine applying the authors' systematic toxicologic analysis procedure using full-scan gas chromatography-mass spectrometry after acid hydrolysis, liquid-liquid extraction of analytes, and microwave-assisted acetylation of extracted analytes.

The effects of arecoline on the release of cytokines using cultured peripheral blood mononuclear cells from patients with oral mucous diseases.

In Taiwan there is a significant correlation between oral precancer diseases and oral cancer associated with the betel quid chewing habit. The carcinogenic components of betel quid are arecoline, arecaidine and safrole. However, it is unknown whether these substances influence the immune functions. This study investigated the effects of betel quid on the immune system using cultured peripheral blood mononuclear cells from patients with oral mucous diseases. In our experiment, mononuclear cells from 10 normal persons, 12 patients with precancer lesions, and 16 patients with squamous cell carcinoma were separated from blood samples and cultured. After stimulation by arecoline, the amounts of IL-2, TNF-alpha, TGF-beta and IFN-gamma secreted by mononuclear cells were measured using the ELISA method. The results showed that IL-2, TNF-alpha, and TGF-beta were significantly lower in mononuclear cells of normal persons as stimulated by arecoline. The TGF-beta amount in cells from oral submucous fibrosis patients with betel quid chewing habit (OSF-B) was lower than normal persons or patients who had long term betel quid chewing habit but were without oral mucosal diseases (N-B), and was also lower than the squamous cell carcinoma with betel quid chewing group (SCC-B). TNF-alpha was significantly lower in the squamous cell carcinoma with long term betel quid chewing group (SCC-B) than in normal persons. TNF-alpha was significantly higher in the squamous cell carcinoma without betel quid chewing group (SCC-N) than in normal persons and SCC-B groups. In addition, IFN-gamma was significantly lower in patients who had long term betel quid chewing but were without oral mucous lesions than the normal person and the OSF group. The results proved that betel quid influences cytokines production by mononuclear cells.

Cytogenetic effects of piperonyl butoxide and safrole in CHO-K1 cells

Recently, hepatocarcinogenicity in rats and mice was reported with regard to the methylenedioxyphenyl compound, piperonyl butoxide (PB), which is used as a synergist for pyrethrins and related insecticides. Induction of sister-chromatid exchanges (SCEs) and chromosomal aberrations (CAs) due to PB were investigated using CHO-K1 cells with or without rat liver S9 fraction (S9); at the same time, the effects of safrole (SF), a methylenedioxyphenyl compound and a weak hepatocarcinogen, were also examined. PB (0.25 and 0.3 mM) and SF (0.8 mM) caused a slight but significant increase in SCEs followed by a cell-cycle delay in the 3-h treatment without S9. In the presence of S9 (4.5%), the cytotoxicity of PB or SF was weakened greatly or slightly, the top dose capable of cell division was raised to 0.6 mM (2-fold) or 1 mM, respectively. PB with S9 induced SCE at doses of 0.4 and 0.5 mM, and caused endoreduplications (ERDs, 7%) at a dose of 0.6 mM, while SF caused a dose-related significant increase in SCE at all doses used (0.4–1 mM) with S9. Genotoxicity of the metabolites of PB or SF was cleared by changing the dose of S9 (1.5–9%) while holding the dose of each chemical constant. In the case of SF (0.6 mM), induction of SCE, ERD and cell-cycle delay intensified almost in a dose-effect relationship, and CAs and a high level of ERD (14%) were caused by a 9% dose of S9. The concentration of unchanged SF in the incubated medium was certainly in inverse proportion to the dose of S9. This strongly suggests that the metabolites of SF are genotoxic. In the case of PB (0.3 mM), no positive responses were produced in the cultures, even with a high level of S9, though the amount of unchanged PB left in the incubated medium was very slight. This indicates that the metabolites of PB may not be genotoxic. In conclusion, PB and SF are possible to somewhat induce SCE at high dose(s) in the absence of S9, and the genotoxic effects of SF are more intensified in the presence of S9 than in its absence, while PB is probably no genotoxic in the presence of sufficient metabolic activation.

SAF's Role and Strategic Priorities

SAF's Role and Strategic Priorities

Differences in Induction of Three P450 Isozymes by Piperonyl Butoxide, Sesamex, Safrole, and Isosafrole

The induction of the P450 isozymes 1a-1, 1a-2, and 2b-10 by four methylenedioxyphenyl (MDP) compounds, safrole (SAF), isosafrole (ISO), piperonyl butoxide (PBO), and sesamex (SES), and by the nonmethylenedioxyphenyl analog of SAF, allyl benzene (AB), was studied using male C57BL/6N mice. P450 1a-1 was not detected in control animals, and was induced by SES and PBO, with SES inducing more 1a-1 protein than PBO. P450 1a-2 mRNA was detected in the livers of control animals and was also increased by all MDP compounds (SES > PBO ≅ ISO > SAF), although SAF treatment did not increase 1a-2 protein. P450 2b-10 mRNA and protein, not detected in untreated animals, were also increased by all MDP compounds (PBO > SES ≅ ISO > SAF). AB treatment did not induce detectable levels of 1a-1, 1a-2, or 2b-10, suggesting that, in this series of compounds, the methylenedioxyphenyl moiety is important in induction.

Metabolism of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone by inducible and constitutive cytochrome P450 enzymes in rats

The tobacco-specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) induces tumor formation in the liver, lung, nasal cavity, and pancreas of rats. Metabolic activation is required for the tumorigenicity of this compound. The involvement of cytochrome P450 enzymes in NNK bioactivation was investigated in rats by studies with chemical inducers and antibodies against P450s. Liver microsomal enzymes catalyzed the formation of 4-oxo-1-(3-pyridyl)-1-butanone (keto aldehyde), 4-hydroxy-1-(3-pyridyl)-1-butanone (keto alcohol), 4-(methylnitrosamino)-1-(3-pyridyl- N-oxide)-1-butanone (NNK- N-oxide), and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) from NNK. When the activity was expressed on a per nanomole P450 basis, treatments of rats with 3-methylcholanthrene (MC), phenobarbital (PB), pregnenolone 16-α-carbonitrile (PCN), Aroclor 1254 (AR), safrole (SA), and isosafrole (ISA) increased the keto aldehyde formation in liver microsomes 2.0-, 2.4-, 3.8-, 2.5-, 2.1-, and 1.8-fold, respectively; PB, AR, SA, and ISA increased the keto alcohol formation 1.7-, 1.3-, 2.0-, and 1.3-fold, respectively. The extents of induction were more pronounced when expressed on a per milligram protein basis, due to the higher microsomal P450 contents in the induced microsomes. The formation of NNK- N-oxide was markedly increased by PB and PCN and slightly increased by AR, SA, and ISA. However, the formation of NNAL, the major metabolite due to carbonyl reduction, was not increased by the treatments but was decreased by AR, ISA, and acetone (AC). The kinetic parameters of NNK metabolism by control, MC-, PB-, and PCN-induced liver microsomes were obtained. A panel of monoclonal (anti-1A1, -2B1, -2C11, and -2E1) and polyclonal (anti-1A2, -2A1, and -3A) antibodies were used to assess the involvement of constitutive hepatic P450 enzymes in NNK metabolism. Keto aldehyde formation was inhibited by anti-1A2 and anti-3A (about 15%) but not by others; the formation of keto alcohol was inhibited by anti-1A2, anti-2A1, and anti-3A (by 13–26%). In incubations with lung microsomes, the formation of keto aldehyde, keto alcohol, NNK- N-oxide, and NNAL were observed. With nasal mucosa microsomes, however, only keto aldehyde and keto alcohol formation were appreciable. SA and AC significantly decreased NNK metabolism in lung and nasal mucosa microsomes. Furthermore, SA inhibited the NNK oxidative metabolism in lung and nasal mucosa microsomes in vitro. AC significantly inhibited the NNK oxidative metabolism in lung microsomes in vitro, but not in nasal mucosa microsomes. The present results demonstrate the important role of P450 enzymes in the metabolism of NNK. Several P450 isozymes can catalyze the oxidation of NNK with different but overlapping regioselectivities and P450 inducers affect the metabolism of NNK differently in the liver, lung, and nasal mucosa.

Safrole as Starting Material for the Synthesis of 7-Metoxy-3',4'-Methylenedioxy Isoflavanone

Safrole as Starting Material for the Synthesis of 7-Metoxy-3',4'-Methylenedioxy Isoflavanone

Continuing Forestry Education--The SAF Role

Continuing Forestry Education--The SAF Role

Mad about safrole

Mad about safrole

Questions of safrole metabolism and carcinogenicity

Questions of safrole metabolism and carcinogenicity

25. A further report on safrole

25. A further report on safrole

Detection of Safrole in Foods and Drugs

Journal Article Detection of Safrole in Foods and Drugs Get access Christiane Genest, Christiane Genest Food and Drug Laboratories, Department of National Health and Welfare, Ottawa, Canada Search for other works by this author on: Oxford Academic Google Scholar D Morison Smith, D Morison Smith Food and Drug Laboratories, Department of National Health and Welfare, Ottawa, Canada Search for other works by this author on: Oxford Academic Google Scholar D G Chapman D G Chapman Food and Drug Laboratories, Department of National Health and Welfare, Ottawa, Canada Search for other works by this author on: Oxford Academic Google Scholar Journal of Association of Official Agricultural Chemists, Volume 44, Issue 4, 1 November 1961, Pages 631–632, https://doi.org/10.1093/jaoac/44.4.631 Published: 01 February 2020

Studies on the Related Compounds of Safrole. I. On the Will gerodt Reaction on the Related Compounds of Isosafrole and Piperonylmethylketone.

Studies on the Related Compounds of Safrole. I. On the Will gerodt Reaction on the Related Compounds of Isosafrole and Piperonylmethylketone.