Radical generation, radical-scavenging activity, and cytotoxicity of eugenol-related compounds.
To clarify the possible link between radicals and cytotoxicity of eugenol-related compounds, dimeric compounds were synthesized from eugenol (4-allyl-2-methoxy-phenol), butylated hydroxyanisole (BHA) (2-t-butyl-4-methoxyphenol) or MMP (2 methoxy-4-methylphenol); bis-EUG (3,3'-dimethoxy-5,5'-di-2-propenyl-1,1'-biphenyl-2,2'-diol), bis-BHA (3,3'-di-t-butyl-5,5'-dimethoxy-1,1'-biphenyl-2,2'-diol), and bis-MMP (3,3'-di-methoxy-5,5'-dimethyl-1,1'-biphenyl-2,2'-diol). The cytotoxic activity of these compounds was determined using a salivary gland tumor cell line (HSG), oral squamous cell carcinoma cell line (HSC-2) and human promyelocytic leukemia cell line (HL-60). A parabolic relationship between the cytotoxicity and log P (the octanol-water partition coefficient) was observed, showing that both BHA and bis-MMP, with a log P of 3-4, were the most cytotoxic. The cytotoxic activity of the 2-methoxy derivatives, eugenol, MMP and bis-MMP, against HSG cells was significantly enhanced by visible-light irradiation, possibly due to their high redox potential. Electron spin resonance (ESR) spectroscopy indicated that eugenol and BHA alone produced radicals under alkaline conditions (pH > 9.5), and eugenol most efficiently scavenges reactive oxygen species (O2-). Antioxidative reactivity of eugenol-related compounds was determined by measuring the inhibiting periods of the AIBN (2,2'-azobisisobutyronitrile)/MMA (methyl methacrylate) polymerization system, and the number of moles of peroxy radical trapped by moles of the relevant phenols (stoichiometric factor, n). It was found that the n values of eugenol and MMP were approximately 1, whereas those of BHA >2, suggesting that eugenol and MMP undergo dimerization through radical-radical couplings through quinone methides, whereas BHA undergoes the competitive interaction with poly-MMA radicals after oxidation by AIBN-peroxy radicals. BHA was an efficient peroxy radical-scavenger, but possibly reacted with polymer radicals of the lipid, thus mediating the cytotoxicity. The n value of bis-BHA was two, whereas those of bis-EUG and bis-MMP were 1.6-1.7, suggesting that the latter were further oxidized. The enthalpies of phenoxyl radical formation were determined using the semi-empirical PM3 quantum-mechanical method and the possible link to redox potential was discussed.
Fujisawa
S
S
Department of Oral Diagnosis, Meikai University School of Dentistry, Saitama, Japan. fujisawa@dent.meikai.ac.jp
Atsumi
T
T
Satoh
K
K
Kadoma
Y
Y
Ishihara
M
M
Okada
N
N
Nagasaki
M
M
Yokoe
I
I
Sakagami
H
H
D000230:Adenocarcinoma; D000975:Antioxidants / Q000378:metabolism / Q000633:toxicity; D005054:Eugenol / Q000031:analogs & derivatives / Q000378:metabolism* / Q000633:toxicity*; D016166:Free Radical Scavengers / Q000378:metabolism*; D005609:Free Radicals / Q000378:metabolism; D018922:HL-60 Cells; D006801:Humans; D009062:Mouth Neoplasms; D017382:Reactive Oxygen Species / Q000378:metabolism; D012468:Salivary Gland Neoplasms; D012709:Serum Albumin / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured
D000975:Antioxidants; D016166:Free Radical Scavengers; D005609:Free Radicals; D017382:Reactive Oxygen Species; D012709:Serum Albumin; D013481:Superoxides; D005054:Eugenol