Eugenol과 1α,25-dihydroxyvitamin D₃의 병합처리에 의한 HL-60 세포의 분화 유도

오미경,박선주,김남훈,조진경,진종률,김인숙,Oh, Mi-Kyung,Park, Seon-Joo,Kim, Nam-Hoon,Cho, Jin-Kyung,Jin, Jong-Youl,Kim, In-Sook Korean Society of Life Science 2007 생명과학회지 Vol.17 No.9

Eugenol은 여러가지 향신료에 있는 필수 오일의 주요 성분으로서 악성 종양 세포의 성장을 저해하고 사멸을 유도한다고 보고되었다. 본 연구에서는 eugenol이 세포 분화 유도에 관여하는지를 조사하기 위하여 HL-60 전골 수성 백혈병 세포의 분화에 미치는 eugenol의 효과를 연구하였다. HL-60 세포에 eugenol (150 ${\mu}M$)을 가했을 때 세포성장이 저해되었으며 $1,25(OH)_{2}$ vit $D_{3}$ (3 nM)와 병합처리시에는 더 큰 저해효과를 보였다. 이 때, eugenol은 $1,25(OH)_{2}$ vit $D_{3}$에 의해 유도되는 세포주기의 $G_{0}/G_{1}$단계 정지를 더욱 증가시킴을 알 수 있었다. 또한, eugenol과 $1,25(OH)_{2}$ vit $D_{3}$를 병합처리 하였을 때에는 $G_{0}/G_{1}$단계의 정지와 관련된 세포주기 조절인자인 p27 level를 상호 협동적으로 증가시켰을 뿐만 아니라 cyclin A, cdk2, cdk4 level를 감소시켰다. 또한 유세포분석실험을 통하여, CD14 (단핵세포 표지 인자)의 발현이 eugenol과 $1,25(OH)_{2}$ vit $D_{3}$를 병합처리한 세포에서 단독처리시보다 더 증가함을 알 수 있었다. 이러한 결과들은 eugenol이 $1,25(OH)_{2}$ vit $D_{3}$와 상호협동적으로 작용하여 저농도의 $1,25(OH)_{2}$ vit $D_{3}$에 의해 자극된 세포 분화 신호를 더욱 더 증대 시킴을 나타낸다. 이러한 eugenol에 의한 세포 분화 유도 작용은 암의 화학적예방 효과에 유용하게 응용될 수 있을 것으로 기대된다. Eugenol (4-allyl-2-methoxyphenol) is a main component of essential oils obtained from various spices. Recent reports have shown that eugenol induces growth inhibition and apoptosis of malignant tumor cells. In this study, the stimulatory effect of eugenol on cell differentiation was investigated in HL-60 promyelocytic leukemia cells. When HL-60 cells were treated in combination with 150 ${\mu}M$ of eugenol and 3 nM of $1{\alpha},25-dihydroxyvitamin$ $D_{3}$, cell growth was slower than that of cells treated with eugenol or $1{\alpha},25-dihydroxyvitamin$ $D_{3}$ alone. Eugenol enhanced low dose of $1{\alpha,25-dihydroxyvitamin }$ $D_{3}-induced$ a $G_{0}/G_{1}$ phase arrest in cell cycle. Consistent with this, combined treatment of eugenol and $1{\alpha},25-dihydroxyvitamin$ $D_{3}$ cooperatively increased p27 level and decreased cyclin A, cdk 2 and cdk 4 levels, which are cell cycle regulators related to $G_{0}/G_{1}$ arrest. According to flow cytometric analysis, the expression of CD14 (monocytic differentiation marker) was more increased in the cells co-treated with eugenol and $1{\alpha},25-dihydroxyvitamin$ $D_{3}$. These results indicate that eugenol potentiates cell differentiation mediated by $1{\alpha},25-dihydroxyvitamin$ $D_{3}$ of suboptimal concentration. The differentiation-inducing property of eugenol maybe contributes to chemopreventive activity of cancer.

사람흑색종세포에서 eugenol에 의한 caspase 의존적 세포자멸사 반응

김규천,최돈수,임장섭,정희찬,,김인령,이무형,박봉수 대한해부학회 2006 Anatomy & Cell Biology Vol.39 No.3

Malignant melanoma is a highly metastatic tumor, resistant to chemotherapy and radiotherapy. Recent studies have suggested that many therapeutic agents used against cancer mediate their effects by induction of apoptosis of the cancer cells. Eugenol enhances the generation of tissue-damaging free radicals and inflammation or allergic reactions. In particular, it is more cytotoxic against cancer cells compared with normal fibroblasts. This study was performed to investigate whether the cytotoxic effect of eugenol is associated with the induction of apoptosis and involves activation of caspase in the human melanoma G361 cells. Eugenol-induced apoptosis was confirmed by MTT assay, Hemacolor stain, Hoechst stain, DNA electrophoresis, and Western blot analysis. Eugenol had a significant dose- and time-dependent inhibitory effect on the viability of G361 cells. Eugenol treatment induced caspase-3 and -6 cleavage, and activation. The caspase-3 substrates PARP and DFF45 are cleaved during eugenol-induced apoptosis. It was found that the casapase-6 substrate lamin A was cleaved, whose cleavage has been reported to be necessary for complete condesation of DNA during apoptosis. These results suggest that eugenol may constitute a potential antitumor compound against melanoma occurring in the skin and oral mucosa. 악성 흑색종은 높은 전이성을 가지며, 화학요법과 방사선요법에 저항성을 보이는 암이다. 최근의 연구 들은 암에 이용되는 많은 치료약물들이 암세포에서 세포자멸사를 유도함으로써 그 효과를 가지는 것임을 제안 하고 있다. Eugenol은 조직에 상처를 주는 free radical의 생성과 염증 또는 알러지 반응을 증가시킨다. 특별히 정상 섬유모세포에 비해 암세포에 더욱 큰 세포독성을 가지고 있는 것으로 알려졌다. 그래서 본 연구는 사람흑 색종세포에서 eugenol이 세포자멸사와 연관된 세포 독성효과가 있는지와 그 과정에서 caspase의 활성이 수반되 는지를 알아보고자 시행되었다. 본 연구는 eugenol이 G361 세포에서 세포자멸사를 일으킴을 확인했으며, eugenol이 유도한 세포자멸사는 MTT 분석법, Hemacolor 염색, Hoechst 염색, DNA electrophoresis 그리고 Western blot 분석법으로 증명되었다. Eugenol은 농도의존적 그리고 시간의존적으로 G361 세포의 생존율을 현격하게 감소시켰으며, eugenol 처리는 caspase-3와 -6의 분절과 활성을 유도했다. Caspase-3의 기질인 PARP와 ICAD/DFF45도 또한 eugenol이 유도한 세포자멸사 과정 중에 분절되었다. 그리고 또한 casapase-6의 기질인 lamin A의 분절이 관찰되었다. Lamin A의 분절된 형태는 DNA의 응축에 필수적인 것으로 알려져 있다. 본 연구결과는 eugenol이 피부나 구강점막에서 발 생하는 사람흑색종에 대해서 잠재력을 가진 항암물질로서 기여할 수 있음을 제시하고 있다.

정향(Eugenia caryophyllata Thunb.) Eugenol 및 그 유도체의 항산화 및 항염증활성

임현희(Hyun-Hee Leem),김은옥(Eun-Ok Kim),서미자(Mi-Jae Seo),최상원(Sang-Won Choi) 한국식품영양과학회 2011 한국식품영양과학회지 Vol.40 No.10

본 연구는 한방스킨의 원료로 널리 사용되고 있는 8가지 생약의 휘발성증류추출액 중 항산화 및 항염증활성이 가장 강한 정향의 증류추출액으로부터 주된 향기성분을 SDE법으로 추출한 후 GC-MS로 확인하였으며, 주된 향기성분인 eugenol과 그 유도체를 합성한 후 항산화 및 항염증활성을 측정하고 비교하였으며, 아울러 HPLC를 이용하여 정향의 eugenol 및 그 유도체를 정량분석 하였다. 8가지 생약의 휘발성증류추출액 중 정향의 증류추출액이 가장 강한 항산화활성(IC₅₀=8.85 μg/mL) 및 COX-2 저해활성(10 μg/mL 농도에서 저해율은 58.15%)을 나타내었으며, 반면 15-LOX 저해활성(25 μg/mL 농도에서 저해율은 86.15%)은 당귀 다음으로 가장 높았다. 정향 증류추출액의 휘발성 향기성분을 SDE법으로 추출한 후 GC-MS를 이용하여 분석한 결과, eugenol, trans-caryophyllene 및 acetyl eugenol을 확인하였다. 한편, eugenol 및 그 유도체(methyl eugenol 및 acetyl eugenol)의 항산화 및 항염증 활성을 측정한 결과, eugenol(IC₅₀=5.99 μg/mL)이 가장 높은 항산화활성을 나타낸 반면, methyl eugenol 및 acetyl eugenol은 거의 활성을 나타내지 않았다. COX-2의 경우 20 μg/mL 농도에서 eugenol(85.35%)이 가장 강한 저해활성을 나타낸 반면, 15-LOX는 20 μg/mL 농도에서 methyl eugenol(83.29%)이 가장 높은 저해활성을 나타내었다. 정향 에탄올추출물의 eugenol 및 유도체의 함량을 HPLC로 분석한 결과, eugenol 및 acetyl eugenol이 각각 6.95%, 1.85% 함유되어 있었으며 methyl eugenol은 검출되지 않았다. 이와 같이 정향 유래의 eugenol 및 그 유도체는 안전성이 문제시되고 있는 합성항산화제 및 항염증제를 대체할 수 있는 천연 유래의 항산화 및 항염증물질로서 잠재적 가치가 있어 향후 기능성식품, 화장품 및 의약품 소재로 널리 사용될 수 있을 것으로 생각된다. Antioxidant and anti-inflammatory activities of eugenol and its derivatives from clove (Eugenia caryophyllata Thunb.) were evaluated using in vitro assay systems by measuring 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, cyclooxygenase-2 (COX-2), and 15-lipoxygenase (15-LOX). Among eight different crude medicinal drugs tested, volatile extracts of clove extracted by steam distillation extraction (SDE) showed potent DPPH radical scavenging activity (IC₅₀=8.85 μg/mL) as well as strong inhibitory activity against COX-2 (58.15%) and 15-LOX (86.15%) at 10 μg/mL and 25 μg/mL, respectively. Major volatile components of clove were identified as eugenol, trans-caryophyllene, and acetyleugenol by GC-MS analysis. Out of three eugenol derivatives, eugenol, methyl eugenol, and acetyl eugenol, eugenol showed the strongest DPPH radical scavenging activity and COX-2 in-hibitory activity, whereas methyl eugenol exhibited the strongest 15-LOX inhibitory activity. Finally, the contents of the three eugenol derivatives in clove were quantified by analytical HPLC. Contents of eugenol and acetyl eugenol in clove were 6.95% and 1.85% per dry weight, respectively. These results suggest that eugenol and its derivatives in steam distilled extract of clove may be useful as potential antioxidant and anti-inflammatory agents.

EFFECT OF EUGENOL AND CAPSAICIN ON THE VOLTAGE-DEPENDENT ION CHANNELS OF TRIGEMINAL AFFERENTS

김주연,박상진,최기운,최호영 대한치과보존학회 2000 Restorative Dentistry & Endodontics Vol.25 No.3

삼차신경절의 뉴런이 구강악안면영역에서의 촉각, 압각,온도각 및 통각 등 다양한 감각을 중추신경계로 전달하는 역할을 하는 것은 주지의 사실이다. 이러한 신경전달에 있어서 이온통로는 감각정보를 전달하는데 핵심적인 역할을 수행하며 특히 소디움 통로는 활동전위의 발생에 중요하다. 소디움 통로는 tetrodotoxin-sensitive(TTX-5) 및 tetrodotox-in-resistant(TTX-r) 통로로 나누어지는 데 이 중 TTX-r 통로에 발생되는 tetrodotoxin-resistant sodium cur-rent(TTX-r INa.)는 capsaicin에 민감한 일차구심신경세포에서 유해자극에 의해 통각신호를 발생시키고 전달하는데 중요하다. 또한 칼슘 통로는 시냅스 전토에 있어서 필수적인 역할을 수행하고 있다. 한편 치과영역에서 치수의 진정 목적으로 eugenol이 흔히 사용되고 있다. 그러나eugenol의 그 작용 기전에 대해서 현재까지 이온통로에 대한 상세한 결과가 없는 실정이며 최근의 보고에 의하면 eugenol이 capsaicin수용기를 통하여 감각신경에 대한 억제작용을 나타낸다고 한다. 따라서 본 실험은 eugenol과 capsaicin이 흰쥐의 삼차신경절의 TTX-r INa와 칼슘통로에 어떠한 영향을 미치는지를 알아보고 eugenol이 capsaicin 수용기를 통하여 작용하는지를 검증하고자 시행되었다 삼차신경절 뉴런은 100-150g의 흰쥐의 삼차신경절로부터 외과적으로 절제하여 통법의 화학적 및 기계적 처리를 통해 단일세포로 분리하였고 이를 Whole-cell patch clamp 방법을 이용하여 시행한 바 다음과 같은 결론을 얻었다. 1.1mM의 dugenol은 흰쥐 삼차신경절 뉴런의 TTX-r INa.와 HVA INa.를 억제하였다. 2.1㎛의 capsaicin은 흰쥐 삼차신경절 뉴런의 TTX-r INa 와 HVA INa 를 억제하였다 3.Capsazepine은 capsaicin의 HVA ICa 에 대한 억제작용을 차단하였다. 4.Capsazepine은 capsaicin의 HVA ICa .에 대한 억제작용을 차단하지 못하였다. 결론적 으로 eugenol과 capsaicin은 tetrodotoxin-resistant sodium current(TTX-r INa )와 high voltage-activated calcium current(HVA ICa )를 모두 억제하는 것으로 나타났으며 , 이러한 작용이 통각의 발생과 시냅스 전달과정을 차단하여 치수 진정 목적으로 많이 사용하는 eugenol의 작용기전으로 판단된다. 한편 capsaicin의 길항제인 capsazepine을 전처치하였을 때에도 eugenol의 HVA ICa 에 대한 억제효과는 변화가 없었다. 이와같은 결과로 보아 HVA ICa 에 관한한 eugenol은 capsaicin 수용기를 통하여 나타나지 않는 것으로 사료된다.

Eugenol Inhibits K⁺ Currents in Trigeminal Ganglion Neurons

Li, H. Y.,Park, C.-K.,Jung, S. J.,Choi, S.-Y.,Lee, S. J.,Park, K.,Kim, J. S.,Oh, S. B. SAGE Publications 2007 Journal of dental research Vol.86 No.9

<P>Eugenol, a natural capsaicin congener, is widely used in dentistry. Eugenol inhibits voltage-activated Na<SUP>+</SUP> and Ca<SUP>2+</SUP> channels in a transient receptor potential vanilloid 1 (TRPV1)-independent manner. We hypothesized that eugenol also inhibits voltage-gated K<SUP>+</SUP> currents, and investigated this in rat trigeminal ganglion neurons and in a heterologous system using whole-cell patch clamping. Eugenol inhibited voltage-gated K<SUP>+</SUP> currents, and the inhibitory effects of eugenol were observed in both capsaicin-sensitive and capsaicin-insensitive neurons. Pre-treatment with capsazepine, a well-known antagonist of TRPV1, failed to block the inhibitory effects of eugenol on K<SUP>+</SUP> currents, suggesting no involvement of TRPV1. Eugenol inhibited human Kv1.5 currents stably expressed in <I>Ltk</I><SUP>−</SUP> cells, where TRPV1 is not endogenously expressed. We conclude that eugenol inhibits voltage-gated K<SUP>+</SUP> currents in a TRPV1-independent manner. The inhibition of voltage-gated K<SUP>+</SUP> currents is likely to contribute to the irritable action of eugenol. Abbreviations: human Kv1.5 channel, hKv1.5; transient receptor potential vanilloid 1, TRPV1.</P>

토마토 'Micro-Tom' 과실의 eugenol synthase 유전자 클로닝, 단백질의 3차 구조 및 생리화학적 특성 예측

강승원,서상규,이태호,이긍표 경상대학교 농업생명과학연구원 2012 농업생명과학연구 Vol.46 No.4

Eugenol은 많은 식물에서 eugenol synthase에 의해 생합성되는 phenylpropene 계통의 휘발성 화합물이다. 그러나, 토마토 과실에서의 특징은 밝혀져 있지 않다. 이에 따라 토마토 ‘Micro-Tom'으로부터 RACE 기법을 이용하여 완전장 cDNA를 클로닝 하여, SlEGS라 명명하였다. SlEGS의 open reading frame은 921bp로, 307개의 아미노산 서열을 갖는 단백질로 번역되었다. BLAST 결과에 따라 SlEGS는 PhEGS1 및 CbEGS2와 각 67.1, 69.4%의 높은 상동성을 갖는 것으로 나타났다. CLC genomics workbench 프로그램을 이용하여 SlEGS의 아미노산 구성을 분석하였고, Swiss-PDB viewer 프로그램에서 homology modeling 기법으로 SlEGS의 3차원 단백질 구조를 구축한 후 ProSA-web 툴로 3차원 구조의 안정성을 확인 하였다. 또한 ExPASy의 ProtParam 툴을 이용하여 SlEGS의 생리화학적 특성을 분석 하였다. SlEGS의 추정 분자량은 33.93kDA이고 등전점(pI)은 5.85로 산성인 것으로 나타났다. 이와 더불어 SlEGS의 흡광 계수(EC), 불안정성 지수(II), alipathic 지수(AI), GRAVY값 등의 생리화학적 특성에 대한 분석을 실시 하였다. Eugenol is a volatile compound synthesized by eugenol synthase in various plants and belongs to phenylpropene compounds. However, characteristics of eugenol synthase in tomato has not been known. Therefore, we cloned a full length cDNA of a putative eugenol synthase from tomato 'Micro-Tom' using rapid amplification of cDNA ends (RACE) technique and named a clone SlEGS. Open reading frame of SlEGS was 921bp long and its deduced amino acid sequence was 307bp. The BLAST analysis indicated that SlEGS shared high similarity with PhEGS1 (67.1%) and CbEGS2 (69.4%). Amino acid composition of SlEGS was determined by CLC genomics workbench tool and 3D structure of SlEGS was constructed by homology modeling using Swiss-PDB viewer and validated using PROCHECK and ProSA-web tool. In addition, the physiochemical properties of SlEGS was evaluated using ExPASy's ProtParam tool. Molecular weight was 33.93kDa and isoelectric point was 5.85 showing acidic nature. Other properties such as extinction coefficient, instability index, aliphatic index, and grand average hydropathy was also analyzed.

백서 척수에서 Capsaicin과 Eugenol이 iCGRP (immunoreactive calcitonin gene-related peptide) 분비 조절에 미치는 영향

오원만,김원재,최남기,박상원,황인남,김선현 大韓齒科保存學會 2001 Restorative Dentistry & Endodontics Vol.26 No.5

Neuropeptide such as calcitonin gene-related peptide and substance P may mediate neurogenic inflammation, but little is known about the regulation of neuropeptide release from rat spinal cord. Eugenol has been reported to reduce odontogenic pain and is known to have a structure similar to capsaicin, a potent stimulant of certain nociceptors. This study was done to examine the effect of capsaicin and eugenol on immunoreactive calcitonin gene-related peptide (iCGRP) release from rat spinal cord and whether eugenol regulates capsaicin-sensitive release of iCGRP or it evokes capsaicin-sensitive release of iCGRP. The dorsal half of rat lumbar spinal cord was chopped into 200㎛ slices. They were superfused (500㎕/min) in vitro with an oxygenated Kreb's buffer. The EC50 of capsaicin on iCGRP release was measured. Eugenol (600㎛ and 1.2mM) and vehicle (0.02% 2-hydroxyl-β-cyclodextrin) were administered prior to stimulation of rat lumbar spinal cord with capsaicin. The amount of iCGRP release from rat lumbar spinal cord was measured by radioimmunoassay. The results were as follows : 1.iCGRP release from rat lumbar spinal cord was dependent on concentration of capsaicin. The EC50 of capsaicin on iCGRP release was 3㎛. 2.In the vehicle treated group, capsaicin (3㎛) evoked a 14-fold increase over basal iCGRP level. 3. Administration of 600㎛ and 1.2mM eugenol evoked a 2.2-fold increase and a 2.3-fold increase over basal iCGRP level respectively. 4.Administration of 600㎛ and 1.2mM eugenol increased capsaicin evoked release of iCGRP by more than 50%. These results indicate that eugenol evoke CGRP release from central nervous system and potentiate the pain-inducing action of capsaicin on it Key words : Eugenol, Capsaicin, Rat lumbar spinal cord, Superfusion, Radioimmunoassay

S Phase Cell Cycle Arrest and Apoptosis is Induced by Eugenol in G361 Human Melanoma Cells

김규천 KOREAN ACADAMY OF ORAL BIOLOGY 2011 International Journal of Oral Biology Vol.36 No.3

Eugenol is an essential oil found in cloves and cinnamon that is used widely in perfumes. However, the significant anesthetic and sedative effects of this compound have led to its use also in dental procedures. Recently, it was reported that eugenol induces apoptosis in several cancer cell types but the mechanism underlying this effect has remained unknown. In our current study, we examined whether the cytotoxic effects of eugenol upon human melanoma G361 cells are associated with cell cycle arrest and apoptosis using a range of methods including an XTT assay, Hoechst staining, immunocytochemistry, western blotting and flow cytometry. Eugenol treatment was found to decrease the viability of the G361 cells in both a time- and dose-dependent manner. The induction of apoptosis in eugenol-treated G361 cells was confirmed by the appearance of nuclear condensation, the release of both cytochrome c and AIF into the cytosol, the cleavage of PARP and DFF45, and the downregulation of procaspase-3 and -9. With regard to cell cycle arrest, a time-dependent decrease in cyclin A, cyclin D3, cyclin E, cdk2, cdk4, and cdc2 expression was observed in the cells after eugenol treatment. Flow cytometry using a FACScan further demonstrated that eugenol induces a cell cycle arrest at S phase. Our results thus suggest that the inhibition of G361 cell proliferation by eugenol is the result of an apoptotic response and an S phase arrest that is linked to the decreased expression of key cell cycle-related molecules.

Mechanisms Underlying the Protective Effect of Eugenol in Rats with Acute Doxorubicin Cardiotoxicity

Amr A. Fouad,Mohamed T. Yacoubi 대한약학회 2011 Archives of Pharmacal Research Vol.34 No.5

The protective effect of eugenol and its possible mechanisms were investigated in rats with acute doxorubicin cardiotoxicity. Cardiac toxicity was induced by a single intraperitoneal injection of doxorubicin (20 mg/kg). Eugenol treatment (5 mg/kg/day, orally) was started 2 days before doxorubicin administration and continued for five consecutive days. Eugenol significantly reduced the elevated serum creatine kinase and lactate dehydrogenase levels, and restored the electrocardiographic disturbances resulted from doxorubicin administration. Also, eugenol reversed doxorubicin-induced deficits in the antioxidant defense mechanisms, decreased lipid peroxidation and attenuated the elevations in cytosolic Ca^2+ and nitric oxide levels in cardiac tissue. In addition, doxorubicin-induced cardiac tissue damage observed by histopathological examination was markedly ameliorated with eugenol. Immunohistochemical analysis revealed that eugenol prevented the doxorubicin-induced activation of caspase-3 in cardiomyocytes. The cardioprotective effect afforded by eugenol was not significantly inhibited by prior administration of capsazepine, the transient potential vanilloid receptor-1 antagonist. It was concluded that eugenol, through its antioxidant activity and its ability to reduce cardiac Ca^2+ accumulation and nitric oxide levels, is a potential candidate to protect against acute doxorubicin cardiotoxicity, a major and dose-limiting clinical problem.

Eugenol Inhibits Sodium Currents in Dental Afferent Neurons

Park, C.-K.,Li, H. Y.,Yeon, K.-Y.,Jung, S. J.,Choi, S.-Y.,Lee, S. J.,Lee, S.,Park, K.,Kim, J. S.,Oh, S. B. Journal of Dental Research, Inc 2006 Journal of dental research Vol.85 No.10

<P>Although eugenol is widely used in dentistry, little is known about the molecular mechanisms responsible for its anesthetic properties. In addition to calcium channels, recently demonstrated by our group, there could be another molecular target for eugenol. Using a whole-cell patch-clamp technique, we investigated the effect of eugenol on voltage-gated sodium channel currents (<I>I</I><SUB><I>Na</I></SUB>) in rat dental primary afferent neurons identified by retrograde labeling with a fluorescent dye in maxillary molars. Eugenol inhibited action potentials and <I>I</I><SUB><I>Na</I></SUB> in both capsaicin-sensitive and capsaicin-insensitive neurons. The pre-treatment with capsazepine, a competitive antagonist of transient receptor potential vanilloid 1 (TRPV1), failed to block the inhibitory effect of eugenol on <I>I</I><SUB><I>Na</I></SUB>, suggesting no involvement of TRPV1. Two types of <I>I</I><SUB><I>Na</I></SUB>, tetrodotoxin (TTX)-resistant and TTX-sensitive <I>I</I><SUB><I>Na</I></SUB>, were inhibited by eugenol. Our results demonstrated that eugenol inhibits <I>I</I><SUB><I>Na</I></SUB> in a TRPV1-independent manner. We suggest that <I>I</I><SUB><I>Na</I></SUB> inhibition by eugenol contributes to its analgesic effect.</P>