Food Compounds Activating Thermosensitive TRP Channels in Asian Herbal and Medicinal Foods

J Nutr Sci Vitaminol, 61, S86–S88, 2015

Tatsuo Watanabe and Yuko Terada

School of Food and Nutritional Sciences, University of Shizuoka, 52–1 Yada, Suruga-ku, Shizuoka 422–8526, Japan

Summary There are several thermosensitive transient receptor potential (TRP) ion channels including capsaicin receptor, TRPV1. Food components activating TRPV1 inhibit body fat deposition through sympathetic nerve stimulation. TRPA1 is another pungency sensor for pungent compounds and is mainly coexpressed with TRPV1 in sensory nerve endings. Therefore, TRPA1 activation is expected to have an anti-obesity effect similar to TRPV1 activation. We have searched for agonists for TRPV1 and TRPA1 in vitro from Asian spices by the use of TRPV1- and TRPA1-expressing cells. Further, we performed food component addition tests to high-fat and high-sucrose diets in mice. We found capsiate, capsiconiate, capsainol from hot and sweet peppers, several piperine analogs from black pepper, gingeriols and shogaols from ginger, and sanshools and hydroxysanshools from sansho (Japanese pepper) to be TRPV1 agonists. We also identified several sulfides from garlic and durian, hydroxy fatty acids from royal jelly, miogadial and miogatrial from mioga (Zingiber mioga), piperine analogs from black pepper, and acetoxychavicol acetate (ACA) from galangal (Alpinia galanga) as TRPA1 agonists. Piperine addition to diets diminished visceral fats and increased the uncoupling protein 1 (UCP1) in interscapular brown adipose tissue (IBAT), and black pepper extract showed stronger effects than piperine. Cinnamaldehyde and ACA as TRPA1 agonists inhibited fat deposition and increased UCP1. We found that several agonists of TRPV1 and TRPA1 and some agonists of TRPV1 and TRPA1 inhibit visceral fat deposition in mice. The effects of such compounds on humans remain to be clarified, but we expect that they will be helpful in the prevention of obesity. Key Words thermosensitive TRP channels, TRPV1, TRPA1, Food compounds

In human, capsaicin, the pungent compound in hot Cells expressing TRPV1 or TRPA1 were seeded in 96-well pepper, is detected through transient receptor potential microplates and cultured for 24 h. After calcium indi- (TRP) ion channel vanilloid 1 (TRPV1). TRPV1 is also cator (e.g., Fluo-4 AM) was loaded, cell-based assays activated by heat (.43˚C) and protons. Since the discov- were performed by the use of multimode fluorescent ery of TRPV1 (1), several thermosensitive TRP channels microplate reader FlexStation II (Molecular Devices, have been found. Food components activating TRPV1 Sunnyvale, CA). The changes of fluorescent value after inhibit abdominal fat deposition through sympathetic sample addition to cells were monitored. To check the nerve stimulation (2). On the other hand, TRPA1 is non-specific responses, antagonist (capsazepine or BCTC another pungency sensor for pungent compounds such for TRPV1, HC030031 for TRPA1) addition tests were as allyl isothiocyanate (AITC, pungent compounds of carried out and the responses in parent HEK cells not mustard or wasabi) and is mainly coexpressed with expressing TRP channels were also checked. TRPV1 in sensory nerve endings. Therefore, both From Capsicum fruits (hot or sweet pepper) we found TRPV1 and TRPA1 activation can prevent obesity in the that some non or very low pungent compounds as same way as capsaicin. TRPV1 agonists (capsiate: (3), capsiconiate: (4), cap- We investigated the activation of both TRPV1 and saicinol: (5)). Capsiate was a strong agonist like capsa- TRPA1 by food components mainly from Asian herbal icin. From black pepper, several piperine analogs were and medicinal plants using cells expressing these chan- found to be TRPV1 agonists (6). In ginger, gingerols and nels in vitro. Further, some TRPA1 agonists were tested shogaols were relatively strong agonists for TRPV1 and if they could induce adrenaline secretion in rats. And the potency for shogaols was slightly stronger than that we performed feeding experiments of several TRPV1 or for gingerols (7). Interestingly, 10-shogaol was a strong TRPA1 agonists in high-energy diets for one month on TRPV1 agonist but had very low pungency. For sansho visceral fat accumulation in mice. (Japanese pepper), we tested 8 kinds of sanshools and hydroxysanshools for TRPV1 activation and found that In Vitro Experiments Using TRPV1- or TRPA1- all of them were weak agonists but had similar potency Expressing Cells as that in human sensory analyses (8). Further, we Human embryonic kidney (HEK) 293 cells were used. found that 1-monoacylglycerol having certain acyl moi- eties from wheat, mioga (Zingiber mioga) and onion are E-mail: [email protected] agonists for TRPV1 (9).

S86 Food Compounds Activating Thermosensitive TRP Channels in Asian Spice S87

From Capsicum fruits Animal Experiments O To show the possibility that TRPA1 agonist could H3CO O capsiate enhance energy metabolism, adrenaline responses to AITC and cinnamaldehyde (CNA) were measured in HO blood from the adrenal vein of anesthetized rats. Intra- H CO O capsiconiate 3 venous administration of AITC or CNA induced adrena- O HO O OH line secretion (13). These responses were blocked by cholinergic blockers showing the participation of adre- H3CO N H capsaicinol nal sympathetic nerve activity. Further, capsaicin-treat-

HO ment, which impairs sensory nerve function, abolished the response due to AITC or CNA. These results suggest From black pepper O that TRPA1 agonists induce adrenaline secretion via O N piperolein A sensory nerve activation like TRPV1 agonist capsaicin. O Effect of food components on visceral fat accumulation in mice fed high-fat and high-sucrose diet was From ginger O

H3CO also evaluated as an in vivo test. Feeding a high-fat 6-shogaol and high-sucrose (HFS) diet for 1 mo induced obesity

HO O OH in C57BL mice. Addition of 1-oleoylglycerol to this diet

H3CO resulted in an increase of UCP1 in interscapular brown 6-gingerol adipose tissue (BAT) (14). With the addition of 0.03 or HO 0.05% of piperine or black pepper extract containing From Japanese pepper (sansho) same amount of piperine, the deposition of visceral fats O was abolished and UCP1 contents in interscapular BAT

N increased (15). In addition, CNA feeding with the HFS H R R = H, α-sanshool diet inhibited the accumulation of visceral fats at least R = OH, hydroxy α-sanshool due to BAT activation (16).

From garlic Conclusion S S S S S S We found that several agonists of TRPV1 and TRPA1 diallyl sulfide diallyl disulfide diallyl trisulfide and some agonists of TRPV1 and TRPA1 inhibit visceral From durian fat deposition in mice. The effects of such compounds S S S S S S S on humans remain to be clarified, but we expect that diethyl disulfide diethyl trisulfide dipropyl disulfide they will be helpful in the prevention of obesity. In fact, From mioga From galangal supplementation of capsiate-containing pepper (CH-19 Sweet) reduces visceral fat in humans (17). CHO AcO OAc REFERENCES

CHO CHO 1) Caterina MJ, Schumacher MA, Tominaga M, Rosen TA, miogatrial 1'-acetoxychavicol acetate Levine JD, Julius D. 1997. The capsaicin receptor: a From royal jelly heat-activated ion channel in the pain pathway. Nature 10-hydroxydeca- 389: 816–824. HO COOH trans-2-enoic acid 2) Iwai K, Yazawa Y, Watanabe T. 2003. Roles as metabolic regulators of the non-nutrients, capsaicin and capsiate, Fig. 1. TRPV1 and/or TRPA1 agonists from Asian supplemented to diets. Proc Japan Acad 79B: 207–212. spices. 3) Iida T, Moriyama T, Kobata K, Morita A, Murayama N, Hashizume S, Fushiki T, Yazawa S, Watanabe T, Tomi- naga M. 2003. TRPV1 activation and induction of noci- We identified several sulfides from garlic (10) and ceptive response by a non-pungent capsaicin-like com- durian (11) as both TRPA1 and TRPV1 agonists but the pound, capsiate. Neuropharmacology 44: 958–967. potency of all compounds to TRPA1 was stronger than 4) Kobata K, Tate H, Iwasaki Y, Tanaka Y, Ohtsu K, Yazawa that to TRPV1. From mioga, we found that miogadial S, Watanabe T. 2008. Isolation of coniferyl esters from and miogatrial are strong agonists for TRPA1. Piperine Capsicum baccatum L., and their enzymatic preparation and agonist activity for TRPV1. Phytochemistry 69: analogs from black pepper were also TRPA1 agonists but 1179–1184. the potency of these analogs for TRPV1 was stronger 5) Kobata K, Iwasawa T, Iwasaki Y, Morita A, Suzuki Y, than that for TRPA1 (6). Acetoxychavicol acetate (ACA) Kikuzaki H, Nakatani N, Watanabe T. 2006. Capsa- is a pungent compound in galangal (Alpinia galanga) and icinol: synthesis by allylic oxidation and its effect on we found that ACA is a TRPA1 agonist (12). Further, we TRPV1-expressing cells and adrenaline secretion in rats. clarified that some of them are stronger than the most Biosci Biotechnol Biochem 70: 1904–1912. well-known TRPA1 agonist, AITC. 6) Okumura Y, Narukawa M, Iwasaki Y, Ishikawa A, Mat- suda H, Yoshikawa M, Watanabe T. 2010. Activation of TRPV1 and TRPA1 by black pepper compounds. Biosci S88 Watanabe T and Terada Y

Biotechnol Biochem 74: 1068–1072. 13) Iwasaki Y, Tanabe M, Kobata K, Watanabe T. 2008, 7) Iwasaki Y, Morita A, Iwasawa T, Kobata K, Sekiwa Y, TRPA1 agonists—allyl isothiocyanate and cinnamal- Morimitsu Y, Kubota K, Watanabe T. 2006. A nonpun- dehyde—induce adrenaline secretion, Biosci Biotechnol gent component of steamed ginger—[10]-shogaol— Biochem 72: 2608–2614. increases adrenaline secretion via the activation of 14) Iwasaki Y, Tamura Y, Inayoshi K, Narukawa M, Kobata TRPV1. Nutr Neurosci 9: 169–178. K, Chiba H, Muraki E, Tsunoda N, Watanabe T. 2011. 8) Sugai E, Morimitsu Y, Iwasaki Y, Morita A, Watanabe T, TRPV1 agonist monoacylglycerol increases UCP1 con- Kubota K. 2005. Pungent qualities of sanshool-related tent in brown adipose tissue and suppresses accumu- compounds evaluated by a sensory test and activation of lation of visceral fat in mice fed a high-fat and high- rat TRPV1. Biosci Biotechnol Biochem 69: 1951–1957. sucrose diet. Biosci Biotechnol Biochem 75: 904–909. 9) Iwasaki Y, Saito O, Tanabe M, Inayoshi K, Kobata K, Uno 15) Okumura Y, Narukawa M, Watanabe T. 2010. Adipos- S, Morita A, Watanabe T. 2008. Monoacylglycerols acti- ity suppression effect in mice due to black pepper and vate capsaicin receptor, TRPV1. Lipids 43: 471–483. its main pungent component, piperine. Biosci Biotechnol 10) Koizumi K, Iwasaki Y, Narukawa M, Iitsuka Y, Fukao Biochem 74: 1545–1549. T, Seki T, Ariga T, Watanabe T. 2009. Diallyl sulfides in 16) Tamura Y, Iwasaki Y, Narukawa M, Watanabe T. 2012. garlic activate both TRPA1 and TRPV1. Biochem Biophys Ingestion of cinnamaldehyde, a TRPA1 agonist, reduces Res Commun 382: 545–548. visceral fats in mice fed a high-fat and high-sucrose diet. 11) Terada Y, Hosono T, Seki T, Ariga T, Ito S, Narukawa M, J Nutr Sci Vitaminol 58: 9–13. Watanabe T. 2014. Sulphur-containing compounds of 17) Kawabata N, Inoue N, Yazawa S, Kawada T, Inoue K, durian activate the thermogenesis-inducing receptors Fushiki T. 2006. Effect of CH-19 Sweet, a non-pungent TRPA1 and TRPV1. Food Chem 157: 213–220. cultivar of red pepper, in decreasing the body weight 12) Narukawa M, Koizumi K, Iwasaki Y, Kubota K, Wata- and suppressing body fat accumulation by sympathetic nabe T. 2010. Galangal pungent component, 1’-ace- nerve activation in humans. Biosci Biotechnol Biochem toxychavicol acetate, activates TRPA1. Biosci Biotechnol 70: 2824–2835. Biochem 74: 1694–1696.