Chapter 14
Capsaicinoid Oxidation by Peroxidases: Kinetic, Structural, and Physiological Considerations
Downloaded by EMORY UNIV on March 11, 2016 | http://pubs.acs.org Publication Date: July 21, 2005 | doi: 10.1021/bk-2005-0909.ch014
Douglas C . Goodwin, Kimberley A . Laband, and Kristen M. Hertwig Department of Chemistry and Program in Cell and Molecular Biosciences, Auburn University, Auburn, A L 36849-5312
Capsaicinoids, biosynthesized exclusively in Capsicum fruits or "hot" peppers, are o-methoxyphenols that have substantial chemopreventive potential. Daily, Capsicum fruits are consumed by vast segments of the world's population. The levels of capsaicinoids in these fruits (and hence their desirability for consumption) are intimately linked with the activity of peroxidases, but several questions surrounding the kinetics of the process have gone unanswered. We applied novel transient- and steady-state methods to address the reduction of peroxidase compounds I and II by capsaicinoids and to obtain steady-state kinetic parameters for capsaicinoid oxidation. Comparison of transient-state and steady-state data identify electron transfer from capsaicinoids to compound II as rate-determining. Ascorbate rapidly reduces capsaicinoid radicals, a property essential for the kinetic studies. However, evidence also suggests that ascorbate is an important factor in capsaicinoid content of Capsicum fruits. The 4-substituent of o-methoxyphenols has a dramatic influence on rates of oxidation by peroxidases. However, our results suggest that kinetic parameters for oxidation of other chemopreventive o-methoxyphenols are likely to be similar to the capsaicinoids.
© 2005 American Chemical Society Shahidi and Ho; Phenolic Compounds in Foods and Natural Health Products ACS Symposium Series; American Chemical Society: Washington, DC, 2005.
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162 As the reports of this symposium firmly establish, evidence continues to accumulate suggesting that naturally-occuring phenolic and polyphenolic compounds have substantial health benefits. The o-methoxyphenols represent an important class of these increasingly prominent phytochemicals. Indeed, recent reports indicate that gingerol (the principle pungent compound in ginger) (1,2) and curcumin (a key component of tumeric) (3-6) show promise as chemopreventive agents through a variety of mechanisms (7). It is also important to note that these compounds impart the desirable or distinctive characteristics to the agricultural/food products that include them, ensuring their consumption in abundance by large sectors of the world's population. In this sense, the capsaicinoids, produced only by plants of the Capsicum genus, are a very important group of o-methoxyphenols. They are the major contributors to the pungency of the Capsicumfruits(i.e., "hot" peppers). The levels of capsaicinoids in Capsicum fruits vary greatly among cultivars, some producing levels as high as 1% (8). Over one-quarter of the world's population consumes Capsicumfruitsor related products on a daily basis, and the primary driving force behind the desirability of these fruits is their capsaicinoid content (9). As with gingerol and curcumin, capsaicinoids (Figure 1A) are effective antioxidant and anti-inflammatory compounds with strong potential for prevention of tumor promotion (7,10-12). Clearly, the factors contributing to the levels of capsaicinoids in Capsicumfruitshave important agricultural, cultural, and biomedical implications.
Capsaicinoids and Peroxidase-Catalyzed Oxidation Peroxidases occupy a central position in capsaicinoid catabolism. Capsaicinoid degradation has been correlated with a rise in peroxidase activity during Capsicum fruit maturation (13). Plant peroxidases have been shown to oxidize capsaicin to the lignin-like dimers, 5,5'-dicapsaicin and 4-0-5 dicapsaicin ether, and further oxidation of these compounds by peroxidases leads to formation of high molecular weight polymers (14). The typical peroxidase catalytic cycle (Figure IB) is initiated by the reaction between ferric peroxidase and H 0 . In the reaction, the heme is oxidized by two electrons to form the ferryl-oxo porphyrin radical intermediate known as compound I. The porphyrin radical of compound I is then reduced by one electron by an exogenous electron donor (i.e., reducing substrate) to produce the ferryl-oxo intermediate compound II and one equivalent of substratefreeradical. 2
2
Shahidi and Ho; Phenolic Compounds in Foods and Natural Health Products ACS Symposium Series; American Chemical Society: Washington, DC, 2005.
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Compound II is then reduced by a second equivalent of exogenous electron donor yielding the ferric peroxidase and a second equivalent of substrate radical (15). Though capsaicinoids are well-known as reducing substrates for plant peroxidases (16-20% several kinetic aspects of the process have remained poorly defined. Until recently, the reactions of peroxidase compound I and compound II with capsaicinoids had not been directly observed nor the rate constants determined. Furthermore, the formation of a broad range of polymeric products (each with its own unique absorption characteristics) had hampered the assignment of steady-state kinetic parameters for capsaicinoid oxidation by peroxidases.
Β
Ferric 2 ° 2
©"
2
/
Cmpdl V I
- G.=°
2
OCH
Dihydrocapsaicin
v
Η 0 + Α·
Capsaicin
OH
H0
H
3
OH
Nonivamide
Figure 1. Structures of some of the capsaicinoids and their proposed interaction with the peroxidase catalytic cycle. Panel A shows the structures of the two most naturally abundant capsaicinoids, capsaicin and dihydrocapsaicin. Nonivamide is often referred to as synthetic capsaicin " but was recently identified as a natural product. Panel Β shows the peroxidase catalytic cycle. The capsaicinoids are proposed to be reducing substrates (AH) for peroxidase compounds I and II f
