Frederick Khachik

Distribution and metabolism of dietary carotenoids in humans as a criterion for development of nutritional supplements

There are approximately 40-50 carotenoids in commonly consumed fruits and vegetables in a typical U.S. diet. These can be divided into carotenoid epoxides, mono- and dihydroxycarotenoids, hydrocarbon carotenoids, and carotenol acyl esters. However, among these, only a selected group of carotenoids are routinely found in human plasma, breast milk, major organs, and ocular tissues. In addition, several carotenoid metabolites have also been isolated and characterized from human plasma, tissues, and ocular tissues. The proposed metabolic transformation of carotenoids in humans will be discussed. Dietary carotenoids and their metabolites have been implicated in the prevention of cancer, cardiovascular disease, and age-related macular degeneration (AMD). An approach for the development of a nutritional supplement that is based on the distribution of carotenoids and their metabolites in humans will be discussed.

Lycopene Uptake and Tissue Disposition in Male and Female Rats

Abstract Epidemiologic and clinical studies suggest that tomato consumption may reduce the risk of cancer. Lycopene, a hydrocarbon carotenoid, is the major carotenoid in tomatoes and, as a potent singlet oxygen quencher, has been considered by some to be the biologically active agent responsible for the reduction of cancer risk associated with tomato consumption. However, little is known concerning Iycopene absorption or biological activity in rodent models of cancer. Therefore, the present study was designed to provide information regarding the uptake and tissue disposition of Iycopene and related carotenoid after feeding a diet containing a carotenoid mixture extracted from tomatoes (Betatene). Betatene was added to the diet at 2.3, 0.9, 0.45, 0.23, 0.09 and 0 (mM/kg diet) and fed to male and female Fischer-344 rats for a period of 10 weeks. Using reverse phase HPLC methods, it was found that approximately 55% of administered Iycopene was excreted in the feces. In both males and females, Iycopene concentrations were highest in the liver (120-42 lJg/g wet wt.); physiologically significant levels were detected in prostate (97-47 ng/g), lung (227-134 ng/g), mammary gland (309-174 ng/g) and serum (285-160 ng/ml). Tissue concentrations were related to dose with the exception of serum, and differences between males and females were minimal. Other carotenoids present in Betatene (l.e., phytoene, phytofluene, z-carotene and l3-carotene) were also absorbed and stored in the liver. These results indicate that Iycopene, when incorporated into the semi purified AIN-76A diet, is absorbed in both male and female rats in a doserelated manner and can be detected at nanogram levels in a variety of target organs.

Effect of dietary lycopene on N-methylnitrosourea-induced mammary tumorigenesis.

Epidemiological studies suggest protective effects of lycopene-rich foods on several types of cancer, including prostate and gastrointestinal tract. Moreover, an inverse association between serum lycopene concentrations and several types of cancer has been reported. However, few studies have focused on breast cancer, and they have shown little association between lycopene consumption and cancer risk. In this report, we used the N-methylnitrosourea (NMU)-induced rat mammary tumor model to compare the effects of pure lycopene with a lycopene-rich tomato carotenoid oleoresin (TCO) on NMU-induced mammary tumorigenesis. Rats were fed diets supplemented with 250 and 500 ppm crystalline lycopene or TCO beginning seven days before initiation with NMU (55 days of age) to termination (18 wk after NMU). Neither pure lycopene nor lycopene in the form of a mixed carotenoid oleoresin exerted an inhibitory effect on tumor incidence, latency, multiplicity, volume, or total tumors per group compared with unsupplemented controls. Weight gains in all groups were similar. Assay of serum lycopene concentrations in lycopene-supplemented groups indicated that median levels of 7,12,60, and 87 ng/ml were attained in blood of groups supplemented with 250 and 500 ppm lycopene and 250 and 500 ppm TCO, respectively. The results of this animal study are consistent with epidemiological reports indicating that lycopene does not protect against breast cancer.

Lycopene Supplementation Prevents the Development of Spontaneous Smooth Muscle Tumors of the Oviduct in Japanese Quail

Abstract: Leiomyomas (fibroids) are benign tumors of the uterus affecting millions of women. Spontaneous leiomyomas of the oviduct are common tumors of the Japanese quail (Coturnix coturnix japonica), which makes it a good animal model for screening potential agents for testing in the prevention and treatment of human myoma uteri. Because dietary intake of lycopene has been associated with a reduced risk of a variety of human cancers, we investigated the effects of lycopene supplementation on the development of leiomyomas in the oviduct of Japanese quail. We also measured serum levels of oxidative stress markers [malondialdehyde (MDA) and homocysteine], lycopene, vitamins C, E, and A, and tissue biomarkers Bcl-2 and Bax expression. One hundred twenty quails (6 mo old) were assigned to 3 treatment groups consisting of 4 replicates of 10 birds in each group. Birds were fed either a basal diet (group C) or the basal diet supplemented with 100 mg (group L1) or 200 mg (group L2) of lycopene per kilogram of diet. The animals were sacrificed after 285 days and the tumors were identified. Lycopene supplementation decreased the number of leiomyomas compared with control subjects (P = 0.056). The tumors in lycopene-fed birds were smaller than those found in control birds (P = 0.01). There were no significant differences in the expression of tissue Bcl-2 and Bax among the study groups. Serum vitamins C, E, and A increased (P = 0.01), whereas MDA and homocysteine concentrations decreased (P = 0.01) with lycopene supplementation. No measurable lycopene could be detected in the serum of control birds, whereas a dose-dependent increase was observed in the serum of lycopene-supplemented birds. The results indicate that dietary supplementation with lycopene reduces the incidence and size of spontaneously occurring leiomyoma of the oviduct in the Japanese quail. Clinical trials should be conducted to investigate the efficacy of lycopene supplementation in the prevention and treatment of uterine leiomyoma in humans.

14 – Dietary Carotenoids and their Metabolites as Potentially Useful Chemoprotective Agents against Cancer

Publisher Summary This chapter emphasizes the importance of other dietary carotenoids in addition to β-carotene, and presents evidence of the nutritional significance of this important class of nutrients in the prevention of cancer as well as macular degeneration, a degenerative eye disease. This was initially based on the knowledge of widespread distribution of carotenoids in fruits and vegetables, as well as the consistent presence of these compounds and their metabolites in human serum, milk, and tissues at relatively high concentrations. Carotenoids are among the most widespread of the naturally occurring groups of pigments and are found in all families of the plant and animal kingdoms. The correlation between dietary carotenoids and carotenoids found routinely in the extracts of human serum/plasma has revealed that only selected groups of carotenoids make their way into the human bloodstream. Some of these carotenoids are absorbed intact and others, such as lutein, zeaxanthin, and lycopene, are converted to several metabolites. Some dietary carotenoids are also present in human tissues, for example, the liver, lung, breast, and cervix. This chapter identifies several new optical isomers of carotenoid metabolities in human plasma, and provides additional evidence of previously proposed metabolic oxidation–reduction reactions of dietary carotenoids in humans. It also presents studies that indicate that carotenoids, in addition to their antioxidant mechanism of action, can exert their biological activity in disease prevention by other mechanisms.

18 – Enhanced Activity of an Oxidation Product of Lycopene Found in Tomato Products and Human Serum Relevant to Cancer Prevention

Lycopene can increase connexin 43 gene expression in 10T1/2 cells and in human keratinocytes in organotypic culture. This chapter investigates that an oxidation product of lycopene; 2,6-cyclolycopene-1,5-diol; is found in tomato products and it increases in human serum after ingestion of these products. This compound is shown to possess greater activity than the parent molecule in its ability to increase connexin 43 levels in mouse and human cells. The result emphasizes that lycopene may function both as an antioxidant, protecting cells from oxidative damage, and as the source of a molecule with biological actions that may additionally protect cells from neoplasia. Oxidative metabolite of lycopene is present in foods at very low concentrations, the preferential uptake of lycopene from foods can account for its disproportionately large presence in serum. The chapter discusses that the in vivo oxidation of lycopene humans may also lead to oxidative metabolite.

Dietary Tomato Powder Supplementation in the Prevention of Leiomyoma of the Oviduct in the Japanese Quail

Abstract Spontaneous leiomyomas of the oviduct are common tumors of the Japanese Quail (Coturnix coturnix japonica), which makes it a good animal model for screening potential agents for testing in the prevention and treatment of human myoma uteri. We have previously reported a decreased incidence of leiomyomas in the oviduct of Japanese quail with lycopene supplementation. Although the major carotenoid in tomatoes is lycopene, tomatoes also contain other compounds, which may contribute to their health benefit. Therefore, in this study, we investigated the effects of tomato powder supplementation on the development of leiomyomas in the oviduct of Japanese quail. We also measured serum levels of malondialdehyde (MDA), carotenoids, and vitamins C, E, and A. A total of 150 quails (3 mo old) were assigned to 3 treatment groups consisting of 5 replicates of 10 birds in each group. Birds were fed either a basal diet (control group) or the basal diet supplemented with 25 g (Treatment I) or 50 g (Treatment II) of tomato powder (0.8 mg lycopene per g of tomato powder) per kg of diet. The animals were sacrificed after 365 days, and the tumors were identified. Tomato powder supplementation significantly decreased the number of leiomyomas as compared to control birds (P < 0.01). The tumors in tomato powder fed birds were smaller than those found in control birds (P < 0.01). Serum lycopene, lutein, zeaxantin, and vitamins C, E, and A increased (P = 0.01), whereas MDA concentrations decreased (P = 0.01) with tomato powder supplementation. No measurable lycopene could be detected in the serum of control birds, whereas a dose-dependent increase was observed in the serum of birds supplemented with tomato powder. The results indicate that dietary supplementation with tomato powder reduces the incidence and size of spontaneously occurring leiomyoma of the oviduct in the Japanese quail. Clinical trials should be conducted to investigate the efficacy of tomato powder supplementation in the prevention and treatment of uterine leiomyoma in humans.

The regiochemistry of trapped radical anion-radical cation pairs from aryl substituted phthalimides

Abstract A series of 4-substituted N-methyl phthalimides was irradiated in the presence of 2,3-dimethyl-2-butene in methanol and the product distributions of the trapped radical anion-radical cation analyzed on the basis of substituent effects on the radical anion.

Cycloadditions of hexachlorocyclopentadiene to 7-substituted norbornadienes: remote substituent effects on reacfivity and stereoselectivity

Abstract The rates and stereoselectivities of the cycloadditions of hexachlorocyclopentadiene to norbornadienes substituted at the 7-position by tert -butyl, trimethylsilyl, methoxy, acetoxy, hydroxy, methoxymethyl, and methoxycarbonyl groups have been measured. The rates correlate with subsfituent electronegativities.

Highlights of carotenoid research from the 17th International Carotenoid Symposium.

Fig. 1. Yearly number of publications in Archives of Biochemistry and B focused on carotenoid science between 1996 and 2015. iophysics This thematic issue dedicated to carotenoids in Archives of Biochemistry and Biophysics (ABB) is the seventh issue predominantly or exclusively focused on the fascinating chemistry, physiology, and spectroscopy of these ubiquitous, colorful pigments synthesized for photosynthesis and photoprotection exclusively by plants and micro-organisms, yet utilized by a wide variety of vertebrates and invertebrates for a diverse array of functions including vision, cancer prevention, and protective coloration. Archives of Biochemistry and Biophysics has had a long history of publishing research and review articles on carotenoids. A keyword search on ‘‘carotenoid’’ reveals that 374 articles were published in ABB on this topic before 1996. Since that time, more carotenoid manuscripts have been published in ABB, but there are obvious periodic spikes of articles (Fig. 1). The initial peak corresponds with ABB Executive Editor Helmut Sies’ ‘‘experiment’’ to invite selected participants from the 2001 Gordon Conference on Carotenoids to contribute minireviews in their fields of expertise. This experiment proved successful enough that special issues on carotenoids have continued to coincide with the Gordon Conference on that topic every three years since then, with spikes of carotenoid publications seen in 2004, 2007, 2010, and 2013. In 2009, another special issue on carotenoids coincided with the International Carotenoid Symposium held one year earlier in Okinawa, Japan. The current issue contains a record number of articles on carotenoids, with seven minireviews and 18 original research publications from participants in the 17th International Carotenoid Symposium held in Park City, Utah, USA from June 29 to July 4, 2014. This triennial meeting organized by the International Carotenoid Society attracted 217 scientists from 33 countries working in an extraordinarily broad array of scientific disciplines who share a common interest in carotenoids. They spent five focused days in the beautiful Wasatch Mountains sharing their latest scientific findings and initiating future productive cross-discipline collaborations. The diversity of interests is clearly evident if one peruses the table of contents where there are articles of interest to carotenoid biologists, biophysicists, botanists, cancer researchers, chemists, epidemiologists, nutritionists, physiologists, and vision scientists. The editors of this special issue would like to express our deep appreciation to Professor Helmut Sies for initiating this ongoing tradition that promotes and recognizes excellence in carotenoid science. The editorial staff at ABB kept us all on task to produce a high quality special issue in a timely manner. We look forward to continuing progress in the field that will be highlighted in future issues of ABB.