Study of Chlorophyll-related Compounds from Dietary Spinach in Human Blood

Pi-Yu CHAO, Meng-Yuan HUANG, Wen-Dar HUANG, Kuan-Hung Robert LIN, Shiau-Ying CHEN, Chi-Ming YANG


Human bioavailability data on chlorophyll (Chl) is very limited. The distribution of Chl-related compounds (CRCs) derived from dietary spinach was investigated in human blood. Eight healthy adults, aged from 21 to 61 year-old, consumed 1.2 kg of just-boiled fresh spinach after an 8-h overnight fast. Before and then 3 h after consuming the spinach, blood samples were taken from each participant. Freeze-dried blood samples were prepared, and 80% acetone was added for grinding. Eight peaks were found in the blood using high-performance liquid chromatography (HPLC), and the main CRCs in the samples were pheophytin (Phe) and pheophorbide (Pho) derivates. Compared to a fasted state, markedly higher levels of blood CRCs were detected in all subjects, except that Pho metabolites were not found in two subjects. No significant differences were seen in most of the peaks between males and females; however, relatively higher CRCs levels were observed in females, particular of Pho derivates. In addition, the blood contained significantly higher levels of Phe in the 36~61-year-old group than in the 21~35-year-old group. These results suggest that the conversion of Chls to CRCs is a rapid process, and Chls obtained by ingestion can be absorbed by the human body.


chlorophyll; high-performance liquid chromatography; human bioavailability; pheophytin; pheophorbide; pigments

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Aprahamian M, Evrard S, Keller P, Tsuji M, Balboni G, Damgé C, Marescaux J (1993). Distribution of pheophorbide A in normal tissues and in an experimental pancreatic cancer in rats. Anticancer Drug Design 8:101-114.

Bellnier DA, Henderson BW, Pandey RK, Potter WR, Dougherty TJ (1993). Murine pharmacokinetics and antitumor efficacy of the photodynamic sensitizer 2-[1-hexyloxyethyl]-2-devinyl pyropheo-phorbide-a (HPPH). Journal of Photochemistry and Photobiology B 20:55-61.

Bellnier DA, Greco WR, Loewen GM, Nava H, Oseroff AR, Dougherty TJ (2006). Clinical pharmacokinetics of the PDT photosensitizers porfimer sodium (Photofrin), 2-[1-hexyloxyethyl]-2-devinyl pyropheo-phorbide-a (photochlor) and 5-ALA-induced protoporphyrin IX. Lasers in Surgery and Medicine 38:439-444.

Chernomorsky S, Segelman A, Poretz RD (1999). Effect of dietary chlorophyll derivatives on mutagenesis and tumor cell growth teratogen. Carcinogens and Mutagens 19:313-322.

Dandler J, Wilhelm B, Scheer H (2010). Distribution of chlorophyll- and bacteriochlorophyll-derived photosensitizers in human blood plasma. Photochemistry and Photobiology 86:182-193.

De Vogel J, Jonker-Termont DS, Van Lieshout EM, Katan MB, Van der Meer R (2005). Green vegetables, red meat and colon cancer: Chlorophyll prevents the cytotoxic and hyperproliferative effects of haem in rat colon. Carcinogens 26:387-393.

Edenharder R, Keller G, Platt KL, Unger KK (2001). Isolation and characterization of structurally novel antimutagenic flavonoids from spinach (Spinacia oleracea). Journal of Agriculture and Food Chemistry 49:2767-2773.

Erlanson-Albertsson C, Per-Åke A (2015). The use of green leaf membranes to promote appetite control, suppress hedonic hunger and loose body weight. Plant Foods and Human Nutrition 70:281-290.

Eva-Lena S, Egecioglu E, Landin-Olsson M, Erlanson-Albertsson C (2015). Consumption of thylakoid-rich spinach extract reduces hunger, increases satiety and reduces cravings for palatable food in overweight women. Appetite 91:209-219.

Fernandes TM, Gomes BB, Lanfer-Marquez UM (2007). Apparent absorption of chlorophyll from spinach in an assay with dogs. Innovative Food Science & Emerging Technologies 8:426-432.

Ferruzzi MG, Blakeslee JD (2007). Absorption, and cancer preventative activity of dietary chlorophyll derivatives. Nutrition Research 27:1-12.

Ferruzzi MG, Failla ML, Schwartz SJ (2001). Assessment of degradation and intestinal cell uptake of carotenoids and chlorophyll derivatives from spinach puree using an in vitro digestion and Caco-2 human cell model. Journal of Agriculture and Food Chemistry 49:2082-2089.

Ferruzzi MG, Lumpkin JL, Schwartz SJ, Failla M (2006). Digestive stability, micellarization, and uptake of beta-carotene isomers by Caco-2 human intestinal cells. Journal of Agriculture and Food Chemistry 54:2780-2785.

Gallardo-Guerrero L, Gandul-Rojas B, Mínguez-Mosquera MI (2008). Digestive stability, micellarization, and uptake by caco-2 human intestinal cell of chlorophyll derivatives from different preparations of pea (Pisum sativum L.). Journal of Agriculture and Food Chemistry 56:8379-8386.

Gandul-Rojas B, Gallardo-Guerrero L, Mínguez-Mosquera MI (2009). Influence of the chlorophyll pigment structure on its transfer from an oily food matrix to intestinal epithelium cells. Journal of Agriculture and Food Chemistry 57:5306-5314.

Hsu CY, Chao PY, Hu SP, Yang CM (2013). The antioxidant and free radical scavenging activities of chlorophylls and pheophytins. Food Nutrition Science 4:1-8.

Hsu CY, Chen YH, Chao PY, Hu SP (2008). Naturally occurring chlorophyll derivatives inhibit aflatoxin B1-DNA adduct formation in hepatoma cells. Mutation Research 657:98-104.

Hsu CY, Yang CM, Chen CM, Chao PY, Hu SP (2005). Effects of chlorophyll-related compounds on hydrogen peroxide induced DNA damage within human lymphocytes. Journal of Agriculture and Food Chemistry 53:2746-2750.

Hsu CY, Yeh TH, Huang MY, Chao PY, Yang CM (2014). Organ-specific distribution of chlorophyll-related compounds from dietary spinach in rabbits. Indian Journal of Biochemistry and Biophyics 51:388-395.

Jonker J, Buitelaar M, Wagenaar E, Van Der Valk M, Scheffer G, Scheper R (2002). The breast cancer resistance protein protects against a major chlorophyll-derived dietary phototoxin and protoporphyria. Proceedings of the National Academy of Sciences 99:15649-15654.

Lanfer-Marquez UM, Barros RM, Sinnecker P (2005). Antioxidant activity of chlorophylls and their derivatives. Food Research International 38:885-891.

Lin KH, Hsu CY, Huang YP, Lai JY, Huang MY, Yang CM, Chao PY (2013). Chlorophyll-related compounds inhibit cell adhesion and inflammation in human aortic cells. Journal of Medicine Food 16:1-13.

Mazor O, Brandis A, Plaks V, Neumark E, Rosenbach-Belkin V, Salomon Y, Scherz A (2005). WST11, a novel water-soluble bacteriochlorophyll derivative; cellular uptake, pharmacokinetics, biodistribution and vascular-targeted photodynamic activity using melanoma tumors as a model. Photochemistry and Photobiology 81:342-351.

McQuistan TJ, Michael T, Simonich M, Pratt M, Pereira C, Hendricks J, Dashwood R, Williams D, Bailey S (2012). Cancer chemoprevention by dietary chlorophylls: A 12,000-animal dose-dose matrix biomarker and tumor study. Food and Chemical Toxicology 50:341-352.

Nakamura Y, Murakami A, Koshimizu K, Ohigashi H (1996). Inhibitory effect of pheophorbide a, a chlorophyll-related compound, on skin tumor promotion in ICR mice. Cancer Letter 108:247-255.

Pandey RK, Shiau FY, Sumlin AB, Dougherty TJ, Smith KM (1992). Structure/activity relationships among photosensitizers related pheophorbides and bacteriopheophorbides. Bioorganic & Medicinal Chemistry Letters 2:491-496.

Sakdarat S, Shuyprom A, Pientong C, Ekalaksananan T, Thongchai S (2009). Bioactive constituents from the leaves of Clinacanthus nutans Lindau. Bioorganic & Medicinal Chemistry 17:1857-1860.

Simonich MT, McQuistan T, Jubert C, Pereira C, Hendricks JD, Schimerlik M, Zhu B, Dashwood RH, Williams DE, Bailey GS (2008). Low-dose dietary chlorophyll inhibits multi-organ carcinogenesis in the rainbow trout. Food and Chemical Toxicology 46:1014-1024.

Song W, Derito CM, Liu MK (2010). Cellular antioxidant activity of common vegetables. Journal of Agriculture and Food Chemistry 58:6621-6629.

Subramoniam A, Asha VV, Nair SA, Sasidharan SP, Sureshkumar PK, Rajendran KN, Karunagaran D, Ramalingam K (2012). Chlorophyll revisited: anti-inflammatory activities of chlorophyll a and inhibition of expression of TNF-a gene by the same. Inflammation 35:959-966.


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