polyphenol-enriched diet for groups at risk of iron loading, for example patients with hereditary haemochromatosis. The flavonol quercetin is the most abundant dietary flavonoid and is especially enriched in onions, tea and apples. It is conservatively estimated that humans consume approximately 40 80 mg of flavonoids/day; and that quercetin contributes approximately 25% of total flavonoid intake . In common with most polyphenols, quercetin is found almost exclusively in foods as glycoside conjugates but can be converted rapidly into PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19657297 the aglycone in the intestinal lumen via the actions of glycosidases. In this study we have investigated the acute and longer-term effects of quercetin on iron metabolism in vivo and in vitro. NaCl, 0.1 mmol/L ascorbic acid, 0.01 mmol/L FeCl3 to displace any 59Fe bound to the mucosal surface. Subsequently, the duodenal mucosa was scraped away, the blood and mucosa samples were weighed and gamma counted for determination of 59 Fe activity. Results were expressed as percentage of absorbed radioactive iron retained in duodenal mucosa or transferred to blood. The percentage of 59Fe 
transferred to the entire blood volume of the animal was calculated using the equation: total blood volume = +0.77. Effects of oral gavage of quercetin on iron metabolism In order to investigate the longer-term effects of quercetin administration on non-haem iron absorption, animals were gavaged with a single dose of quercetin or vehicle eighteen hours before experiments. Animals were anesthetized with pentobarbitone sodium injected IP. The duodenum and liver were collected, snap frozen in liquid N2, and stored at 280uC for real-time PCR analysis and tissue non-haem iron determination. RNA extraction and RT-PCR Duodenal RNA was extracted with TRIzol purchase A-83-01 according to the manufacturer’s instructions. Total RNA was reverse transcribed using the Verso cDNA reverse transcription kit, according to the manufacturer’s instructions. Real-time PCR reactions were performed using a Roche Lightcycler using GAPDH as internal standard. Each reaction was performed in duplicate and contained 10 pmol of specific primers, 16SYBR Green Mastermix, and 1 mL of cDNA in a 20 mL reaction. Samples without cDNA were included as negative controls. Cycle threshold values were obtained for each gene of interest and the GAPDH internal standard. Gene expression was normalized to GAPDH and represented as DCt values. For each sample the mean of the DCt values was calculated. Relative gene expression was normalized to 1.0 of controls. Material and Methods Animals and treatments Rats were supplied by the Comparative Biology Unit, Royal Free Campus, UCL Medical School, London, UK. All experimental procedures were approved by the University College London local animal ethics committee and were conducted in accordance with the UK Animals Act, 1986. After weaning Sprague-Dawley male rats were placed on low iron diet for two weeks and allowed free access to water throughout. At the end of the experimental procedure animals were killed by administering a terminal dose of pentobarbitone sodium. Levels of tissue non-haem iron Quantitative measurement of non-haem iron was performed according to the modified method of Torrance and Bothwell. Briefly, 2050 mg of duodenum was dried at 55uC for 72 hours and subsequently weighed. Dried samples were digested in 1 mL acid mixture HCl and 10% trichloroacetic acid) at 65uC for 20 hours. 1 mL of chromogen reagent bathophenanthrolinesulfonate