Dose effect on the uptake and accumulation of hydroxytyrosol and its metabolites in target tissues in rats.

Abstract

The aim of the present study was to investigate the relationship between the 34 hydroxytyrosol (HT) dose intake and the HT metabolites tissue uptake in order to assess 35 if the HT levels detected in tissues after the administration of different doses are 36 relevant and sufficiently compatible with those that have been reported as in vitro 37 modulators of biological functions. Rats were given a refined olive oil enriched with HT 38 at different doses (1, 10 and 100 mg/kg) and after 5 hours they were sacrificed. Plasma 39 samples and different organs as liver, kidney, heart and brain were obtained, and HT  In recent years, hydroxytyrosol (HT) and its derivatives have led to a great interest from 50 the virgin olive oil (VOO) producers and manufacturers of nutraceutical supplements. 51 The increasing interest in HT is mainly due to the EFSA Panel on Dietetic Products, 52 Nutrition and Allergies (NDA) inform that established a cause-and-effect relationship   In this context, the present study aimed to investigate the relationship between 75 the HT dose intake and tissue uptake in rats, and thus, providing complementary 76 information in relation to the target/dose relationship. For this purpose, liver, kidney, 77 brain and heart were collected after the administration of three different doses: 1 mg 78 HT/kg rat weight (HT-1) compatible with human dietary habits [7], and two higher 79 doses (10 mg/kg rat weight: HT-10 and 100 mg/kg rat weight: HT-100) that could  Results of the plasma analysis showed that free HT and its metabolites were 83 detected and were measurable after the three administered doses (Table 1), except for 84 the glucuronide conjugates, which were only detected at the highest doses (HT-10 and 85 HT-100). In the control plasma (vehicle), none of these compounds were detected, 86 which validates them as products of HT metabolism. Sulfation was the most relevant 87 conjugation pathway at the three administered doses compared with the glucuronide 88 conjugates. It is important to highlight that the free form of HT was detected in the 89 plasma after the administration of all three doses and that the recovery of free HT 90 appeared to rise in a dose-dependent fashion (from 0.05 µmol/L HT-1 to 12.9 µmol/L 91 HT-100) as with the metabolites (p trend<0.001).

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The dose effect on the tissue uptake of HT was first studied on two metabolic 93 tissues (liver, kidney) obtained five hours after the intake of the vehicle (refined olive 94 oil) and the different doses of HT ( Table 2). In general, the nature of the HT 95 metabolites was similar to the plasma and a significant dose-dependent uptake was 96 observed for all metabolites studied (p trend<0.001) except for HVAlc in the kidney 97 ( Table 2). Additionally, significant differences were observed for the metabolites 98 quantified at all HT doses compared to the vehicle group (p<0.001).

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Besides the metabolic tissues, two target tissues (brain and heart) were studied 114 showing a lower deposition of HT metabolites ( Table 3). Regarding the brain, some of 115 the HT metabolites were also detected in the vehicle group suggesting that the low 116 concentrations detected could be produced endogenously from dopamine metabolism 117 [15]. It is remarkable, however, that at the highest dose (HT-100), HT-S, HV-Ac, 118 HVAc-S and HVAlc-S presented significant increases compared to the other 119 administered doses, indicating that only at higher doses some metabolites from the HT 120 intake could cross the blood-brain barrier. In fact, dose-dependent accumulation of HT-121 S and HVAlc-S was observed in the brain (p trend<0.001) ( hypothesized that the accumulation of these metabolites in the brain after a 125 pharmacological dose of HT could exert a neuro-protective activity in the central nerve 126 system. In the present study, the free form of HT was not detected in the brain, in 127 contrast to a previous study, in which 100 mg/kg of HT was administered through the 128 femoral vein [17] and considerable amounts of free HT were detected in the brain.

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Compared with the oral route of our study, intravenous administration could explain the 130 higher exposure of HT to the brain tissue and its detection in its free form.

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In contrast to other tissues, HT metabolites were not detected in the heart in HT-1 132 and HT-10 groups (   Summarizing, our study showed that HT and its metabolites could be accumulated 158 in a dose-dependent manner basically in the liver, kidney and brain and were detected in 159 these tissues even at nutritionally-relevant human doses, a dose that was not previously 160 studied in tissue disposition. The detection of free HT in liver and kidney is noteworthy.