Waleed Faisal, Caitriona O'Driscoll, Brendan Griffin1 Comparison of the intestinal lymphatic transport of lycopene in anaesthetised rat from a range of lipid based formulations. Waleed Faisal, Caitriona O'Driscoll, Brendan Griffin1 1Pharmaceutics, School of Pharmacy, University College Cork, Ireland w.faisal@mars.ucc.ie INTRODUCTION Mounting epidemiological evidence over the past decade suggests that a high intake of lycopene, the major carotenoid in tomatoes, may reduce the occurrence or progression of prostate cancer [1]. Absorption of lycopene from dietary food stuffs however is highly variable, appears to be affected by co-administered food stuffs and is non-dose dependent. The reasons for this variable absorption from the gastro-intestinal tract have not been elucidated. Lycopene is an extremely lipophilic carotenoid, and it is frequently hypothesized that lycopene is transported, to some extent, via the intestinal lymphatics. Lipid based formulations are known to enhance the oral bioavailability of highly lipophilic compounds, such as lycopene [2]. The current study proposes a systematic evaluation of the factors influencing uptake of lycopene from the gastro-intestinal tract using a lipid formulation approach. Aim: To assess the degree of intestinal lymphatic transport of lycopene in simulated fasted and fed state conditions in the rat. To elucidate the effect of bile salt mixed micelle formulations on the degree of intestinal lymphatic transport. To compare the compare the effect of long chain fatty acid (LCFA) versus medium chain fatty acid (MCFA) on intestinal lymphatic transport. Methods: The intestinal lymphatic transport of lycopene from a range of lipid based formulations were assessed in the mesenteric lymph duct cannulated anaesthetised rat model [2]. The lipid formulations administered were: fasted state stimulated intestinal fluid (FASSIF) and fed state stimulated intestinal fluid (FESSIF), oleic acid bile salt mixed micelles and decanoic acid bile salt mixed micelles. Lymph samples were collected, extracted and analysed by HPLC. Triglyceride levels in intestinal lymph were analysed by colorimetric assay. Solubility experiments were performed to determine saturation solubility of lycopene in simulated gastric fluids. RESULTS In an effort to explore further the mechanisms for enhanced lymphatic transport, a comparison of the effect of lipid formulations on intestinal triglyceride (TG) levels was performed (Table 2) Table 1 presents a comparison in terms of lipid composition and saturation solubility of lycopene for four lipid formulations used in the lymphatic transport experiments. Table 2: Cumulative transport of triglyceride into the mesenteric lymph (mean  S.E., n4) and cumulative lymph flow as a function of lipid formulation after 8 hours Formulation Composition Saturation Solubility of lycopene at 220C (µg/ml) Mean ± SE LCFA mixed micelles Oleic acid (C18) 1% Sodium cholate 2% 22.00±0.187 MCFA mixed micelles Decanoic acid (C12) 1% 13.12±0.706 FASSIF Sodium taurocholate 3mM Lecithin 0.75mM 8.29±0.682 FESSIF Sodium taurocholate 15mM Lecithin 3.75mM 10.97±0.898 Formulation Cumulative Mass of total TG (mg) in Mesenteric lymph (Mean  SE) Cumulative exogenous TG transport (mg) Cumulative lymph flow (ml) 0-8 hr Saline , Control 14.42.16 7.29 1.04 LCFA bile salt mixed micelles 52.624.66 38.21 9.31 1.18 MCFA bile salt mixed micelles 30.932.24 16.52 6.860.83 FASSIF 21.311.41 6.91 5.650.83 FESSIF 27.824.41 13.42 6.470.67 The extent of intestinal lymphatic transport following administration of FESSIF is 1.62% compared to 1.07% of the administered dose for FASSIF. The differences however was not statistically significant. A B Figure 1: Comparison of cumulative % of dose of lycopene transported in intestinal lymph after administration of FASSIF and FESSIF formulations in the rat. Figure 3: Correlation between lycopene transport rate (ng/hr) and TG transport rate (mg/hr) in rat lymph after administration of (A) bile salt LCFA and MCFA mixed micelle formulation and (B) FASSIF and FESSIF. The extent of intestinal lymphatic transport for the LCFA mixed micelle was 1.17% of the dose administered, which was significantly higher than that for the MCFA mixed micelles (0.42%). LCFA are transported primarily via the intestinal lymphatic transport pathway (i.e. chylomicron bound), whereas MCFA are generally considered to be more predominantly absorbed via the portal (blood) absorption route. Conclusion The extent of intestinal lymphatic transport of lycopene was consistently higher in fed state simulated intestinal fluids compared to fasted state intestinal fluids in the anaesthetised rat. The differences however were not statistically significant. LCFA bile salt mixed micelles promote the extent of intestinal lymphatic uptake of lycopene to a greater extent than MCFA bile salt mixed micelles. A strong positive correlation between intestinal lycopene levels and intestinal triglyceride levels was demonstrated for the bile salt mixed micelle formulations, which confirms that lycopene is transported in association with TG rich intestinal lipoproteins. The choice of formulation lipid can significantly affect the extent of intestinal lymphatic transport. The degree of correlation between lycopene and TG transport rate was lower for FASSIF and FESSIF formulations, which may reflect the influence of other mechanisms affecting lymphatic transport Reference 1. Giovannucci E. et al. Journal of the National Cancer Institute (Bethesda) 94: 391-398, 2002. 2. Griffin BT and O'Driscoll CM. Journal of Pharmacy and Pharmacology 58: 917-925, 2006. Figure 2: Comparison of cumulative % dose of lycopene transported in rat lymph after administration of LCFA and MCFA bile salt mixed micelle formulations.