The mammalian lignan, enterolactone (EL), is a gut microbe metabolite of plant lignan secoisolariciresinol diglucoside (SDG), which is most abundant in flaxseed. Numerous epidemiological, experimental and clinical studies suggest the protective effects of EL against various chronic diseases such as cancer, cardiovascular disease, and inflammation. However, EL's oral bioavailability is low and highly variable due to extensive first-pass metabolism, especially glucuronidation, which results in the large amount of glucuronide metabolites but low levels of free EL in human plasma. Hepatocytes and enterocytes express UDP-glucuronosyltransferases (UGT), the enzymes responsible for the conjugation of glucuronic acid to EL. To better understand the contribution of liver and intestine to the first-pass glucuronidation, I conducted an in vitro enzyme kinetic analysis of EL glucuronidation using hepatic and intestinal microsomal fractions from both human and rat.An intrinsic clearance (CL int ) value was calculated using the substrate depletion approach. In addition to monitoring substrate depletion, high-pressure liquid chromatography (HPLC) analysis allowed detection of EL glucuronides, which were further substantiated by LC-MS.EL monoglucuronide was identified in rat and human intestinal and liver microsomes.Enzyme kinetic studies indicated the extent of hepatic microsomal glucuronidation exceeded intestinal glucuronidation in both human and rat, while the human liver CL int value was slightly higher than that of rat liver. The CL int value generated in human intestinal microsomes was only one third of the value of human liver, whereas, the CL int of rat jejunum or colon was one-twentieth of rat liver, suggesting the human intestine makes a greater contribution to EL glucuronidation than rat intestine. These results suggest that both liver and intestine contribute to EL glucuronidation and the human intestine may exert a greater influence on the first-pass glucuronidation of EL than rat intestine, thereby significantly decreasing EL's oral bioavailability. The rat might underestimate the extent of intestinal metabolism of EL relative to human.iii

Introduction t-Butylamine is a versatile and important building block for compounds with biological activities. Finasteride, a 5a-reductase inhibitor for use in treating acne, female hirsutism and benign prostatic hyperplasia, was prepared by the reaction of the carboxylate of 4-aza-5a-androst -1-ene-3-one with t-butylamine. 1It was also built into an apoptosis inhibitor, 2 an HIV protease inhibitor, 3 various antibotics 4 and nucleosides. 5 Moreover t-butylamine residues are commonly in adrenergic drugs to increase affinity for b-adrenoceptors. 6-8Here we describe an efficient synthesis of [15 N]t-butylamine hydrochloride, which can be used as biomarker for biologically active compounds containing this motif or incorporated in the synthesis of drug candidates for use as analytical internal standards or for metabolic studies. H-NMR (400 MHz) and 13 C-NMR (100 MHz) spectra were recorded on a JNM-ECA-400 NMR spectrometer in CDCl 3 or DMSO-d 6 (TMS as internal standard). FT-IR spectra were recorded on a Nicolet FT-IR 5700 spectrometer using KBr pellets. EI-MS Spectra were obtained with ZAB-HS spectrometer. : 3390, 3199, 2961, 2923, 2756, 1648, 1485, 1455, 1408, 1376, 1356, 1221, 1103, 863, 816, 732, 615 45 mmol) in DMF (8 mL), followed by the addition of a solution of NBS (0.48 g, 2.7 mmol) in DMF (2 mL) at room temperature under nitrogen. After reaction at this temperature for 30 h, the reaction mixture was poured into water and extracted with CH 2 Cl 2 (15 mL Â 3). The combined extracts were washed with water, saturated NaHCO 3 , dried over anhydrous magnesium sulfate and evaporated. The crude product was purified by flash chromatography (petroleum ether: ethyl 3341, 2969, 1713, 1498, 1454, 1394, 1365, 1266, 1210, 1072, 914, 776, 738, 696 Results and discussion