Software implementing the methods for network generation presented in this paper is available for academic use by request from the authors in the form of compiled linux binary executables.

ABSTRACT:Human liver microsomes have typically resulted in marked underprediction of in vivo human intrinsic clearance (CL int ); therefore, the utility of cryopreserved hepatocytes as an alternative in vitro system has become an important issue. In this study, 10 compounds (tolbutamide, diclofenac, S-warfarin, S-mephenytoin, dextromethorphan, bufuralol, quinidine, nifedipine, testosterone, and terfenadine) were selected as substrate probes for CYP2C9, 2C19, 2D6, and 3A4, and the kinetics of metabolite formation (n ‫؍‬ 14 pathways) were investigated in three individual lots of cryopreserved hepatocytes and in a pool of human liver microsomes. For the majority of the compounds, lower unbound K M or S 50 values were observed in hepatocytes compared with microsomes, on average by 50% over a 200-fold range (0.5-140 M). Expressed on an equivalent liver weight basis, a good correlation between microsomal and hepatocyte V max values was observed for most pathways greater than 5 orders of magnitude (0.16-216 nmol/min/g liver). Unbound hepatocyte CL int (CL int,u ) values, when scaled to the whole liver (range 0.38-4000 ml/min/kg), were on average 2.5-fold higher than microsomal CL int,u values, with the exception of tolbutamide and diclofenac, for which lower hepatocellular CL int,u values were observed. Hepatocyte predicted CL int values were compared with human in vivo CL int values, and to supplement our data, in vitro data from cryopreserved hepatocytes were collated from four other published sources. These data show that for 37 drugs, there is, on average, a 4.5-fold under-prediction of the in vivo CL int using cryopreserved hepatocytes, representing a significant reduction in prediction bias compared with human microsomes.Human liver microsomes have traditionally been the most commonly used in vitro system for the prediction of metabolic clearance, in particular, for new chemical entities within drug discovery programs in the pharmaceutical industry. However, the use of this system has typically resulted in an underestimation of clearance, as illustrated by a data set of 55 compounds in which a 9-fold under-prediction of the in vivo intrinsic clearance (CL int ) was observed . As a result of this, attention in recent years has been placed on the use of alternative in vitro systems for clearance prediction. Fresh human hepatocytes are envisaged as a potentially more accurate system because of the full complement of both phase I and phase II metabolizing enzymes, along with the presence of transporter proteins, which should result in drug concentrations within the hepatocyte that are equivalent to in vivo concentrations within the liver. However, the limited availability of fresh human tissue and the cost implications involved in the preparation of freshly isolated human hepatocytes have resulted in cryopreserved hepatocytes emerging as the favored alternative, which also have the added advantage of being readily available commercially and more convenient to use (Li et al., 1999).Two major issues concerning the use ...

Zfp521 is a novel antiadipogenic transcription factor that helps to determine the identity of a mesenchymal cell as bone or fat.

An endophytic fungus, Gliocladium roseum (NRRL 50072), produced a series of volatile hydrocarbons and hydrocarbon derivatives on an oatmeal-based agar under microaerophilic conditions as analysed by solid-phase micro-extraction (SPME)-GC/MS. As an example, this organism produced an extensive series of the acetic acid esters of straight-chained alkanes including those of pentyl, hexyl, heptyl, octyl, sec-octyl and decyl alcohols. Other hydrocarbons were also produced by this organism, including undecane, 2,6-dimethyl; decane, 3,3,5-trimethyl; cyclohexene, 4-methyl; decane, 3,3,6-trimethyl; and undecane, 4,4-dimethyl. Volatile hydrocarbons were also produced on a cellulose-based medium, including heptane, octane, benzene, and some branched hydrocarbons. An extract of the host plant, Eucryphia cordifolia (ulmo), supported the growth and hydrocarbon production of this fungus. Quantification of volatile organic compounds, as measured by proton transfer mass spectrometry (PTR-MS), indicated a level of organic substances in the order of 80 p.p.m.v. (parts per million by volume) in the air space above the oatmeal agar medium in an 18 day old culture. Scaling the PTR-MS profile the acetic acid heptyl ester was quantified (at 500 p.p.b.v.) and subsequently the amount of each compound in the GC/MS profile could be estimated; all yielded a total value of about 4.0 p.p.m.v. The hydrocarbon profile of G. roseum contains a number of compounds normally associated with diesel fuel and so the volatiles of this fungus have been dubbed 'myco-diesel'. Extraction of liquid cultures of the fungus revealed the presence of numerous fatty acids and other lipids. All of these findings have implications in energy production and utilization.