Genetic drift in animal populations has been a recognized concern for many years. Less understood is the potential for phenotypic "drift" or variation that is not related to any genetic change. Recently, stock Sprague-Dawley (Crl:CD(SD)) rats obtained from the Charles River Raleigh facility demonstrated a distinct endogenous urinary metabonomic profile that differed from historical control SD urine spectral profiles obtained over the past several years in our laboratory. In follow-up studies, the origin of the variant phenotype was narrowed down to animals of both sexes that were housed in one specific room (Room 9) in the Raleigh facility. It is likely that the two phenotypes are related to distinct populations of gut flora that particularly impact the metabolism of aromatic molecules. The most pronounced difference between the two phenotypes is the relative amounts of hippuric acid versus other aromatic acid metabolites of chlorogenic acid. Though both molecular species are present in either phenotype, the marked variation in levels of these molecules between the two phenotypes has led to the designation of high hippuric acid (HIP) and high chlorogenic acid metabolites (CA) phenotypes. Specific urinary components that distinguish the phenotypes have been thoroughly characterized by NMR spectroscopy with additional, limited characterization by LC-MS (high performance liquid chromatography coupled with mass spectrometry). Co-habitation of rats from the two phenotypes rapidly facilitated a switch of the CA phenotype to the historical Sprague-Dawley phenotype (HIP). The impact of these variant phenotypes on drug metabolism and long-term safety assessment studies (e.g., carcinogenicity bioassays) is unknown.

A NMR spectroscopic method is described that enables the quantitation of specific lipid classes and components, independent of fatty acid composition. We demonstrate this method for measuring cholesterol, squalene, and pools of sterol esters, wax esters (WEs), and triglyceride (TG) components in sebum and meibum. When 600 MHz NMR equipment is used in conjunction with highly sensitive cryogenically cooled probes, this method has adequate sensitivity, and for some applications, advantages over commonly used HPLC-evaporative light-scattering detection and mass spectrometry-based approaches. This method is shown to be useful for preclinical and clinical monitoring of the efficacy of sebum-reducing agents in animals and humans. In Syrian hamsters, 3% topical flutamide and 20 mg/kg oral isotretinoin reduced sterol esters by 18.7% and 30.0%, respectively, and reduced WEs by 32.9% and 31.8%, respectively, as measured in a punch biopsy of the ear. In a 72 patient clinical methodology study, the assay delivered reproducible and noninvasive measurements of WEs, cholesteryl esters, TGs, and squalene from Sebutape: skin blots. The quantitative results of sebum analysis obtained by the NMR method correlate well with those obtained with HPLC-based approaches. This approach may be broadly applicable to cases in which fatty acid-independent quantification of lipid classes is desired. Mammalian skin is composed of three primary layers: the stratum corneum, the epidermis, and the dermis. The outer layer of the skin, the stratum corneum, primarily functions as a barrier to the external environment, preventing water loss and the invasion of microorganisms. Sebum secreted to the stratum corneum from the sebaceous glands is a key component of the skin surface and has various demonstrated and postulated functions (1). Sebum itself is a complex mixture of lipids and is produced by the sebaceous glands. At maturation, the acinar cells of the sebaceous glands lyse and release sebum into the lumenal duct, from which the sebum is secreted. Cholesterol, sterol esters, wax esters (WEs), and triglycerides (TGs) are the primary lipids found in the sebum of many mammals, and squalene is also a major component of human sebum. In humans, the predominant sterol esters are cholesteryl esters (CEs), whereas in male Syrian hamster (a common sebum model), they are chiefly fatty acid esters of a single predominant noncholesterol sterol (2). The identity of the sterol constituting these esters in this species has not been identified, and herein we will refer to these as hamster sterol esters (HSEs). WEs are unique to sebum in that they are not synthesized by other cells in the body. Sebum composition can be modified from its native state by the action of xenobiots, which can greatly increase the complexity of its biochemical makeup; for example, bacterial hydrolytic enzymes are known to break down TGs to produce free fatty acids (3).A number of methods to isolate and qualitatively and/ or quantitatively analyze the lipid components of sebum and/...

Pulsed-field-gradient NMR methods are used for determining the partitioning of surfactants between unimeric and micellar forms in mixed surfactant systems. The method allows determination of the composition dependence of activity coefficients in binary surfactant mixtures. Application of the regular solution approach to NMR data for several mixtures gives β parameters which are dependent on concentration and which differ systematically among the surfactants within the same binary mixtures. The β parameter summarizes interactions among different pairs of surfactants and, in principle, should not be composition dependent. The observed behavior is therefore not consistent with expectations from the regular solution model. Use of the van Laar expressions, on the other hand, accounts well for the composition dependence of the activity coefficients. The van Laar expressions also account for the often-observed composition dependence of regular solution β parameters determined from CMC measurements. Though similar to the symmetric regular solution model, the van Laar expressions contain an additional parameter which reflects differences in the sizes of the mixture components. The results therefore suggest that headgroup size and headgroup packing are important contributors to nonideal surfactant behavior. Computer algorithms are described for extracting the van Laar interaction energy-and size-related parameters from NMR-derived results, from mixed critical micelle concentrations, or from heats of mixing. Data for several binary surfactant mixtures are presented and discussed. The results emphasize that when accurate data are available, the single-parameter regular solution model will not always fully account for nonideal surfactant mixing.

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Metabonomics is an emerging technology that enables rapid in vivo screening for toxicity, disease state, or drug efficacy. The technology combines the power of high-resolution nuclear magnetic resonance (NMR) techniques with statistical data analysis methods to rapidly evaluate the metabolic "status" of an animal. Complimentary to other profiling technologies like proteomics and genomics, metabonomics provides a fingerprint of the small-molecules contained in a given biofluid through the time course of a study. This article reviews the steps in implementing a metabonomics-based screening program from study design through data analysis. While metabonomics is still a relatively new technology in comparison to the other "omics", published results from metabonomics studies demonstrate its potential impact in the drug discovery process by enabling the incorporation of safety endpoints much earlier in the drug discovery process, reducing the likelihood (and cost) of later stage attrition.