The neonatal rat cardiomyocyte model enables heart researchers to study and understand the morphological, biochemical and electrophysiological characteristics of the heart. This model offers a broad spectrum of experiments, such as studies of contraction, ischaemia, hypoxia and the toxicity of various compounds. This review examines the methodology for the isolation and cultivation of primary neonatal rat cardiomyocyte cultures, considers possible problems and pitfalls, and presents an optimized protocol for cardiomyocyte preparation.

Liquid chromatography/electrospray ionization mass and tandem mass spectrometry (MS/MS) techniques were used to identify two minor components and one new compound in the polyphenolic extract of green tea (Camellia sinensis). Identification and structure assignments were based on previously reported sub-structural features in the MS/MS product, precursor and neutral loss scans of reference samples. The structures of two minor components, related to the known green tea components epicatechin gallate (ECG, 5) and epigallocatechin gallate (EGCG, 6), are formed by methylation at the 3"-O-position of the gallic acid moiety. The new compound contained a gallic acid ester group, but had only one phenolic group in either the A- or B-ring, relative to the structure of 5. High-resolution mass measurements supported the empirical formula assigned to the new compounds. An important fragmentation for defining the position of methylation of the ester function involves ionization of the phenolic group at the 4"-position of the gallic acid, followed by elimination of the ester function as a neutral with concomitant formation of the m/z 169 ion. If the 4"-position is blocked by methylation, the formation of m/z 169 incorporating the gallic acid group would be blocked. Thus, the presence of an ion representing the ester group indicates a free 4"-phenol and the absence of this ion would signify the 4"-position as a site of methylation. The operation of this mechanism should be general and useful in assigning the site of methylation of any polyphenolic ester group in natural products. A similar conclusion can be drawn concerning alkylation or esterification of the 4'-position of the catechins, i. e. blocking the 4'-phenol would prevent formation of the m/z 125 ion common to all of the catechin compounds thus far examined. Therefore, mass spectral data are of considerable importance in deducing the sites of alkylation or esterification in the structures of the components of green tea.

Mass spectrometric methods including EIMS, FABMS, and LC/ESIMS have been surveyed as tools for the detection of catechins in extracts of green tea (Camellia sinensis). EIMS provide both molecular weight and structure information, including epimer differentiation, on compounds 1, 2, and 4 and some structural information with compounds 5 and 6. FABMS gives both molecular weight and structure information, by an retro-Diels-Alder mechanism, on all compounds. LC/ESIMS provides unambiguous molecular weight information on all compounds and some additional structural data are evident in the mass spectra of 5 and 6. LC/ESIMS is, thus, shown to be an appropriate method for the direct analysis of crude extracts of green tea. The information obtained in this work will be of importance in future studies involving identification of the components of green tea and for characterization of synthetic analogues.

Central nervous system (CNS) treatment contributes to improved long-term disease-free survival from childhood acute lymphoblastic leukemia (ALL) by significantly decreasing the rate of disease relapse. Methotrexate (MTX), a drug commonly used for CNS treatment, has been associated with cognitive and academic problems, white-matter changes, perfusion defects, and brain atrophy. This study investigated oxidative stress as a possible mechanism of chemotherapy induced CNS injury. Unoxidized and oxidized components of phosphatidylcholine (PC), the most prevalent phospholipid in CNS cellular membranes, were measured in cerebral spinal fluid (CSF) samples obtained from 21 children diagnosed with low (n = 7), standard (n= 7), or high (n= 7) risk ALL. Children with high-risk ALL received the most MTX, especially during the most intensive phase of treatment (consolidation). Phospholipids were extracted from CSF samples obtained at diagnosis and during the induction, consolidation, and continuation treatment phases. Unoxidized and oxidized PC were measured by normal phase high-performance liquid chromatography at 2 ultraviolet wavelengths (206 and 234 nm, respectively). Data were analyzed by 2-way repeated-measures analysis of variance. Results support the hypotheses that the highest levels of oxidized PC would be observed during the most intensive phase of ALL therapy and in the high-risk ALL group. Findings provide preliminary evidence for chemotherapy-induced oxidative stress in CNS membrane phospholipids.