Influences on breastfeeding rates in low income communities in Ontario.

The purpose of this paper was to identify factors associated with breastfeeding among mothers of children born in 1994 in five of the low income communities participating in the longitudinal prevention initiative "Better Beginnings, Better Futures." Household income was < or = poverty line for 76%, 63% had completed high school or beyond, and 29% were born outside of Canada. The breastfeeding initiation rate was 77% (380 of 493). Of the 270 women who initiated breastfeeding and were interviewed up to five months postpartum, 63% continued for at least three months. Women with higher education, married, not experiencing financial stress and who attended prenatal classes were more likely to initiate breastfeeding. Continuation of breastfeeding was associated with older age, higher education, not smoking, and participation in a home visitor program. Breastfeeding promotion strategies should include ongoing support as well as education components.

Generation and analysis of 280,000 human expressed sequence tags.

We report the generation of 319,311 single-pass sequencing reactions (known as expressed sequence tags, or ESTs) obtained from the 5' and 3' ends of 194,031 human cDNA clones. Our goal has been to obtain tag sequences from many different genes and to deposit these in the publicly accessible Data Base for Expressed Sequence Tags. Highly efficient automatic screening of the data allows deposition of the annotated sequences without delay. Sequences have been generated from 26 oligo(dT) primed directionally cloned libraries, of which 18 were normalized. The libraries were constructed using mRNA isolated from 17 different tissues representing three developmental states. Comparisons of a subset of our data with nonredundant human mRNA and protein data bases show that the ESTs represent many known sequences and contain many that are novel. Analysis of protein families using Hidden Markov Models confirms this observation and supports the contention that although normalization reduces significantly the relative abundance of redundant cDNA clones, it does not result in the complete removal of members of gene families.

Changes in glutathione content and resistance to anticancer agents in human stomach cancer cells induced by treatments with melphalan in vitro.

A clone of a human gastric carcinoma cell line was used to determine whether cells which had survived a treatment with Melphalan would express altered survival responses when treated again with this agent 1 week or more later. Cells were treated for 1 h each week with 2 micrograms/ml (99% lethal dose). After the first Melphalan treatment, the cells exhibited a 10-fold reduction in sensitivity to Melphalan. This was preceded by a 2-fold increase in intracellular glutathione content. By the end of 10 weekly treatments, the cells were 50 times more resistant than controls (based on changes in survival fractions). They also demonstrated collateral resistance to Actinomycin D, 1-(2-chloroethyl)-3-(4-methylcyclohexyl)-1-nitrosourea, galactitol, and X-rays, but showed no change in sensitivity to 5-fluorouracil, bleomycin, and Adriamycin. The resistance to Melphalan was not reversible when treatment was withheld for 4 weeks on two different occasions. The results suggest that treatment with a high dose of Melphalan either selects an existing population of cells with a high GSH content or induces mutations leading to increased GSH content or both, and this results in the expression of greater Melphalan resistance at the time of other treatments. Furthermore, Melphalan treatment stimulates a 50% increase in GSH content in resistant cells in just 6 h, an 85% increase in 36 h, and a 150% increase in 72 h. L-Buthionine sulfoximine partially reversed the expression of resistance to Melphalan by inducing a 60% reduction in intracellular glutathione content.

Experience with routine applications of liquid chromatography-mass spectrometry in the pharmaceutical industry.

The combination of liquid chromatography and mass spectrometry (LC-MS) has been established to complement gas chromatography (GC)-MS in the analysis of non-volatile and labile drugs in complex materials. The possibilities of LC-MS in the pharmaceutical industry for the analysis of drug substances and dosage forms, metabolism studies and the elucidation of the structures of materials of biological origin are discussed. Instrumental requirements, limitations and applications of LC-MS are considered and experiences with LC-MS in routine applications are reported. Preliminary results obtained with thermospray LC-MS are compared with those using a direct liquid inlet interface.

Radioprotective effect of polyethylene glycol.

Polyethylene glycol of molecular weight 400 (PEG-400) had a radioprotective effect of about 20% against lethality when given ip 20 min prior to single or fractionated X-ray doses to the head and neck. Dose modification factors (DMF) based on LD50/15 values ranged from 1.14 to 1.24. A similar DMF of 1.12 based on LD50/30 values was obtained using single doses of whole-body X irradiation. Mice given head and neck irradiation had significantly reduced rectal temperatures (31.3 +/- 3.0 degrees C) 9 days post irradiation compared with unirradiated controls (35.4 +/- 0.6 degrees C). No such reduction was observed when PEG-400 was given with radiation (36.3 +/- 0.9 degrees C). PEG-400 also lessened, but not significantly, the frequency of shivering in irradiated animals. Histopathologic examination of the oral structures demonstrated only marginal protection by PEG-400. Estimation of the alpha/beta ratio from LD50 data on head and neck-irradiated mice yielded values of 4.4 +/- 1.9 (95% confidence limits) Gy without PEG-400 and 7.9 +/- 1.4 Gy with PEG-400. Since it is a non-thiol radioprotector, PEG-400 may be more useful when combined with more conventional thiol-containing radioprotectors.

Formation of methyl ester of 2-methylglyceric acid from thymine glycol residues: a convenient new method for determining radiation damage to DNA.

Thymine glycol residues in DNA or thymidine were converted to methyl 2-methylglycerate by reaction with alkaline borohydride followed by methanolic HCl. The product was labeled either from [3H]DNA or from [3H]borohydride and was followed by cochromatography with authentic 14C-labeled material. Following acid hydrolysis, the identity of 2-methylglyceric acid was confirmed by high-resolution mass spectrometry, NMR, IR, and elemental analysis. Treatment of DNA or thymidine with X-irradiation, with H2O2 and Fe2+, with H2O2, Cu2+, and ascorbate, and with H2O2 and ultraviolet light, permanganate, or sonication all produced methyl 2-methylglycerate in varying amounts after alkaline borohydride and methanolic HCl, whereas untreated DNA did not. The data indicate that certain oxidants including hydroxyl radicals generated by chemical means or from radiolysis of water convert thymine residues to thymine glycols in DNA, which can be determined as methyl 2-methylglycerate.

Polyethylene glycol as a protector against head and neck irradiation.

Mice injected i.p. with polyethylene glycols (PEG) 20 min prior to head and neck X irradiation with 1650 rad showed improved survival, increased food and water consumption, and retention of body weight compared with irradiated controls. The LD 50/15 for PEG-treated mice was 1900 +/- 108 rad compared to 1527 +/- 56 for the controls. PEG of molecular weights 200, 400, and 600 afforded significant levels of radioprotection; PEG of molecular weights 1000, 1450, 4000, and 20,000 when given at maximum tolerated doses (approximately 0.5 LD 50) did not. The degree of radioprotection by PEG with molecular 400 given 20 min before irradiation increased with dose up to the maximum tolerated dose of 6.4 g/Kg. Significant, but lower, levels of radioprotection were observed when the PEG was given 5 min after irradiation. Mice injected i.p. with PEG, cystamine, 5-thioglucose, chlorpromazine, polyethylene glycol monomethyl ether or polyvinylpyrrolidone all had comparable survival levels. Polypropylene glycol, polyethylene glycol diacrylate, and polycaprolactonediol were more toxic than PEG and showed no radioprotection.

Mobile-phase and temperature effects in the reversed phase chromatographic separation of proteins.

This paper explores the influence of mobile-phase and temperature effects on the gradient elution reversed phase chromatographic behavior of proteins. Using LiChrospher SI 500, bonded with n-butyl groups and a gradient in 1-propanol, 10 mM H3PO4, rapid separation and high mass recovery were obtained for a series of globular proteins. This protein separation and recovery are compared to those obtained when acetonitrile and acetonitrile plus 10 mM H3PO4 are used as eluting gradient solvents. In general, acetonitrile yielded lower recovery than 1-propanol, particularly for the more hydrophobic proteins, e.g., ovalbumin. For all three gradient solvents, little difference was observed in retention or recovery when the n-alkyl chain of the bonded phase varied. On the other hand, relative to the n-alkyl phases, a significantly lower retention of all proteins was found on more hydrophilic phases, e.g., cyano and nonendcapped n-butyl, when acetonitrile was the organic modifier, while in the case of 1-propanol, no retention difference was observed. Thus, column comparisons depend on the protein/mobile-phase combinations examined. The role of column temperature was also studied, and it was found that for certain proteins dramatic changes in peak shape occurred as a function of temperature. The influences of ionic strength and salt type were also studied. Protein mass recovery was shown to decrease with an increase in salt concentration; moreover, perchlorate was shown to have a larger effect in this regard than phosphate. In addition, salt concentration and type were found to influence peak shape greatly in certain cases. The results indicate the strong influences of mobile phase and temperature on chromatographic behavior, and some of the options available when this behavior is not satisfactory. Several protein separations are presented illustrating the power of the reversed phase approach.

Characterization of radiation damage to DNA by reaction with borohydride.

Irradiation of aqueous solutions of native calf thymus DNA with x-rays produced functional groups that reacted with sodium borohydride. The DNA was labeled with tritium from NaB3H4 to the extent of 2.0 x 10(-10) atom/dalton/rad. The presence of cysteamine or other radical scavengers, or saturation of the solution with nitrogen during irradiation decreased the labeling. After mild acid hydrolysis, the major tritium-containing moiety was identical with 2,3-dihydroxy-2-methylpropanoic acid in all chromatographic systems tested. The suggested mechanism of labeling involved reduction by borohydride of the potential aldehyde at carbon 6 of thymine glycol residues present in the irradiated DNA.