Supercritical fluid chromatographic analysis for on-line monitoring of hexane removal from soybean oil miscella using liquid carbon dioxide.

Liquid carbon dioxide (L-CO2) can be used to separate hexane from hexane/soybean oil (SBO) mixtures (i.e., miscella). An on-line supercritical fluid chromatographic (SFC) method was developed to monitor this separation. L-CO2 (25 degrees C and 9.31 MPa) was passed through 50 mL of a 25% (w/w) hexane miscella and then directed on-line through a SFC injector. After passing 300-L expanded CO2, the hexane concentrations in the L-CO2 were 0.05% and 0.04% for n-hexane and isohexane, respectively and the residual hexane concentrations in the SBO were 3.8 and 3.3 ppm, respectively. This technique provided real time on-line monitoring of the hexane separation process.

Pressurized fluids for extraction of cedarwood oil from Juniperus virginianna.

The extraction of cedarwood oil (CWO) using liquid carbon dioxide (LC-CO(2)) was investigated and compared to supercritical fluid extraction, including the effects of extraction pressure and length of extraction. The chemical composition of the extracts was monitored over the course of the extraction as well. The cumulative yields of CWO from cedarwood chips using 80 L of carbon dioxide varied very little treatment to treatment, with all temperature/pressure combinations yielding between 3.55 and 3.88% CWO, and the cumulative yields were statistically equivalent. The rate of extraction was highest under the supercritical extraction conditions (i.e., 100 degrees C and 6000 psi). Under the liquid CO(2) conditions (i.e., 25 degrees C), the extraction rates did not vary significantly with extraction pressure. However, there were differences in the chemical composition of the collected CWO. Extractions at 100 degrees C gave a much lower ratio of cedrol/cedrene than extractions at 25 degrees C. The highest ratio of cedrol/cedrene was obtained using 25 degrees C and 1500 psi. The use of subcritical water was also investigated for the extraction of CWO as well. Although some CWO was extracted using this method, the temperature/pressure combinations that gave the highest weight percentage yields also gave oils with an off odor while those combinations that gave a higher quality oil had very low yields. It appears that the high temperatures and acidic conditions cause a dehydration of the tertiary alcohol, cedrol, to its hydrocarbon analogue, cedrene, during CO(2) or pressurized water extractions of cedarwood.

Supercritical Fluid Extraction of Aflatoxin M1 from Beef Livert †.

Supercritical fluid extraction (SFE) and a pressurized-fluid-extraction process were applied for the removal of aflatoxin M1 from beef liver samples. Various pressures, temperatures, quantity of supercritical carbon dioxide, and organic modifiers were investigated to optimize the extraction methods. Organic modifier was found to be essential for quantitative recovery of aflatoxin M1. Extracts were cleaned up by solid-phase extraction and were analyzed via high-performance liquid chromatography coupled with fluorescence detection of the trifluoroacetic acid derivative. Solvent-modified carbon dioxide SFE achieved recoveries comparable to an AOAC-approved method involving organic solvent extraction. SFE allowed the traditional amounts of sample and organic solvent to be reduced. Also, the supercritical-fluid extraction permitted the use of carbon dioxide modified with acetonitrile: methanol (2:1) to replace methylene chloride as the organic solvent for the extraction step.

Evolutionarily conserved sequence elements that positively regulate IFN-gamma expression in T cells.

Our understanding of mechanisms by which the expression of IFN-gamma is regulated is limited. Herein, we identify two evolutionarily conserved noncoding sequence elements (IFNgCNS1 and IFNg CNS2) located approximately 5 kb upstream and approximately 18 kb downstream of the initiation codon of the murine Ifng gene. When linked to the murine Ifng gene (-3.4 to +5.6 kb) and transiently transfected into EL-4 cells, these elements clearly enhanced IFN-gamma expression in response to ionomycin and phorbol 12-myristate 13-acetate and weakly enhanced expression in response to T-bet. A DNase I hypersensitive site and extragenic transcripts at IFNgCNS2 correlated positively with the capacity of primary T cell subsets to produce IFN-gamma. Transcriptionally favorable histone modifications in the Ifng promoter, intronic regions, IFNgCNS2, and, although less pronounced, IFNgCNS1 increased as naïve T cells differentiated into IFN-gamma-producing effector CD8+ and T helper (TH) 1 T cells, but not into TH2 T cells. Like IFN-gamma expression, these histone modifications were T-bet-dependent in CD4+ cells, but not CD8+ T cells. These findings define two distal regulatory elements associated with T cell subset-specific IFN-gamma expression.

Genomic organization of the interleukin-1 locus.

Recent additions have expanded the interleukin (IL)-1 gene family to 10 members. We have determined the order, orientation, and intergenic distance of the nine IL-1 family genes that lie on human chromosome 2. We report cDNA sequences for the mouse orthologs of three of these genes. The order and orientation of the mouse genes have been mapped, and the mouse locus compared with the human locus. There is a break in the mouse locus of > 100 kb, compared with the human locus, located between Il1b and the most centromere-proximal of the novel mouse genes. The mouse seems to be missing an ortholog of human IL1F7.

Sequence polymorphism at the human apolipoprotein AII gene ( APOA2): unexpected deficit of variation in an African-American sample.

A 3.3-kb region, encompassing the APOA2 gene and 2 kb of 5' and 3' flanking DNA, was re-sequenced in a "core" sample of 24 individuals, sampled without regard to the health from each of three populations: African-Americans from Jackson (Miss., USA), Europeans from North Karelia (Finland), and non-Hispanic European-Americans from Rochester, (Minn., USA). Fifteen variable sites were identified (14 SNPs and one multi-allelic microsatellite, all silent), and these sites segregated as 18 sequence haplotypes (or nine, if SNPs only are considered). The haplotype distribution in the core African-American sample was unusual, with a deficit of particular haplotypes compared with those found in the other two samples, and a significantly (P<0.05) low level of nucleotide diversity relative to patterns of polymorphism and divergence at other human loci. Six of the 14 SNPs, whose variation captured the haplotype structure of the core data, were then genotyped by oligonucleotide ligation assay in an additional 2183 individuals from the same three populations (n=843, n=452, and n=888, respectively). All six sites varied in each of the larger "epidemiological" samples, and together, they defined 19 SNP haplotypes, seven with relative frequencies greater than 1% in the total sample; all of these common haplotypes had been identified earlier in the core re-sequencing survey. Here also, the African-American sample showed significantly lower SNP heterozygosity and haplotype diversity than the other two samples. The deficit of polymorphism is consistent with a population-specific non-neutral increase in the relative frequency of several haplotypes in Jackson.

The effects of scale: variation in the APOA1/C3/A4/A5 gene cluster.

While there is considerable appeal to the idea of selecting a few SNPs to represent all, or much, of the DNA sequence variability in a local chromosomal region, it is also important to quantify what detail is lost in adopting such an approach. To address this issue, we compared high- and low-resolution depictions of sequence diversity for the same genomic region, the APOA1/C3/A4/A5 gene cluster on chromosome 11. First, extensive re-sequencing identified all nucleotide and sequence haplotype variation of the linked apolipoprotein genes in 72 individuals from three populations: African-Americans from Jackson, Miss., Europeans from North Karelia, Finland, and European-Americans from Rochester, Minn. We identified 124 SNPs in 17.7 kb and significant differences in variation among genes. APOC3 gene diversity was particularly distinctive at high resolution, showing large allele frequency differences ( F(ST) values >0.250) between Jackson and the other two samples, and divergent population-specific haplotype lineages. Next, we selected haplotype-tagging SNPs (htSNPs) for each gene, at a density of approximately one SNP per kb, using an algorithm suggested by Stram et al. (2003). The 17 htSNPs identified were then used to reconstruct low-resolution haplotypes, from which inferences about the structure of variation were also drawn. This comparison showed that while the htSNPs successfully tagged common haplotype variation, they also left much underlying sequence diversity undetected and failed, in some cases, to co-classify groups of closely related haplotypes. The implications of these findings for other haplotype-based descriptions of human variation are discussed.