Inlet hydrogenation gas chromatography to predict mass % linear paraffin content.

The light condensate fraction obtained from the low temperature Fischer-Tropsch (LT-FT) process is very complex and it is processed further by hydrotreating to produce hydrocarbon products that can be sold as final products. The mass% linear paraffins in some of the final paraffin products is listed as a required specification. Usually gas chromatography equipped with a flame ionisation detector (GC-FID) is used for the analysis of the condensate feeds to estimate the mass% linear paraffins that can be expected in the final products after commercial hydrogenation. This is an important parameter used in the blending of suitable condensate feeds. Due to the complexity of the condensate feeds, significant peak overlap occurs in the GC-FID analysis, making it difficult to accurately estimate the mass% linear paraffin content that will be obtained in the hydrogenated products. Inlet hydrogenation GC-FID analysis simplifies the prediction of the mass% linear content that can be expected in the paraffin product fractions from the analysis of a plant feed since the feed is hydrogenated in the GC inlet before GC-FID analysis. The results from this study showed that sufficient hydrogenation without significant peak tailing can be obtained in the GC inlet when using the appropriate mass and particle size Pd/Al2O3 catalyst with the optimum bed height. Inlet hydrogenation GC-FID analysis simplifies the prediction of the mass% linear content that can be expected in the paraffin product fractions. The method can be implemented on routine GC-FID instrumentation by simply installing an inlet liner containing an appropriate catalyst, that could be re-used at least 20 times, and avoids the purchasing of additional instrumentation and complex data processing and is suitable for commercial process control.

Analysis of oxidised heavy paraffininc products by high temperature comprehensive two-dimensional gas chromatography.

Heavy petroleum fractions are produced during crude and synthetic crude oil refining processes and they need to be upgraded to useable products to increase their market value. Usually these fractions are upgraded to fuel products by hydrocracking, hydroisomerization and hydrogenation processes. These fractions are also upgraded to other high value commercial products like lubricant oils and waxes by distillation, hydrogenation, and oxidation and/or blending. Oxidation of hydrogenated heavy paraffinic fractions produces high value products that contain a variety of oxygenates and the characterization of these heavy oxygenates is very important for the control of oxidation processes. Traditionally titrimetric procedures are used to monitor oxygenate formation, however, these titrimetric procedures are tedious and lack selectivity toward specific oxygenate classes in complex matrices. Comprehensive two-dimensional gas chromatography (GC×GC) is a way of increasing peak capacity for the comprehensive analysis of complex samples. Other groups have used HT-GC×GC to extend the carbon number range attainable by GC×GC and have optimised HT-GC×GC parameters for the separation of aromatics, nitrogen-containing compounds as well as sulphur-containing compounds in heavy petroleum fractions. HT-GC×GC column combinations for the separation of oxygenates in oxidised heavy paraffinic fractions are optimised in this study. The advantages of the HT-GC×GC method in the monitoring of the oxidation reactions of heavy paraffinic fraction samples are illustrated.

Two-dimensional gas chromatography-online hydrogenation for improved characterization of petrochemical samples.

The Fischer-Tropsch (FT) process produces a variety of hydrocarbons over a wide carbon number range and during subsequent product workup a large variety of synthetic fuels and chemicals are produced. The complexity of the product slate obtained from this process is well documented and the high temperature FT (HT-FT) process products are spread over gas, oil and water phases. The characterization of these phases is very challenging even when using comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC×GC-TOFMS). Despite the increase in separation power, peak co-elution still occurs when samples containing isomeric compounds are analysed by comprehensive two dimensional GC. The separation of isomeric compounds with the same double bond equivalents is especially difficult since these compounds elute in a similar position on the GC×GC chromatogram and have identical molecular masses and similar fragmentation patterns in their electron ionization (EI) mass spectra. On-line hydrogenation after GC×GC separation is a possible way to distinguish between these isomeric compounds since the number of rings and alkene double bonds can be determined from the mass spectra of the compounds before and after hydrogenation. This paper describes development of a GC×GC method with post column hydrogenation for the determination of the backbone of cyclic/olefinic structures enabling us to differentiate between classes like dienes and cyclic olefins in complex petrochemical streams.

Hyphenation of supercritical fluid chromatography and two-dimensional gas chromatography-mass spectrometry for group type separations.

The Fischer-Tropsch (FT) process produces a variety of compounds over a wide carbon number range and the synthetic crude oil produced by this process is rich in highly valuable olefins and oxygenates, which crude oil only contains at trace levels. The characterization of these products is very challenging even when using comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GC×GC-TOF-MS). The separation between cyclic paraffins and olefins is especially difficult since they elute in similar positions on the GC×GC chromatogram and since they have identical molecular masses with indistinguishable fragmentation patterns. Previously, a high performance liquid chromatography (HPLC) fractionation procedure was used prior to GC×GC-TOF-MS analysis to distinguish between alkenes and alkanes, both cyclic and non-cyclic, however, there was co-elution of the solvents used in the HPLC fractionation procedure, and the volatile components in the gasoline sample and the dilution introduced by the off-line fractionation procedure made it very difficult to investigate components present at very low concentrations. The hyphenation of supercritical fluid chromatography (SFC) to GC×GC is less complicated and the removal of the supercritical CO2 can be easily achieved without any loss of the volatile sample components, eliminating the introduction of co-eluting solvents as well as the dilution effect. This paper describes the on-line hyphenation of SFC to a GC×GC system in order to comprehensively characterize the chemical groups (saturates, unsaturates, oxygenates and aromatics) in an FT sample.

Total-body digital X-ray in trauma. An experience report on the first operational full body scanner in Europe and its possible role in ATLS.

When patients enter our emergency room with suspected multiple injuries, Statscan provides a full body anterior and lateral image for initial diagnosis, and then zooms in on specific smaller areas for a more detailed evaluation. In order to examine the possible role of Statscan in the management of multiply injured patients we implemented a modified ATLS((R)) algorithm, where X-ray of C-spine, chest and pelvis have been replaced by single-total a.p./lat. body radiograph. Between 15 October 2006 and 1 February 2007 143 trauma patients (mean ISS 15+/-14 (3-75)) were included. We compared the time in resuscitation room to 650 patients (mean ISS 14+/-14 (3-75)) which were treated between 1 January 2002 and 1 January 2004 according to conventional ATLS protocol. The total-body scanning time was 3.5 min (3-6 min) compared to 25.7 (8-48 min) for conventional X-rays, The total ER time was unchanged 28.7 min (13-58 min) compared to 29.1 min (15-65 min) using conventional plain radiography. In 116/143 patients additional CT scans were necessary. In 98/116 full body trauma CT scans were performed. In 18/116 patients selective CT scans were ordered based on Statscan findings. In 43/143 additional conventional X-rays had to be performed, mainly due to inadequate a.p. views of fractured bones. All radiographs were transmitted over the hospital network (Picture Archiving and Communication System, PACS) for immediate simultaneous viewing at different places. The rapid availability of images for interpretation because of their digital nature and the reduced need for repeat exposures because of faulty radiography are also felt to be strengths.

Teratogenicity of 3-hydroxynorvaline in chicken and mouse embryos.

3-Hydroxynorvaline (HNV; 2-amino-3-hydroxypentanoic acid), a microbial L-threonine analogue, is toxic to mammalian cells and displays antiviral properties. In view of this, we investigated the toxicity and/or potential teratogenicity of HNV in developing chicken and mouse embryos. HNV was administered to chicken embryos (in ovo; dose 75-300 mumole/egg; 48 h post-incubation) and pregnant Hanover NMRI mice (per os; total dose 900-1800 mg/kg body mass; gestation days 7-9). Control animals received sterile saline solutions. Harvested embryos (chicken embryos, 10 days post-incubation; mouse embryos; gestation day 18) were fixed in glutaraldehyde and stereomicroscopically inspected for signs of dysmorphogenesis. Body mass, body and toe length and mortality of chicken embryos, and the body mass and mortality of mouse embryos were recorded. HNV exposure significantly increased the incidence of embryotoxic (growth retardation, toxic mortality) and congenital defects in both chicken and mouse embryos. All the observed effects were dose-dependent. In conclusion, HNV is an embryotoxic and teratogenic compound, which caused significant developmental delay and congenital defects in developing chicken and mouse embryos.

The effect of different apo(a) isoforms on plasminogen activation in black South African subjects.

OBJECTIVE: The present study evaluates the effect of different apolipoprotein(a) [apo(a)] isofroms on plasminogen activation. DESIGN: A cross-sectional study. SETTING: A rural village (Dikgale district) in the Northen Province of South Africa. SUBJECT: A total of 90 apparently healthy subjects (64 females and 36 males) aged 43 to 67 years participated in the study. RESULTS: The mean lipoprotein(a) [ Lp(a)] level in the subjects was 38.14 +/- 22.34 mg/dl. No association was found between Lp(a) isoforms. When the ratio of Lp(a):plasminogen was less then 1.3, a competitive inhibition was observed, but when the ratio exceeded 1.3, an uncompetitive inhibition was observed with all isoforms. CONCLUSION: The results of the present study suggest that the inhibition of plasminogen activation by Lp(a) is not dependent of apo(a) size.

Report on a new type of trauma full-body digital X-ray machine.

The purpose of this study was to evaluate the diagnostic equivalence, radiation dose, clinical usefulness and radiographic aspects of a low-dose, full-body digital X-ray machine in a busy trauma unit. A digital trauma X-ray machine known as "LODOX" was compared with conventional radiography between June 1999 and November 2001 in the Groote Schuur Hospital Trauma Unit, Cape Town. Digital images of a variety of body regions commonly imaged in trauma were compared for diagnostic image quality in a number of categories with equivalent conventional radiographs. A seven-point equivalence scoring system ranging from much inferior (-3) through equivalent (0) to much superior (+3) was used in each category. Radiation dose was recorded and compared with that in conventional measurements. Turnaround times of patients undergoing digital and conventional X-rays were evaluated. Clinical and radiographic issues were assessed by staff feedback. The digital images when compared with conventional film had an overall mean equivalence score of -0.429, with a standard deviation (SD) of 0.77. The best digital performance was in the mediastinum (mean 0.346, SD 0.49) and the weakest was for bony detail (mean -0.654, SD 0.81). Relative digital radiation dose compared to conventional varied from 72% (chest) to 2% (pelvis), with a simple average of 6%. Radiographic points included full-body imaging capability and differing positioning, penetration, workflow and practicality considerations. The digital images required overall patient times of 5-6 min, compared with 8-48 min for conventional X-rays. New installations are under way, and computed tomography and angiography applications are being explored. FDA approval is awaited. Projected cost is similar to that of flat-panel digital units. This digital unit was felt to be diagnostically substantially equivalent to conventional radiographs, with low-dose full-body imaging, improved workflow, digital technology and long-term cost benefits as potentially favourable contributions to trauma imaging.

Steroidogenic correlates of pregnancy in the rock hyrax (Procavia capensis).

In pregnant rock hyraxes isolated leucocytes metabolise both [3H]pregnenolone and [3H]progesterone while whole blood, erythrocytes and an erythrocyte/leucocyte mixture only metabolised [3H]progesterone. Plasma displayed no tendency to metabolically convert any one of these two steroids. In whole blood [3H]progesterone appears to be converted to 5alpha-pregnane-3,20-dione and a compound with chromatographic properties similar to that of 5alpha-pregnan-3alpha-ol-20-one. 5Alpha-pregnane-3,20-dione exhibited a high relative binding affinity for the uterine progesterone eceptor (94%), but 5alpha-pregnan-3alpha-ol-20-one displayed very little affinity for the same receptor (0.4%). 5Alpha-pregnane-3,20-dione may therefore aid in the maintenance of pregnancy. Corpora lutea metabolised progesterone to 17alpha-hydroxyprogesterone, a compound exhibiting no progestational function because of its low relative binding affinity for the uterine progesterone receptor (2%). Progesterone appears to be the main product of the corpus luteum. However, 5alpha-pregnane-3,20-dione circulated at concentrations approximately 8.5 times higher than progesterone, probably due to the metabolic conversion of progesterone to 5alpha-pregnane-3,20-dione by the blood. We conclude that in the hyrax progesterone, produced by the corpora lutea, enters the circulation, where it is reduced to 5alpha-pregnanes. 5Alpha-pregane-3,20-dione may then be transported to the uterus where it binds to the progesterone receptor to assist in the maintenance of pregnancy. This mechanism appears to be analogous to that of the African elephant which is phylogenetically related to the hyrax, except that in the elephant the 5alpha-reduced metabolites are produced by luteal tissue and not the blood.

Folate status, homocysteine metabolism, and methylene tetrahydrofolate reductase genotype in rural South African blacks with a history of pregnancy complicated by neural tube defects.

The birth incidence of neural tube defects (NTDs) in South Africa is threefold to sixfold higher in rural compared with urban blacks. We investigated whether folate deficiency and aberrant homocysteine metabolism could explain the high NTD incidence in rural black populations. Plasma folate and total homocyst(e)ine (tHcy) concentrations were determined in apparently healthy rural black women (n = 107), rural black women with a history of pregnancy complicated by NTDs (n = 54), and urban blacks (n = 101). Methionine load tests were performed on the 54 women with a history of NTD-affected pregnancy and 54 controls matched for age and body mass. The presence of the 677C --> T mutation in the methylene tetrahydrofolate reductase (MTHFR) gene was investigated in both groups by a polymerase chain reaction (PCR) of genomic DNA and HinfI digestion of the PCR product. Apparently healthy urban black women (n = 101) had a lower (P < .001) plasma folate concentration compared with rural black women (n = 107). Women with a history of NTD-affected pregnancy did not differ significantly from controls with respect to plasma folate, fasting homocyst(e)ine, methionine, and the post-methionine load increase in plasma homocyst(e)ine. More than 50% of both of the latter groups had a post-methionine load increase in plasma tHcy less than the fifth percentile as observed in a healthy white control group. No homozygotes for the 677C --> T mutation in the MTHFR gene were found in black mothers with NTD-affected offspring or controls. It is concluded that black urbanization is characterized by a diminished folate status that is paradoxically associated with a lower NTD birth incidence. Homozygosity for the 677C --> T mutation in the gene coding for MTHFR does not constitute a genetic risk factor for NTDs in blacks. No aberrant homocysteine metabolism could be demonstrated in black women with NTD-affected pregnancies.

Influence of gestation on uterine endometrial steroid receptor concentrations in the African elephant, Loxodonta africana.

The modulatory effects of gestational age and circulating concentrations of progesterone, 5alpha-pregnane-3,20-dione, and estradiol-17beta on the uterine sex steroid hormone receptor levels of the African elephant were investigated. Uterine tissue biopsies and blood samples were obtained from animals culled in the Kruger National Park. Estrogen and progesterone receptor concentrations were determined in uterine biopsies from subadult, lactating, early-, mid-, and late-pregnant elephants, by equilibrium binding assays. Circulating estradiol-17beta and progesterone concentrations were measured by means of RIAs, while plasma concentrations of 5alpha-pregnane-3,20-dione were determined with an amplified ELISA. Significant inverse correlations of the concentrations of estrogen and progesterone receptors with the gestational stage of the elephants were observed. Pregnant uterine horns of individual animals contained lower levels of estrogen and progesterone receptors than the nonpregnant horns of the same animals. A strong positive correlation existed between uterine estrogen and progesterone receptors levels. Circulating concentrations of 5alpha-pregnane-3,20-dione and progesterone decreased with an increase in the concentrations of progesterone receptors as well as with fetal age. We conclude that the progesterone receptor concentrations are down-regulated with progressing gestation in the African elephant. This down-regulation appears to be linked to an increase in circulatory 5alpha-pregnane-3,20-dione concentration in the plasma of pregnant animals.

The effect of Simvastatin on the plasma antioxidant concentrations in patients with hypercholesterolaemia.

The aim of this study was to monitor the antioxidant status of patients with hypercholesterolaemia during treatment with Simvastatin. Forty-seven patients, of whom 25 had confirmed familial hypercholesterolaemia (FH), were treated with 10 or 20 mg of Simvastatin per day for 14 weeks. As expected, total cholesterol and LDL cholesterol concentrations decreased considerably, while HDL cholesterol concentrations increased during drug treatment. In neither FH nor non-FH patients were any significant changes observed for retinol status, while plasma vitamin C concentrations were also not adversely affected by the drug therapy. In both patient groups Simvastatin therapy led to a significant decrease in plasma alpha-tocopherol (P < 0.05) concentrations, however, the alpha-tocopherol/total cholesterol ratio increased by 9.1 (P < 0.01) and 12.1% (P < 0.01) in FH and non-FH patients, respectively, during the 14-week treatment period. The coenzyme Q10/total cholesterol ratio did not change significantly in non-FH patients, but was significantly lower (P < 0.05) than the baseline ratio after 4 and 14 weeks of Simvastatin treatment in FH patients. The alpha-tocopherol/total cholesterol ratio of FH patients remained consistently and significantly lower (P < 0.01) compared with non-FH patients, indicating that LDL from the former group may be more vulnerable to free radical-mediated damage and lipid peroxidation. Our results suggest that the significant decline in circulating alpha-tocopherol and coenzyme Q10 concentrations was mainly a function of the decrease in serum total cholesterol concentrations.

Spontaneous oxidation of methionine: effect on the quantification of plasma methionine levels.

Plasma methionine (Met), methionine sulfoxide (MSO), and total Met concentrations were determined by reversed-phase chromatography and fluorescence detection after automated precolumn derivatization with an o-phthalic aldehyde mercaptoethanol reagent. Addition of pure, MSO-free L-Met to plasma samples resulted in the anticipated linear increase in plasma Met concentrations, but simultaneously effected a dose-dependent, linear increase in MSO levels. In contrast, the addition of pure L-MSO to plasma samples rendered linear calibration curves for MSO, while the Met concentration remained constant. A strong buffering effect against the spontaneous or hydrogen peroxide induced oxidation of Met to MSO was observed in plasma samples. This protective effect could be neutralized by preincubating the plasma samples with sodium azide. The addition of relatively low concentrations of red cell lysates to plasma samples, prior to hydrogen peroxide oxidation, strongly inhibited the conversion of Met to MSO. Plasma samples from 127 healthy female volunteers were analyzed: MSO concentrations (mean, 3.6 +/- 2.1 microM) exhibited a weak positive correlation (r = 0.352) with Met levels (mean, 21.3 +/- 6.1 microM) but, after the exclusion of two probable outliers from the data set, no correlation was observed. Our results suggest that plasma Met concentrations should be corrected for oxidative losses incurred during storage, sample processing and because of the action of a variety of in situ oxidants, present in plasma, in order to obtain a reliable estimate of the methionine status of an individual.

Ligand specificity of uterine oestrogen and progesterone receptors in the subadult African elephant, Loxodonta africana.

The ligand specificity of progesterone and oestrogen receptors in the uteri of four nonpregnant, nonlactating African elephants, killed during routine culling in the Kruger National Park, were determined. The mean (+/-SEM) Kd values of the oestrogen (0.18 +/- 0.019 x 10(-9) mol l-1, n = 12) and progesterone (0.22 +/- 0.025 x 10(-9) mol l-1, n = 12) receptors were essentially similar when [3H]promegestone was used as radioligand in the progesterone receptor assays. However, when [3H]progesterone was used as radioligand, the progesterone receptor exhibited a significantly higher Kd value (1.03 +/- 0.132 x 10(-9) mol l-1, n = 12) than that of the oestrogen receptor. The use of the different radioligands did not significantly affect the quantitative values obtained for the progesterone receptor. Both the oestrogen and the progesterone receptors displayed a high ligand specificity. The 5 alpha-reduced metabolites of progesterone exhibited a high relative binding affinity for the progesterone receptor (5 alpha-pregnane-3,20-dione: relative binding affinity = 43%; 5 alpha-pregnane-3 alpha-ol-20-one: relative binding affinity = 20%) but the synthetic antiprogestin RU 486 did not compete successfully with progesterone in competitive binding studies. However, norethindrone (relative binding affinity = 293%) competed successfully for binding to the progesterone receptor, and may have some potential in the future development of a technique to control reproductive output in the African elephant.

Correlation between essential fatty acids and parameters of bone formation and degradation.

There are two types of essential fatty acids (EFAs), the n-6 derived from linoleic acid (LA) and the n-3, derived from alpha-linolenic acid (ALA). Most of the functions of the EFAs require the conversion of LA and ALA to their metabolites including, gammalinolenic (GLA), dihomogammalinolenic (DGLA), arachidonic (AA) (n-6) and eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids (n-3). Supplementing specific GLA:EPA ratios has effects on bone formation and degradation. A study was designed to investigate the effect of various dietary ratios of n-6:n-3 on calcium homeostasis. Female Sprague Dawley rats were ovariectomised (OVX) at age =11 weeks, and were supplemented from age 12 weeks for six weeks with different ratios (9:1; 3:1; 1:3; 1:9) of GLA:EPA. Bone parameters and red blood cell (RBC) fatty acid profiles were measured at age=18 weeks. RBC GLA and DGLA increased in groups 9:1 and 3:1(p<0.05). EPA and DGLA increased in 1:3 and 1:9 while AA decreased (p<0.05). Correlations were calculated between bone calcium, deoxypyridinoline (Dpyd) and specific fatty acids. DGLA was positively correlated with femur calcium and negatively with Dpyd excretion while DHA and EPA were correlated with femur calcium.

Designing a transmandibular implant with the aid of biostereometric analysis.

A biostereometric survey of severely resorbed dry mandibles was used to determine whether it was possible to design a transmandibular implant that could be used for the majority of clinical situations. Computerized profiles of the relevant mandibular areas were generated and superimposed to examine the feasibility of a universal shape. It would appear that the crestal shape of the mandible between the mental foramina is on the circumference of a circle and that the surface of the inferior border is so variable that a flat surface would have to be used for a transmandibular implant. A design of a transmandibular implant is proposed on the basis of the common anatomic features of the mandible.

Effective homocysteine metabolism may protect South African blacks against coronary heart disease.

L-Methionine (0.1 g/kg body wt) was administered to young white [n = 18; mean (+/- SD) age 20.0 +/- 1.0 y] and black [n = 12; mean (+/- SD) age 22.0 +/- 1.3 y] volunteers who had a similar lifestyle and who did not differ significantly from each other with respect to plasma folate or vitamin B-12 concentrations. Blacks, however, had significantly lower plasma pyridoxal-5'-phosphate concentrations compared with whites (P < 0.001). Fasting plasma homocysteine concentrations in blacks and whites were not significantly different. The mean (+/- SD) maximum increase in plasma homocysteine concentration measured after methionine loading was significantly lower (P < 0.01) in blacks (11.0 +/- 3.6 mumol/L) than in whites (18.0 +/- 6.2 mumol/L). Six weeks of vitamin supplementation (1.0 mg folic acid, 400 micrograms vitamin B-12, and 10 mg pyridoxine/d) reduced the mean (+/- SD) fasting plasma homocysteine concentration from 9.6 +/- 3.5 to 7.2 +/- 1.6 mumol/L in whites (P < 0.05) and from 8.4 +/- 2.4 to 5.6 +/- 1.4 mumol/L in blacks (P < 0.01). The mean (+/- SD) maximum increase in plasma homocysteine concentration after methionine loading declined from 18.0 +/- 6.2 to 11.1 +/- 2.3 mumol/L (P < 0.01) in whites, but vitamin supplementation did not have a significant effect on the methionine-load test in black volunteers. A significant race-by-time interaction shows that blacks metabolized homocysteine more effectively than did whites, which may partly explain their relative resistance against coronary heart disease despite a high prevalence of obesity, hypertension, and smoking.

Oestrogen and progesterone receptor status and PgR/ER ratios in normal and myomatous human myometrium.

The concentration and molecular properties of the oestrogen (ER) and the progesterone (PgR), present in normal myometria and uterine leiomyometria, obtained from a group of age-matched, pre-menopausal, negroid female patients were investigated. Serum oestrogen and progesterone levels did not differ significantly in the two groups. Significant differences were detected in ER and PgR levels between normal and leiomyomatous myometria. Both ER (154%; p < 0.0001) and PgR (33%; p < 0.05) were significantly increased in uterine leiomyomas. PgR levels were less affected than the ER levels, causing a significant decrease (44%; p < 0.05) in the PgR/ER ratio in myomatous myometria. Dissociation and sedimentation constants, as well as iso-electric points of ER and PgR were essentially similar in normal and in myomatous myometria. According to our results, tissue pathology does not appear to be associated with defects in the molecular properties of ER and PgR, but with differential changes in the ER and PgR levels, subsequently affecting the PgR/ER ratio.

Supplemented gamma-linolenic acid and eicosapentaenoic acid influence bone status in young male rats: effects on free urinary collagen crosslinks, total urinary hydroxyproline, and bone calcium content.

The effect of different ratios of the prostaglandin precursors gamma-linolenic (GLA) and eicosapentaenoic (EPA) acids on bone status in growing rats measured as a function of free urinary pyridinium crosslinks and hydroxyproline levels was investigated. Male Sprague-Dawley rats were weaned onto an essential fatty acid deficient diet and from their fifth week, different groups of rats received a balanced, semisynthetic diet, supplemented with different ratios of GLA:EPA supplied as a mixture of evening primrose oil (EPO) and fish oil (FO). Controls were supplemented with linoleic (LA; sunflower oil) and alpha-linolenic (ALA; linseed oil) acids (3:1) or a commercially available rat chow. Animals were terminated at 84 days and femur length, ash weight, calcium content, free urinary pyridinium crosslinks (Pyd and Dpyd), total hydroxyproline (Hyp), and creatinine levels measured. Free urinary Pyd and Dpyd are good indicators of bone status and they correlated well with Hyp. Pyd and Dpyd excretion were significantly decreased in the higher GLA:EPA dietary groups and correlated well (r = 0.7) with Hyp levels. Concomitantly, bone calcium content increased significantly in the same dietary groups. These results suggest that diet supplementation with relatively high GLA:EPA ratios are more effective in inhibiting bone resorption than LA:ALA.

Cholate and pH reduce interference by sodium dodecyl sulfate in the determination of DNA with Hoechst.

The use of the fluorescent dye 33258 Hoechst (Hoe) to quantitatively determine DNA in cell culture in the presence of lysing agents like sodium dodecyl sulfate (SDS) is limited by the masking effect of high levels of nonspecific fluorescence, caused by the binding of Hoe to micelles. The masking effect can be reduced substantially by increasing the concentration of the counterion, the addition of cholate, or the pH of the buffer. An optimized method was developed, combining the antimasking effects of sodium chloride, cholate, and pH to accurately determine DNA concentrations as low as 15 ng/ml in the presence of up to 6.9 mM (0.2%) SDS. The effectiveness of SDS in cell dissolution can now be combined with the specificity and sensitivity of Hoe to determine cellular DNA.