Sterols and oxysterols

The ingredients of animal feed formulation, especially the fatty sources, play a significant role in the production of high-quality food of animal origin. Industrial fat by-products are a major source of feeding fats in Europe. The main objectives of the project were firstly to assess feed fats and oils for their composition and content of sterols (cholesterol and phytosterols) and oxysterols, and secondly to assess the levels of cholesterol and oxycholesterols of chicken and rabbit tissues after feeding with especially formulated feeds containing trans fatty acids and oxidized lipids. The lack of standardized analytical procedure prompted the evaluation of an in-house purification method by comparing it with a number of commonly used methods for the analysis of oxysterols. The saponification and transesterification steps showed rather comparable results. A two-dimensional capillary GC column with different stationary phases (a 35% phenyl column coupled to an apolar 5% phenyl column) was used for better resolution of a large number of oxysterols. This new system improved the separation efficiency and reduced the analytical time for a wide range of oxysterols. The satisfactory purification method and the reliable separation of oxysterols facilitated the qualitative and quantitative assessment of sterols and oxysterols in samples of by-products from chemical and physical refining. A large variation in the levels of sterols and oxysterols was observed in the fat by-products from chemical and physical refining processes for edible fats and oils. The observed variations in the contents and composition of sterols and oxysterols were mainly due to the origins, production facility and different processing conditions of the by-product samples. The highly oxidized lipid and trans fatty acid feeds significantly increased the contents of cholesterol and oxycholesterols in edible parts, e.g. the muscles and livers of chickens and rabbits (0.01< p ≤0.05). Hence, the consumption of products from animals fed with higher levels of trans fatty acids and oxidized lipid feeds may contribute to higher ingestion of cholesterol and oxycholesterols by humans

Cholesterol oxidation products : analytical methods and levels in sweets containing heated butter oil

Various methods are used to analyse cholesterol oxidation products (COP) due to the unavailability of a standard method. In order to select a suitable method for the enrichment of COP, three methods of saponication (A – C),and transesterication (D) of tallow with three levels (5,10 and 20 µg) of spiked COP, were evaluated. Further enrichment of COP was done by solid phase extraction, quantied by GC, and confirmed by GC–MS. The in-house method A, and method D, showed the best results among the four methods evaluated. The recoveries at all levels of spiked COP were generally higher than 60% in method A. The recoveries of all spiked COP at 5 µg level were consistently lower in method D compared with method A. From the results of this study it can be concluded that method A may be more suitable for the analysis of very low levels of COP in foods. Ghee (clarified butter oil), a major ingredient in Indian sweets, is an important source of saturated fatty acids, cholesterol and COP that are considered as risk factors for atherosclerosis. The high frequency of atherosclerotic complications reported among the Indian immigrants in England prompted determination of lipids and lipid oxidation status of a ghee sample and fifteen Indian sweets that are available in London supermarkets. The fatty acid profile of the samples shows saturated fats (about 73%), mainly composed of myristic, palmitic and stearic acids, except in two samples, which had levels of oleic acid in excess of 60%. TBARS values ranging from 19 to 260 µg/100 g might be due to the different oxidative status of different sweets samples. Cholesterol (approximately 0.22%) was the main sterol in all samples, except two where unspecified oils were used instead. Variation of total COP was from 0.94 to 38.41 µg/g sample. Some of these sweets may be a source of considerable amounts of saturated fatty acids, cholesterol and COP in the diet that possibly contribute to atherosclerosis

Concentrations of canine prostate specific esterase, CPSE, at baseline are associated with the relative size of the prostate at three-year follow-up

BackgroundEnlargement of the prostate is associated with prostatic diseases in dogs, and an estimation of prostatic size is a central part in the diagnostic workup. Ultrasonography is often the method of choice, but biomarkers constitute an alternative. Canine prostate specific esterase (CPSE) shares many characteristics with human prostate specific antigen (PSA) and is related to prostate size. In men with clinical symptoms of prostatic disease, PSA concentrations are related to prostate growth. The aims of the present follow-up study were to evaluate if the concentration of CPSE is associated with future growth of the prostate, and if analysis of a panel of 16 steroids gives further information on prostatic growth. Owners of dogs included in a previous study were 3 years later contacted for a follow-up study that included an interview and a clinical examination. The prostate was examined by ultrasonography. Serum concentrations of CPSE were measured, as was a panel of steroids.ResultsOf the 79 dogs included at baseline, owners of 77 dogs (97%) were reached for an interview, and 22 were available for a follow-up examination. Six of the 79 dogs had clinical signs of prostatic disease at baseline, and eight of the remaining 73 dogs (11%) developed clinical signs between baseline and follow-up, information was lacking for two dogs. Development of clinical signs was significantly more common in dogs with a relative prostate size of >= 2.5 at baseline (n=20) than in dogs with smaller prostates (n=51). Serum concentrations of CPSE at baseline were not associated with the change in prostatic size between baseline and follow-up. Serum concentrations of CPSE at baseline and at follow-up were positively associated with the relative prostatic size (S-rel) at follow-up. Concentrations of corticosterone (P = 0.024), and the class corticosteroids (P = 0.0035) were positively associated with the difference in S-rel between baseline and follow-up.ConclusionsThe results support the use of CPSE for estimating present and future prostatic size in dogs >= 4years, and the clinical usefulness of prostatic size for predicting development of clinical signs of prostatic disease in the dog. The association between corticosteroids and prostate growth warrants further investigation

Acute effects of methcathinone and manganese in mice: A dose response study

An intravenously injectable illicit drug made by mixing pseudoephedrine, potassium permanganate, vinegar and water, yielding methcathinone (Mcat) and manganese (Mn), induces an extrapyramidal syndrome with parkinsonism, dystonia, gait and balance disorders similar to manganism. Although the cause of the syndrome is largely attributed to Mn, the interaction of the drug's individual components is not known and the role of Mcat is possibly underestimated. Aim of the present study was to analyze dose-dependent behavioral effects of the mixture and its two main active components Mcat and Mn in an acute setting and determine the lethal doses of each substance. Three groups of C57BL/6 mice were injected intraperitoneally with (1) the drug mixture containing 10, 25, 50, 100 or 150 mg of Mcat and respectively 1.6, 3.8, 6.9, 17.1 and 22.6 mg of Mn per kilogram of body weight; (2) 10, 25, 50, 100, 150, 200 or 300 mg of racemic Mcat/kg of body weight; (3) MnCl2 10, 25 or 50 mg/kg of body weight. Locomotor activity of the animals, various signs and time of death were recorded. Lower doses (10 and 25 mg/kg) of Mcat had a clear motor activity stimulating effect and this was clearly dose-dependent. High doses of Mcat produced epileptic seizures in 74% of the animals and became lethal with the highest doses. Similarly, the mixture had a clear dose-dependent stimulating effect and the higher doses became lethal. The LD50 of the pseudoephedrine mixture was 110.2 mg of Mcat/kg and for pure Mcat 201.7 mg/kg. Mn did not prove to be lethal in doses up to 50 mg/kg, but had a strong dose dependent inhibitory effect on the animals’ behavior. Our data reveal that both Mn and Mcat have a significant role in the toxicity of the mixture

A Novel Targeted Analysis of Peripheral Steroids by Ultra-Performance Supercritical Fluid Chromatography Hyphenated to Tandem Mass Spectrometry

Ultra-performance supercritical fluid chromatography-tandem mass spectrometry (UPSFC-MS/MS) is an alternative method for steroid analysis. Continuous development of analytical methodologies for steroid profiling is of major importance in the clinical environment to provide useful and more comprehensive data. The aim of this study was to identify and quantify a large number of endogenous steroids from the four major classes (estrogens, androgens, progestogens and corticosteroids) simultaneously within a short analytical time. This novel UPSFC-MS/MS method with electrospray in positive ionisation (ESI+) mode is robust, selective and present sufficiently high sensitivity to profile nineteen steroids in 50 mu L human plasma. Under optimised conditions, nineteen different steroids were separated with high efficiency in the multiple reaction monitoring (MRM) mode. The linearity of the method was good with correlation coefficients (R-2) in the range of 0.9983-0.9999 and with calibration range from 0.05-500 ng/mL in human plasma. The intraday and interday precision of the method, as RSD, was less than 15%. The accuracy of the nineteen analytes varied between 80 to 116%. Finally, the novel method was successfully applied for the determination of nineteen steroids within 5 minutes providing the possibility to use it for research as well as routine healthcare practice

Hyperandrogenic Symptoms Are a Persistent Suffering in Midlife Women with PCOS; a Prospective Cohort Study in Sweden

Polycystic ovary syndrome (PCOS) is a common endocrine disorder among women, and the majority suffers from hyperandrogenism. Hyperandrogenism causes psychological morbidity and impaired quality of life in women with PCOS during the reproductive years, but data on prevalence and impact during midlife are lacking. Thus, this study aimed to address whether hyperandrogenism persists into midlife and, if so, what impact it has on quality of life. In order to answer this question, we performed a multicenter prospective cohort study, where we included women already diagnosed with PCOS who had reached the age of 45 years or more and age-matched controls. All participants underwent a physical exam, structured medical interview, biochemical testing and filled out self-assessment questionnaires. More than 40% of the women with PCOS and 82% of those who presented with the hyperandrogenic phenotype at the diagnostic work-up still suffered from hirsutism. Circulating testosterone levels were similar between women with PCOS and controls while free androgen index was higher in women with PCOS, independent of weight. Women with hyperandrogenic PCOS expressed persisting concerns regarding hirsutism at the follow-up assessment. In conclusion, women with PCOS who present with hyperandrogenic symptoms at the time they are diagnosed with PCOS have a higher risk of persistent androgenic symptoms and impaired quality of life in midlife

Hyperandrogenic symptoms are a persistent suffering in midlife women with pcos; a prospective cohort study in sweden

Polycystic ovary syndrome (PCOS) is a common endocrine disorder among women, and the majority suffers from hyperandrogenism. Hyperandrogenism causes psychological morbidity and impaired quality of life in women with PCOS during the reproductive years, but data on prevalence and impact during midlife are lacking. Thus, this study aimed to address whether hyperandrogenism persists into midlife and, if so, what impact it has on quality of life. In order to answer this question, we performed a multicenter prospective cohort study, where we included women already diagnosed with PCOS who had reached the age of 45 years or more and age-matched controls. All participants underwent a physical exam, structured medical interview, biochemical testing and filled out self-assessment questionnaires. More than 40% of the women with PCOS and 82% of those who presented with the hyperandrogenic phenotype at the diagnostic work-up still suffered from hirsutism. Circulating testosterone levels were similar between women with PCOS and controls while free androgen index was higher in women with PCOS, independent of weight. Women with hyperandrogenic PCOS expressed persisting concerns regarding hirsutism at the follow-up assessment. In conclusion, women with PCOS who present with hyperandrogenic symptoms at the time they are diagnosed with PCOS have a higher risk of persistent androgenic symptoms and impaired quality of life in midlife

Plasma levels of arginines at term pregnancy in relation to mode of onset of labor and mode of childbirth

PROBLEM: The exact biochemical mechanisms that initiate labor are not yet fully understood. Nitric oxide is a potent relaxant of uterine smooth muscles until labor starts, and its precursor is L-arginine. Asymmetric (ADMA) and symmetric (SDMA) dimethylarginines, are potent NO-inhibitors. However, arginines (dimethylarginines and L-arginine) are scarcely studied in relation to labor and childbirth. We aimed to investigate arginines in women with spontaneous (SLVB) and induced (ILVB) term labor with vaginal birth and in women undergoing elective caesarean section (ECS). METHOD OF STUDY: Women at gestational week 16-18 were recruited to the population-based prospective cohort study BASIC at the Uppsala University Hospital, Sweden. Plasma samples taken at start of labor were analyzed for arginines, from SLVB (n = 45), ILVB (n = 45), and ECS (n = 45), using Ultra-High Performance Liquid Chromatography. Between-group differences were assessed using Kruskal-Wallis and Mann-Whitney U-test. RESULTS: Women with SLVB and ILVB had higher levels of ADMA (p &lt; .0001), SDMA (p &lt; .05) and lower L-arginines (p &lt; .01), L-arginine/ADMA (p &lt; .0001), and L-arginine/SDMA (p &lt; .01, respectively &lt;.001) compared to ECS. However, ILVB had higher ADMA (p &lt; .0001) and lower L-arginine (p &lt; .01), L-arginine/ADMA (p &lt; .0001), and L-arginine/SDMA (p &lt; .01) compared to SLVB. Results are adjusted for gestational length at birth and cervical dilatation at sampling. CONCLUSION: Our novel findings of higher levels of dimethylarginines in term vaginal births compared to ECS give insights into the biochemical mechanisms of labor. These findings might also serve as a basis for further studies of arginines in complicated pregnancies and labor

Domestication Effects on Stress Induced Steroid Secretion and Adrenal Gene Expression in Chickens

Understanding the genetic basis of phenotypic diversity is a challenge in contemporary biology. Domestication provides a model for unravelling aspects of the genetic basis of stress sensitivity. The ancestral Red Junglefowl (RJF) exhibits greater fear-related behaviour and a more pronounced HPA-axis reactivity than its domesticated counterpart, the White Leghorn (WL). By comparing hormones (plasmatic) and adrenal global gene transcription profiles between WL and RJF in response to an acute stress event, we investigated the molecular basis for the altered physiological stress responsiveness in domesticated chickens. Basal levels of pregnenolone and dehydroepiandrosterone as well as corticosterone response were lower in WL. Microarray analysis of gene expression in adrenal glands showed a significant breed effect in a large number of transcripts with over-representation of genes in the channel activity pathway. The expression of the best-known steroidogenesis genes were similar across the breeds used. Transcription levels of acute stress response genes such as StAR, CH25 and POMC were upregulated in response to acute stress. Dampened HPA reactivity in domesticated chickens was associated with changes in the expression of several genes that presents potentially minor regulatory effects rather than by means of change in expression of critical steroidogenic genes in the adrenal.Funding agencies: Swedish Research Council (VR) [621-2011-4731]; Swedish Research Council for Environment, Agricultural Sciences and Spatial Planning (FORMAS) [221-2011-1088]; SRC [621-2011-5523]; ERC [322206]; Swedish Centre of Excellence in Animal Welfare</p

Functional properties of low-modulus PMMA bone cements containing linoleic acid

Acrylic bone cements modified with linoleic acid are a promising low-modulus alternative to traditional high-modulus bone cements. However, several key properties remain unexplored, including the effect of autoclave sterilization and the potential use of low-modulus cements in other applications than vertebral augmentation. In this work, we evaluate the effect of sterilization on the structure and stability of linoleic acid, as well as in the handling properties, glass transition temperature, mechanical properties, and screw augmentation potential of low-modulus cement containing the fatty acid. Neither 1H NMR nor SFC-MS/MS analysis showed any detectable differences in autoclaved linoleic acid compared to fresh one. The peak polymerization temperature of the low-modulus cement was much lower (28–30 °C) than that of the high-modulus cement (67 °C), whereas the setting time remained comparable (20–25 min). The Tg of the low-modulus cement was lower (75–78 °C) than that of the high-stiffness cement (103 °C). It was shown that sterilization of linoleic acid by autoclaving did not significantly affect the functional properties of low-modulus PMMA bone cement, making the component suitable for sterile production. Ultimately, the low-modulus cement exhibited handling and mechanical properties that more closely match those of osteoporotic vertebral bone with a screw holding capacity of under 2000 N, making it a promising alternative for use in combination with orthopedic hardware in applications where high-stiffness augmentation materials can result in undesired effects.Authors in thesis list of papers: C. Robo, D. Wenner, M. Nilsson, J. Hilborn, C. Öhman-Mägi, C. Persson</p