Determination of phytosterols in oenological matrices by liquid chromatography-atmospheric pressure chemical ionization and ion-trap mass spectrometry

The aim of the present study is the identification of plant sterols and the development of an analytical method that allows for the quantification of such family of compounds in oenological matrices. The application of liquid chromatography-atmospheric pressure chemical ionization ion-trap mass spectrometry (LC-APCI-ITMS) to sterol characterization is a useful tool and was selected to perform this research. Sterol separation was achieved using a C8 column with a mobile phase consisting of water and acetonitrile under gradient conditions and column temperature of 35 °C, which leads to analyte elution in less than 25 min. Retention times, precursor ions and MRM transitions of analytes allowed for the identification and sensitive quantitative determination of phytosterols in oenological matrices at trace levels. The method showed a dynamic linear range over the concentration ranges from 0.02 to 320 mg kg⁻¹ for the different parts of grapes and from 8 to 100 ng mL⁻¹ in case of wine. The most abundant phytosterol in all samples was β-sitosterol. The seeds are the richest source of phytosterols having a great amount of β-sitosterol, 314 mg kg⁻¹ fresh berry mass, followed by stigmasterol, fucosterol and campesterol at much lower concentrations (ranging from 3 to 10 mg kg⁻¹).     

A nutraceutical based approach to reduce cholesterolaemia in patients previously intolerant of more than a statin: a pilot study

This study was aimed at evaluating the tolerability of a nutraceutical combination with anticholesterolaemic action in patients intolerant of statin treatment. A total of 32 hypercholesterolaemic patients, all intolerant to at least two statin treatments, were enrolled in the ambulatory service. None of the enrolled patients was diabetic or undergoing secondary prevention of cardiovascular disease. Patients consumed one yoghurt with 2 g of added phytosterols each morning (Pro-Activ, Unilever, Milan, Italy) and one tablet of a registered combined nutraceutical (Armolipid Plus, Rottapharm, Monza, Italy). We tested the efficacy of the treatment after 4 and 6 months. One tablet of Armolipid Plus contains: berberine 500 mg, monacolin 3 mg, and policosanol 10 mg. After 3 months of treatment the patients showed a significant decrease in total cholesterol, low-density lipoprotein cholesterol and triglycerides, and a significant increase in the plasma level of high-density lipoprotein cholesterol. These results were maintained after 6 months of treatment without a significant change in the mean values.     

我国三城市老年妇女膳食植物甾醇摄人量与血脂含量的关系

目的 对我国3个城市老年妇女膳食植物甾醇的摄入量进行调查,比较其主要膳食来源,初步探讨不同膳食模式下植物甾醇摄入量与血脂含量的关系.方法 根据不同膳食结构和特点,选择北京、合肥、乌鲁木齐市为调查点,各调查点选择50岁以上的妇女80～100名,利用2 d连续24 h回顾法对其膳食进行调查.同时对其膳食调查中所涉及的常见植物食物进行采样,气相色谱法分析食物中植物甾醇的含量(包括β-谷甾醇、菜油甾醇、豆甾醇、谷甾烷醇)并计算总植物甾醇含量.在样品分析的基础上,计算被调查者膳食植物甾醇的摄入量并分析其主要来源,同时对被调查者进行体格检查和血液生化指标测定.结果 北京、合肥、乌鲁木齐市符合条件的被调查者各有100、101和84名.其中北京和合肥市被调查者膳食植物甾醇的摄入量平均值分别为340.3 mg/d和313.5mg/d,主要来源是植物油类和谷类食物;乌鲁木齐市被调查者膳食植物甾醇平均摄入量为550.4mg/d,高于北京和合肥市(t值分别为9.369、10.420,P值均<0.01),其主要来源为谷类食物(提供总摄入量的53.1%).血生化指标结果显示,乌鲁木齐市被调查者的血脂血糖含量低于其余2个城市的被调查者,其中血清总胆固醇(TC)含量为(4.04±0.78)mmol/L,低于北京[(4.89±0.91)mmol/L]和合肥市[(4.71±0.83)mmoL/L](t值分别为6.766、5.401,P值均<0.01);血清甘油三酯(TG)含量为(1.01±0.48)mmol/L,低于北京[(1.3l±0.53)mmol/L]和合肥市[(1.66±0.75)mmol/L](t值分别为3.343、7.293,P值均<0.01);同时乌鲁木齐市被调查者的血糖含量平均值[(5.02±2.18)mmoL/L]也低于北京[(5.69±1.53)mmol/L,t=2.561,P<0.05]和合肥市[(5.78±1.53)mmol/L,t=2.934,P<0.01].结论 膳食结构不同导致不同地区老年妇女植物甾醇摄入量差别较大,摄入较多的植物甾醇有助于降低老年妇女血脂水平.     

Effect of dalcetrapib, a CETP modulator, on non-cholesterol sterol markers of cholesterol homeostasis in healthy subjects

Objective

Subjects with high HDL-C show elevated plasma markers of cholesterol absorption and reduced markers of cholesterol synthesis. We evaluated the effect of dalcetrapib, a cholesteryl ester transfer protein modulator, on markers of cholesterol homeostasis in healthy subjects.

Methods

Dalcetrapib was administered daily with or without ezetimibe in a randomized, open-label, crossover study in 22 healthy subjects over three 7-day periods: dalcetrapib 900 mg, ezetimibe 10 mg, dalcetrapib 900 mg plus ezetimibe 10 mg. Plasma non-cholesterol sterols lathosterol and desmosterol (cholesterol synthesis markers) and campesterol, β-sitosterol and cholestanol (intestinal cholesterol absorption markers) were measured. A hamster model was used to compare the effect of dalcetrapib and torcetrapib with or without ezetimibe on these markers and determine the effect of dalcetrapib on cholesterol absorption.

Results

Dalcetrapib increased campesterol, β-sitosterol, and cholestanol by 27% (p = 0.001), 32% (p < 0.001), and 12% (p = 0.03), respectively, in man (non-cholesterol sterol/cholesterol ratio). Dalcetrapib + ezetimibe reduced campesterol by 11% (p = 0.02); β-sitosterol and cholestanol were unaffected. Lathosterol and desmosterol were unchanged with dalcetrapib, but both increased with ezetimibe alone (56–148%, p < 0.001) and with dalcetrapib + ezetimibe (32–38%, p < 0.001). In hamsters, dalcetrapib and torcetrapib increased HDL-C by 49% (p = 0.04) and 72% (p = 0.003), respectively. Unlike torcetrapib, dalcetrapib altered cholesterol homeostasis towards increased markers of cholesterol absorption; cholesterol synthesis markers were unaffected by either treatment. Dalcetrapib did not change plasma ³H-cholesterol level but increased ³H-cholesterol in plasma HDL vs non-HDL, after oral dosing of labeled cholesterol.

Conclusion

Dalcetrapib specifically increased markers of cholesterol absorption, most likely reflecting nascent HDL lipidation by intestinal ABCA1, without affecting markers of synthesis.     

Association of plasma markers of cholesterol homeostasis with metabolic syndrome components. A cross-sectional study

Background and aims

Increased plasma phytosterols, which reflect enhanced cholesterol absorption, have been related to an increased risk of cardiovascular disease (CVD). However, high CVD risk conditions, such as obesity, diabetes and the metabolic syndrome (MetS) have been associated with reduced cholesterol absorption. We investigated associations between plasma noncholesterol sterols and MetS components.

Methods and results

With a cross-sectional design, we related MetS components to plasma noncholesterol sterol-to-cholesterol ratios measured by gas chromatography in 674 dyslipidemic patients and 361 healthy subjects participating in a prospective cohort study.Plasma phytosterol-to-cholesterol ratios were inversely associated with all components of the MetS. In the dyslipidemic group, multivariable analyses showed that a 1-SD increase in sitosterol-to-cholesterol ratio was associated with a reduced risk for any MetS feature, ranging from 0.57 (95% CI, 0.45 to 0.71) for visceral adiposity to 0.82 (95% CI, 0.69 to 0.98) for high blood pressure. The risk of having MetS was nearly halved, with ORs of 0.49 (95% CI, 0.38 to 0.64) or 0.56 (95% CI, 0.44-0.70), depending on the definition. Results were opposed for plasma lathosterol, a marker of cholesterol synthesis. Most findings were reproduced in the healthy cohort. ApoE genotype was unrelated to plasma noncholesterol sterols.

Conclusion

In both dyslipidemic and healthy populations, MetS is associated with increased plasma lathosterol, a cholesterol synthesis marker, and decreased plasma sitosterol, a marker of cholesterol absorption. Elevated plasma phytosterols related to a lower frequency of cardiometabolic risk factors, suggesting that they are associated with a reduced CVD risk.     

Antiplatelet effects of aspirin with phytosterols: Comparison with non-enteric coated aspirin alone

The novel combination of aspirin and phytosterols may be a potential strategy to treat patients with cardiovascular disease. We sought to determine if the antiplatelet effects of a combination caplet of 81 mg aspirin with 400 mg phytosterols differed from the antiplatelet effects of non-enteric coated aspirin. The first five days of aspirin therapy alone (T1) produced marked reductions in collagen-induced, ADP-induced, and archidonic acid- induced platelet aggregation, and in serum and urine TxB₂ compared to baseline. Five days after randomization to aspirin alone versus aspirin + phytosterols (T2), there were no differences in any measurement of platelet function within each group compared to T1 or between groups. The present study suggests that the antiplatelet effect of non-enteric coated 81 mg twice-daily aspirin therapy alone is not affected by the addition of phytosterols in a combination product.     

Differential effects of cholesterol and phytosterols on cell proliferation, apoptosis and expression of a prostate specific gene in prostate cancer cell lines

Background: The purpose of our study was to show the apoptotic and anti-proliferative effects of phytosterols as distinct from cholesterol effects on prostate cancer cell lines, and also their differential expression of caveolin-1, and a prostate specific gene, PCGEM1. Methods: PC-3 and DU145 cells were treated with sterols (cholesterol and phytosterols) for 48 h, followed by trypan blue dye exclusion measurement of cytotoxicity and MTT cell proliferation assays, respectively. Cell cycle analysis was carried out microscopically, and by propidium iodide uptake using flow cytometry. Sterol induction of oncogenic gene expression was evaluated by RT-PCR. Apoptotic cells were identified by immunocytochemistry using DNA fragmentation method, and by annexin V adhesion using flow cytometry. Results: Physiological doses (16 μM) of these sterols were not cytotoxic in these cells. Cholesterol-enrichment promoted mitosis (54 and 61% by microscopy; 40.8 and 34.08% by FACS analysis in PC-3 and DU145, respectively) and cell growth (P < 0.05), while phytosterols suppressed mitosis (29 and 35% by microscopy; 27.71 and 17.37% by FACS analysis in PC-3 and DU145, respectively), and significantly induced tumor-suppression (P < 0.05) and apoptosis. We demonstrated for the first time that cholesterols upregulated the expression of PCGEM1 even in androgen-insensitive prostate cancer cell lines. Phytosterols reversed this effect, while upregulating the expression of caveolin-1, a known mediator of androgen-dependent proto-oncogene signals that presumably control growth and anti-apoptosis. Conclusions: Phytosterol inhibition of PCGEM1 and cell growth and the overexpression of caveolin-1, suggests that poor disease prognosis anchors on the ability of caveolin-1 to regulate downstream oncogene(s) and apoptosis genes. Sterol intake may contribute to the disparity in incidence of prostate cancer, and elucidation of the mechanism for modulation of growth and apoptosis signaling may reveal potential targets for cancer prevention and/or chemotherapeutic intervention. Sterol regulation of PCGEM1 expression suggests its potential as biomarker for prediction of neoplasms that would be responsive to chemoprevention by phytosterols.