Sex affects the response of Wistar rats to polyvinyl pyrrolidone (PVP)-coated silver nanoparticles in an oral 28 days repeated dose toxicity study.

BACKGROUND: Silver nanoparticles (AgNPs) are widely used in biomedicine due to their strong antimicrobial, antifungal, and antiviral activities. Concerns about their possible negative impacts on human and environmental health directed many researchers towards the assessment of the safety and toxicity of AgNPs in both in vitro and in vivo settings. A growing body of scientific information confirms that the biodistribution of AgNPs and their toxic effects vary depending on the particle size, coating, and dose as well as on the route of administration and duration of exposure. This study aimed to clarify the sex-related differences in the outcomes of oral 28 days repeated dose exposure to AgNPs. METHODS: Wistar rats of both sexes were gavaged daily using low doses (0.1 and 1 mg Ag/kg b.w.) of polyvinylpyrrolidone (PVP)-coated small-sized (10 nm) AgNPs. After exposure, blood and organs of all rats were analysed through biodistribution and accumulation of Ag, whereas the state of the liver and kidneys was evaluated by the levels of reactive oxygen species (ROS) and glutathione (GSH), catalase (CAT) activity, superoxide dismutase (SOD) and glutathione peroxidase (GPx), expression of metallothionein (Mt) genes and levels of Mt proteins. RESULTS: In all animals, changes in oxidative stress markers and blood parameters were observed indicating the toxicity of AgNPs applied orally even at low doses. Sex-related differences were noticed in all assessed parameters. While female rats eliminated AgNPs from the liver and kidneys more efficiently than males when treated with low doses, the opposite was observed for animals treated with higher doses of AgNPs. Female Wistar rats exposed to 1 mg PVP-coated AgNPs/kg b.w. accumulated two to three times more silver in the blood, liver, kidney and hearth than males, while the accumulation in most organs of digestive tract was more than ten times higher compared to males. Oxidative stress responses in the organs of males, except the liver of males treated with high doses, were less intense than in the organs of females. However, both Mt genes and Mt protein expression were significantly reduced after treatment in the liver and kidneys of males, while they remained unchanged in females. CONCLUSIONS: Observed toxicity effects of AgNPs in Wistar rats revealed sex-related differences in response to an oral 28 days repeated exposure.

Comparison of in vitro toxicity of silver ions and silver nanoparticles on human hepatoma cells.

Scientific information on the potential harmful effects of silver nanoparticles (AgNPs) on human health severely lags behind their exponentially growing applications in consumer products. In assessing the toxic risk of AgNP usage, liver, as a detoxifying organ, is particularly important. The aim of this study was to explore the toxicity mechanisms of nano and ionic forms of silver on human hepatoblastoma (HepG2) cells. The results showed that silver ions and citrate-coated AgNPs reduced cell viability in a dose-dependent manner. The IC50 values of silver ions and citrate-coated AgNPs were 0.5 and 50 mg L(-1) , respectively. The LDH leakage and inhibition of albumin synthesis, along with decreased ALT activity, indicated that treatment with either AgNP or Ag ions resulted in membrane damage and reduced the cell function of human liver cells. Evaluation of oxidative stress markers demonstrating depletion of GSH, increased ROS production, and increased SOD activity, indicated that oxidative stress might contribute to the toxicity effects of nano and ionic forms of silver. The observed toxic effect of AgNP on HepG2 cells was substantially weaker than that caused by ionic silver, while the uptake of nano and ionic forms of silver by HepG2 cells was nearly the same. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 679-692, 2016.

Food Safety is an Important Public Health Issue: Chloramphenicol Residues Determination by Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS) in honey.

BACKGROUND: Honey is used for nutritional, medicinal and industrial purposes and antibiotic residues may harm its quality and constitute a danger to human health. The broad spectrum antibiotic chloramphenicol (CAP) was used for curative purposes in veterinary medicine, but is now forbidden in European Union (EU) because of its many serious side effects (e.g. aplastic anaemia, grey syndrome, severe bone marrow depression and hypersensitivity). The aim of this study was to facilitate analyses of the quality and safety of Croatian honey distributed to whole European Union market; an assessment that has not previously been made. SUBJECTS AND METHODS: CAP in honey was qualifying and quantifying by validated liquid chromatography tandem mass spectrometry with negative electrospray ionisation method (LC-MS/MS). The target antibiotic was separated on chromatographic column Zorbax SB C18 (150 mm x 2.1 mm, 3.5 µm) with a gradient elution using acetonitrile - 0.1% formic acid mobile phase at a flow rate of 0.3 mL/min, with column temperature 35°C for CAP and 5D-CAP as internal standard. Homogenised honey samples were diluted with acetate buffer solution and extracted on Oasis Hydrophilic-Lipophilic-Balanced (HLB) sorbents. The method was used to analyse 280 domestic honey samples collected throughout Croatia between 2005.-2013. RESULTS: Recoveries of the method for real (acacia, chestnut, linden and flower) honey samples were 102% with RSD 8.4%. The value CCα and CCß were 0.09 and 0.12 µg/kg, respectively. Results showed only three subsequent positive detections (1.1%) of CAP in honey. CONCLUSIONS: Analysed honey samples from Croatia showed good quality and safety what is the one of the main objective in consumer health policy in EU.

Coffee Silver Skin-Health Safety, Nutritional Value, and Microwave Extraction of Proteins.

The aim of this research was to evaluate the health safety (concentrations of pesticide residues and heavy metals) and nutritional parameters (macro- and microminerals and crude fibre) of coffee silver skin (CS), as well to isolate proteins from this by-product using an optimised microwave extraction method. The CS by-product samples showed the highest amount of potassium, followed by calcium, magnesium, and sodium. Iron was found in the highest quantity among the microminerals, followed by copper, manganese, zinc, and chromium. The CS sample showed a large amount of fibre and a moderate quantity of proteins obtained by the optimised microwave extraction method. Four heavy metals (nickel, lead, arsenic, and cadmium) were detected, and all were under the permitted levels. Among the 265 analysed pesticides, only three showed small quantity. The results for the proteins extracted by microwave showed that the total protein concentration values ranged from 0.52 ± 0.01 mg/L to 0.77 ± 0.07 mg/L. The highest value of the concentration of total proteins (0.77 ± 0.07 mg/L) was found in the sample treated for 9 min, using a power of 200 W. Based on these results, it can be concluded that CS is a healthy and nutritionally rich nutraceutical that could be used in the production of new products in the food industry and other industries.

Knowledge and Attitudes of Cannabidiol in Croatia among Students, Physicians, and Pharmacists.

Due to cannabidiol's health benefits and absence of serious side effects, its use is constantly growing. This is a survey-based cross-sectional study that was conducted to determine Croatian pharmacists', physicians', and students' knowledge and attitudes about cannabidiol (CBD). Two questionnaires were created, one for students and the other for physicians and pharmacists. Our participants (in total 874: 473 students and 401 physicians and pharmacists) generally had positive attitudes towards CBD therapy as approximately 60% of them believe that CBD treatment is generally efficacious. Participants had positive attitudes toward the therapeutic value of CBD, especially pharmacists and pharmacy students (63.8% and 72.2%, respectively). Pharmacists were significantly more convinced that CBD could reduce the use of opioids prescribed for chronic pain (p < 0.05). Only 17.5% of students had read scientific papers about CBD, compared to a significantly higher percentage of physicians and pharmacists (43.0% and 47.8%, respectively) (p < 0.05). This study revealed a gap in knowledge regarding CBD, since 89.3% of pharmacists and physicians, as well as 84.8% of students, believe they need more education about CBD. We conclude that it is important to improve the educational curricula so that medical professionals can recommend CBD use to their patients when needed.

Effect of natural biostimulants on yield and nutritional quality: an example of sweet yellow pepper (Capsicum annuum L.) plants.

BACKGROUND: Modifications in growing techniques can affect the yield and nutritional quality of various cultivated plant species. Owing to its high nutritional value, pepper (Capsicum annuum L.) was used in this study as a model plant to investigate the effect of natural biostimulants on yield and fruit quality parameters under conditions of reduced fertilisation. RESULTS: A positive influence of biostimulant treatment on yield parameters was observed. The overall increase in the pigment content of leaves after biostimulant application agreed well with the higher total and commercial yields of treated pepper cultivars compared with their controls. The results showed that natural biostimulants had a positive effect on the vitamin C and total phenolic contents in pepper fruits during the hot summer season. The 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonate) (ABTS) antioxidant activities were also significantly higher (P < 0.05) in treated plants and correlated strongly with all measured quality parameters except total phenolic content. CONCLUSION: Generally, biostimulants improved the antioxidant activity, vitamin C and phenolic contents in fruits as well as the pigment content in leaves of treated compared with non-treated pepper plants grown hydroponically. Thus the application of biostimulants could be considered as a good production strategy for obtaining high yields of nutritionally valuable vegetables with lower impact on the environment.

The controversies of parabens - an overview nowadays.

Effects of paraben toxicity, i.e., endocrine-disruption properties, are in the focus of researchers for decades, but still - they are a hot subject of debate. Parabens are aliphatic esters of p-hydroxybenzoic acid, which are widely used as antimicrobial agents for the preservation of cosmetics, pharmaceuticals and foods. Mostly used parabens are methyl-, ethyl-, propyl- and butylparaben. Although the toxicity of parabens is reported in animals and in in vitro studies, it cannot be taken for granted when discussing hazards for human health due to an unrealistic exposure -safety profile. Many studies have demonstrated that parabens are non-teratogenic, non-mutagenic, non-carcinogenic and the real evidence for their toxicity in humans has not been established. For now, methyl-, ethyl- and propylparaben are considered safe for use in cosmetics and pharmaceuticals within the recommended range of doses. Regarding alternatives for parabens, a variety of approaches have been proposed, but every substitute would need to be tested rigorously for toxicity and safety.

Food-Drug Interactions with Fruit Juices.

Fruit juices contain a large number of phytochemicals that, in combination with certain drugs, can cause food-drug interactions that can be clinically significant and lead to adverse events. The mechanisms behind such interactions are in most cases related to phytochemical interference with the activity of cytochrome P450 metabolizing enzymes (CYPs) or drug transporters. Moreover, alterations in their activity can have a clinical relevance if systemic exposure to the drug is decreased or increased, meaning that the pharmacological drug effects are suboptimal, or the drug will cause toxicity. In general, the common pharmacokinetic parameters found to be altered in food-drug interactions regarding fruit juices are the area under the concentration-time curve, bioavailability, and maximum plasma concentration. In most cases, the results from the drug interaction studies with fruit juices provide only limited information due to the small number of subjects, which are also healthy volunteers. Moreover, drug interactions with fruit juices are challenging to predict due to the unknown amounts of the specific phytochemicals responsible for the interaction, as well as due to the inter-individual variability of drug metabolism, among others. Therefore, this work aims to raise awareness about possible pharmacological interactions with fruit juices.

Safety of Probiotics: Functional Fruit Beverages and Nutraceuticals.

Over the last decade, fruit juice consumption has increased. Their rise in popularity can be attributed to the belief that they are a quick way to consuming a dietary portion of fruit. Probiotics added to fruit juices produce various bioactive compounds, thus probiotic fruit juices can be considered as a new type of functional foods. Such combinations could improve nutritional properties and provide health benefits of fruit juices, due to delivering positive health attributes from both sources (fruit juices and probiotics). However, this review discusses the other side of the same coin, i.e., the one that challenges general beliefs that probiotics are undoubtedly safe. This topic deserves more acknowledgments from the medical and nutritional literature, as it is highly important for health care professionals and nutritionists who must be aware of potential probiotic issues. Still, clinical trials have not adequately questioned the safety of probiotics, as they are generally considered safe. Therefore, this reviews aims to give an evidence-based perspective of probiotic safety, focusing on probiotic fruit beverages and nutraceuticals, by providing documented clinical case reports and studies. Finally, the paper deals with some additional insights from the pharmacological and toxicological point of views, such as pharmacological repercussions of probiotics on health.

Innovative Hurdle Technologies for the Preservation of Functional Fruit Juices.

Functional nutrition, which includes the consumption of fruit juices, has become the field of interest for those seeking a healthy lifestyle. Functional nutrition is also of great interest to the food industry, with the aims of improving human health and providing economic prosperity in a sustainable manner. The functional food sector is the most profitable part of the food industry, with a fast-growing market resulting from new sociodemographic trends (e.g., longer life expectancy, higher standard of living, better health care), which often includes sustainable concepts of food production. Therefore, the demand for hurdle technology in the food industry is growing, along with the consumption of minimally processed foods, not only because this approach inactivates microorganisms in food, but because it can also prolong the shelf life of food products. To preserve food products such as fruit juices, the hurdle technology approach often uses non-thermal methods as alternatives to pasteurization, which can cause a decrease in the nutritional value and quality of the food. Non-thermal technologies are often combined with different hurdles, such as antimicrobial additives, thermal treatment, and ultraviolet or pulsed light, to achieve synergistic effects and overall quality improvements in (functional) juices. Hence, hurdle technology could be a promising approach for the preservation of fruit juices due to its efficiency and low impact on juice quality and characteristics, although all processing parameters still require optimization.

Malonaldehyde and erythrocyte antioxidant status in children with controlled asthma.

In the pathogenesis of asthma, oxidative stress appears to play an important role and existence of an oxidant/antioxidant imbalance is evident. In this study the key markers of oxidative stress and lipid peroxidation in the pathogenesis of asthma in childhood in comparison to healthy subjects were investigated. Plasma marker of the lipid peroxidation: malondialdehyde (MDA), the erythrocytes antioxidative enzymes: glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), glutathione reductase (GR) and cysteine-containing tripeptide glutathione (GSH) were evaluated by spectrophotometric methods using blood samples collected from 37 healthy children and 44 asthmatic patients. The GSH-Px activity was significantly lower in asthmatic children (3.99 +/- 1.0 IU/g Hb) than in healthy controls (4.61 +/- 1.3 IU/g Hb; p < 0.034). Significant difference in activity of the SOD, GR, and concentration of cysteine-containing tripeptide GSH was not confirmed (p > 0.05). Lower GSH-Px activity in children with controlled asthma showed deficient erythrocyte antioxidant defence and evidence of association between oxidative stress and asthma in childhood. Preserved activity of GR and SOD, together with concentration of GSH and MDA, still seems to be crucial in controlling antioxidant/oxidant balance of the disease.

Phenolic and Antioxidant Analysis of Olive Leaves Extracts (Olea europaea L.) Obtained by High Voltage Electrical Discharges (HVED).

BACKGROUND: The aim of this study was to evaluate high voltage electrical discharges (HVED) as a green technology, in order to establish the effectiveness of phenolic extraction from olive leaves against conventional extraction (CE). HVED parameters included different green solvents (water, ethanol), treatment times (3 and 9 min), gases (nitrogen, argon), and voltages (15, 20, 25 kV). METHODS: Phenolic compounds were characterized by ultra-performance liquid chromatography-tandem mass spectrometer (UPLC-MS/MS), while antioxidant potency (total phenolic content and antioxidant capacity) were monitored spectrophotometrically. Data for Near infrared spectroscopy (NIR) spectroscopy, colorimetry, zeta potential, particle size, and conductivity were also reported. RESULTS: The highest yield of phenolic compounds was obtained for the sample treated with argon/9 min/20 kV/50% (3.2 times higher as compared to CE). Obtained results suggested the usage of HVED technology in simultaneous extraction and nanoformulation, and production of stable emulsion systems. Antioxidant capacity (AOC) of obtained extracts showed no significant difference upon the HVED treatment. CONCLUSIONS: Ethanol with HVED destroys the linkage between phenolic compounds and components of the plant material to which they are bound. All extracts were compliant with legal requirements regarding content of contaminants, pesticide residues and toxic metals. In conclusion, HVED presents an excellent potential for phenolic compounds extraction for further use in functional food manufacturing.

Comparing the effects of thermal and non-thermal technologies on pomegranate juice quality: A review.

The consumption of pomegranate juice (PJ) has increased substantially since scientific literature reported its therapeutic benefits that are attributable to antioxidant, antimicrobial, anti-carcinogenic, and anti-inflammatory properties. The growing consumer demands for fresh and high-quality food products have led to the interest in non-thermal technologies for preservation of fruit juices. Although thermal pasteurization is the most widely used preservation technology, it has adverse effects on the nutritional and sensory quality of juices. Conversely, non-thermal technologies are suitable alternatives for preservation and without negative effects on the quality. However, there is limited scientific literature concerning the use of non-thermal technologies for preservation of PJ. Therefore, this review focuses on PJ preservation by using non-thermal technologies. In conclusion, pomegranate is an economical crop that can justify the use of advanced non-thermal technologies for PJ preservation, as consumers' interest can offset the expenses associated with investments in alternative technological options and processing adjustments.

Differences in the levels of pesticides, metals, sulphites and ochratoxin A between organically and conventionally produced wines.

Organic products are generally recognized to be healthier and safer than conventional products. However, the actual scientific data regarding the importance of organic production on particular contaminant/additive content of wines is scarce. This study aimed to evaluate contents of pesticides, metals, sulphites and ochratoxin A in organically (org) and conventionally (conv) produced wines from eleven locations in different winegrowing regions of Croatia. All wines contained significantly lower levels of residues as compared to the maximum limits (MLs) with the exception of excessive amounts of Cu and Zn in one sample. Pb and Mg were mainly significantly less represented in org wines. There were no significant differences in the content of sulphite or ochratoxin A between org and conv wines. Significantly lower total pesticide concentrations and average number of pesticides per sample were obtained in org wines. The majority of ochratoxin A positive wines were from conv wine producers.

Monitoring of selected pharmaceuticals in surface waters of Croatia.

Sulfonamides, macrolides, torasemide, fumagillin, and chloramphenicol were simultaneously analyzed in surface water samples by using solid-phase extraction (SPE) and reversed-phase (RP) liquid chromatography-electrospray tandem mass spectrometry (LC-ESI-MS/MS). In the pre-concentration and clean-up process, the pH value of samples and volume of the solvent for extraction of analytes from cartridge were optimized. Extraction recoveries were high with values in the range from 62 to 115%. Limits of quantification (LoQ) were in the range from 0.02 to 0.2 µg L-1. Repeatability of the method was evaluated at LoQ and expressed as relative standard deviation (RSD). Calculated RSDs were low with values in the range from 2.4 to 14.5%. The method was successfully applied for analysis of surface water real samples. Samples were collected along the rivers in Croatia on four sampling sites in 2012 in Danube catchment areas, 19 sampling sites in Danube and Adriatic catchment areas in 2013, and another 19 places in 2014. Altogether, 20 target compounds were analyzed in 148 water samples and detected in 31 samples in range (0.1-5.3) µg L-1 or in 20.1% of samples. The most frequent and highest concentrations were detected for macrolide antibiotics. This is the first attempt of such monitoring in surface waters in Croatia.

Qualitative and quantitative analysis of flavonoids of the strawberry tree - Arbutus unedo L. (Ericaceae).

The leaves and fruits of strawberry tree - Arbutus unedo L., collected from two separate geographic locations in Croatia were investigated to determine their flavonoid composition and content. Quercitrin, isoquercitrin, hyperoside and rutin were identified in all leaf samples by means of thin-layer chromatography; the fruits contained only isoquercitrin. Chlorogenic acid was present in some leaf samples. The content of flavonoids depended on the plant organ investigated, date of collection and the locality. Spectrophotometric determination of the flavonoids indicated that the leaves are richer in flavonoids (0.52-2.00%) than fruits (0.10-0.29%).

Association of methylenetetrahydrofolate (MTHFR) and apolipoprotein E (apo E) genotypes with homocysteine, vitamin and lipid levels in carotid stenosis.

The aim of the study was to investigate the association between methylenetetrahydrofolate (MTHFR) genotypes and levels of homocysteine (Hcy), folate, vitamin B12 and lipids as well as the association between apolipoprotein E (apo E) genotypes and levels of lipids in a Croatian healthy control group and a group of patients with > 70% carotid stenosis (CS). The study included 98 Croats, 38 patients with > 70% carotid stenosis and 60 age- and sex-matched controls. The MTHFR and apo E genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), Hcy by enzyme immunoassay, vitamins by immunochemiluminiscence, and lipids by spectrophotometric method. There was no difference between control subjects and CS patients in the distribution of C677T MTHFR genotypes (p=0. 786) and alleles (p=0.904), however, differences in the frequencies of apo E genotypes (p=0.012) and alleles (p=0.029) were statistically significant. The odds ratio for apo E 3/4 genotype was 3.93 (95% CI 1.23-12.61). Hyperhomocysteinemia (> or =15 micromol/L) was found in 11% of CS patients and 5% of control subjects. Total cholesterol, triglycerides, vitamin B12 and folate were statistically different in "all MTHFR genotypes" (p<0.001, p<0.01, p=0.044 and p=0.036, respectively), and in TC/TT (p<0.001, p=0.003, p=0.030 and p=0.032, respectively) groups. The levels of total cholesterol, LDL cholesterol and triglycerides in the apo E 3/3, and total cholesterol in the apo E 3/4 group yielded statistical difference. An association was found of apo E 3/4 genotype but not of MTHFR genotypes with the risk of CS. MTHFR and apo E affect blood lipid levels, which was statistically confirmed. An association was also recorded between hyperhomocysteinemia and patients with CS. Vitamin status in CS showed a statistically verified association with TC/TT MTHFR genotype. In the group of patients with TC/TT MTHFR genotype, lower vitamin B12 and higher folate values were recorded. The results of multiple logistic analysis showed that there was no statistical significance of Hcy levels (OR 2.403, p=0.334) or conventional vascular risk factors such as smoking habit (OR 0.505, p=0.149), age (OR 1.048, p=0.087) or sex (OR 2.037, p=0.112) in predicting CS.

Response of biochemical biomarkers in the aquatic crustacean Daphnia magna exposed to silver nanoparticles.

The proliferation of silver nanoparticle (AgNP) production and use owing to their antimicrobial properties justifies the need to examine the resulting environmental impacts. The discharge of biocidal nanoparticles to water bodies may pose a threat to aquatic species. This study evaluated the effects of citrate-coated AgNPs on the standardized test organism Daphnia magna Straus clone MBP996 by means of biochemical biomarker response. AgNP toxicity was compared against the toxic effect of Ag(+). The toxicity endpoints were calculated based upon measured Ag concentrations in exposure media. For AgNPs, the NOAEC and LOAEC values at 48 h were 5 and 7 µg Ag/L, respectively, while these values were 0.5 and 1 µg Ag/L, respectively, for Ag(+). The EC50 at 48 h was computed to be 12.4 ± 0.6 and 2.6 ± 0.1 µg Ag/L for AgNPs and Ag(+), respectively, with 95 % confidence intervals of 12.1-12.8 and 2.3-2.8 µg Ag/L, respectively. These results indicate significant less toxicity of AgNP compared to free Ag(+) ions. Five biomarkers were evaluated in Daphnia magna neonates after acute exposure to Ag(+) or AgNPs, including glutathione (GSH) level, reactive oxygen species (ROS) content, and catalase (CAT), acetylcholinesterase (AChE), and superoxide dismutase (SOD) activity. AgNPs induced toxicity and oxidative stress responses in D. magna neonates at tenfold higher concentrations than Ag. Biochemical methods revealed a clear increase in AChE activity, decreased ROS level, increased GSH level and CAT activity, but no significant changes in SOD activity. As Ag(+) may dissolve from AgNPs, these two types of Ag could act synergistically and produce a greater toxic response. The observed remarkably high toxicity of AgNPs (in the parts-per-billion range) to crustaceans indicates that these organisms are a vulnerable link in the aquatic food chain with regard to contamination by nanosilver. Graphical Abstract ᅟ.

Croatian population data for the C677T polymorphism in methylenetetrahydrofolate reductase: frequencies in healthy and atherosclerotic study groups.

BACKGROUND: The aim of this study was to investigate the frequency of C677T methylenetetrahydrofolate reductase (MTHFR) mutation in healthy Croatian volunteers and in patients with atherosclerosis. METHODS: The C677T MTHFR gene mutation was determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in 640 subjects, residents of the Zagreb city or Zagreb surroundings. Control group (n=298) was healthy blood donors. Patients (n=342) were divided into two groups of those with coronary heart disease, CAD (n=247) and those with >60% carotid stenosis, CS (n=95). RESULTS: CC genotype was recorded in 45% of healthy volunteers and 46% of patients (46.3% with CS and 46.2% with CAD). TC genotype was found in 49% of healthy volunteers and 45% of patients (46.3% with CS and 44.9% with CAD). There was no significant difference (p>0.05) from the control group in the genotype or allele frequency either for the overall group of patients with atherosclerosis or for the patient subgroups. CONCLUSION: The preliminary study of MTHFR polymorphism in control subjects and cardiovascular disease/carotid stenosis patients revealed that in Croats there was a low frequency of TT genotype (6% in controls vs. 9% in patients) and T allele (31% for cases and controls). Additionally, our results did not show significantly higher frequency of MTHFR mutation in CAD and CS studied groups.

Paraoxonase activity and concentration of indicators of lipids status in the serum of cardiac patients.

In this research, we measured the activity of paraoxonase (basal and activated) enzyme, and components of lipid status components (total cholesterol, LDL cholesterol, HDL cholesterol and Apo A I) in the serum of patients, undergoing bypass surgery. We also tested how the applied EKC affected changes of defined indicators. Measuring of all the given parameters was conducted prior to the operation, 90 minutes, 1.5 hour, 6 hours, 24 hours and 72 hours, on 29 patients (11 of them did undergo myocardium revascularization with the application of EKC, while the rest of them did not). Activity of paraoxonase (both basal and activated) changes significantly during the postoperative period, in relation to pre-operative values, p < 0.05. Total cholesterol concentration is reduced in both examined groups, regardless of the application of EKC. This trend is also accompanied by LDL cholesterol concentration. On the other hand, HDL cholesterol concentration during post-operative period does not indicate any significant statistical change in relation to pre-operative values, while we noticed difference with regard to EKC application, 90 minutes after surgery. This change of lipid status indicator is partly due to heparin, a stimulator of lipoprotein lipase that was applied during the surgery. Our conclusion is that lipid profile changes significantly after the bypass surgery, mostly regardless of the application of EKC.