Placental transport of parabens studied using an ex-vivo human perfusion model.

INTRODUCTION: Parabens are a group of chemicals widely used as preservatives in daily consumer products such as cosmetics, food items, pharmaceuticals and household commodities. They have been broadly detected in human samples indicating a general human exposure, and concerns arose from their potential endocrine disrupting effect. Especially the exposure to parabens during pregnancy is concerning, as the time of fetal development is a particularly vulnerable period. The aim of this study was to investigate the transport and metabolism of four commonly used parabens: methyl-, ethyl-, propyl- and butylparaben (MeP, EtP, PrP and BuP) and the metabolite para-hydroxybenzoic acid (PHBA) across the human placenta. METHODS: An ex-vivo human placental perfusion model was used. The test compounds were added in the maternal compartment (with initial concentrations of 1 mM or 0.1 mM). Placental transport was evaluated by fetal-maternal concentration ratios (FM-ratio), transport index (TI) and indicative permeability (IP). RESULTS: Information about parabens kinetics was taken from 10 perfusions and PHBA from 7 perfusions. Paraben metabolism was not detected. The placental transport of MeP, EtP, PrP, BuP and PHBA revealed a transfer from maternal to fetal circulations with FM120 of 0.86 ± 0.27 (MeP), 0.98 ± 0.28 (EtP), 1.00 ± 0.28 (PrP), 1.12 ± 0.59 (BuP) and 0.82 ± 0.37 (PHBA). The test substances accumulated in the perfused tissue in some degree. The average kinetic parameters FM-ratio, TI and IP were not different between chemicals. DISCUSSION: The present study shows that the placenta barrier is permeable to all four parabens and the metabolite, which implies potential fetal exposure.

Pharmacokinetic characterization of carnosol from rosemary (Salvia Rosmarinus) in male C57BL/6 mice and inhibition profile in human cytochrome P450 enzymes.

Rosemary (Salvia Rosmarinus) is a rich source of dietary diterpenes with carnosol as one of the major polyphenols used to standardize rosemary extracts approved as a food preservative, however, at present there is not any information on the murine pharmacokinetic profile of carnosol or its potential for drug interactions. The present study utilizes cell-free, cell-based, and animal-based experiments to define the pharmacokinetic profile of the food based phytochemical carnosol. Mice were administered carnosol (100 mg/kg body weight) by oral gavage and plasma levels were analyzed by LC-MS/MS to establish a detailed pharmacokinetic profile. The maximum plasma concentration exceeded 1 µM after a single administration. The results are significant as they offer insights on the potential for food-drug interactions between carnosol from rosemary and active pharmaceutical ingredients. Carnosol was observed to inhibit selected CYP450 enzymes and modulate metabolic enzymes and transporters in in vitro assays.

Synergistic chemopreventive effect of allyl isothiocyanate and sulforaphane on non-small cell lung carcinoma cells.

Isothiocyanates from cruciferous vegetables are known for their potential anti-carcinogenic activities. These isothiocyanates are frequently consumed together as part of a regular diet, but their combined effects on carcinogenesis have not been well studied. Herein, we tested the hypothesis that combination of two isothiocyanates, i.e. allyl isothiocyanate and sulforaphane, produced a synergy in inhibiting the growth of A549 lung cancer cells. Our results showed that the combination treatment led to a stronger growth inhibition than the singular treatment. Isobologram analysis proved that the enhanced inhibitory effect of the combination treatment was synergistic. Flow cytometry demonstrated that the combination treatment caused more extensive cell cycle arrest and apoptosis than the singular treatment with modified expression of key proteins regulating these cellular processes. The combined treatment resulted in the production of intracellular reactive oxygen species, which might contribute to the inhibitory effects on cancer cells. Moreover, a synergy between allyl isothiocyanate and sulforaphane was also observed in anti-cell migration. Collectively, our results have demonstrated the potential of different isothiocyanates used in combination to produce enhanced protective effects against carcinogenesis.

Phenolic Acids Derivatives - Biotechnological Methods of Synthesis and Bioactivity.

Phenolic acids are secondary plant metabolites belonging to polyphenol classes, widely spread throughout the plant kingdom. The name "phenolic acids", in general, describes phenols that possess one carboxylic acid functionality and they could be divided into three major subclasses: hydroxybenzoic, hydroxycinnamic and hydroxyphenylacetic acids. The great interest in phenolic acids is associated with their high potential for food preservation (antioxidant and antimicrobial activity) and, last but not least, high therapeutic potential. The aim of this review is to summarise the current knowledge concerning phenolic acids and the topics discussed include natural sources of phenolic acids, biosynthesis and metabolism, health benefits of phenolic acids (first of all the antioxidant activity of dietary phenolics and also anticarcinogenic and anti-inflammatory effect), their antimicrobial activity for selected groups of bacteria and fungi (yeasts) and structural modifications of the molecule especially those increasing lipophilicity and improving solubility in lipids. Recent advances in the methods of enzymatic synthesis of phenolic acid derivatives are described and some conclusions referring to Structure-Property-Activity Relationships of phenolic acids, particularly important from the point of view of their pharmaceutical and nutritional applications, are discussed.

The tissue residues of sodium dehydroacetate used as feed preservative in swine.

BACKGROUND: Sodium dehydroacetate (Na-DHA) is a food and feed additive with antimicrobial effects. There is little information on Na-DHA residue levels in foods derived from animals. In this study, Na-DHA residue levels in swine tissues were determined by HLPC, and the pharmacokinetics of Na-DHA in tissues were determined. RESULTS: The Na-DHA residue levels in swine tissues were <1.2 mg kg-1 at different withdrawal time after thirty-two Duroc × Landrace × Yorkshire pigs were administered 200 mg Na-DHA kg-1 through the feed for 30 days. In decreasing order of Na-DHA residue levels, the tissues were kidney > liver > muscle > fat. The pharmacokinetics of Na-DHA followed a binomial regression model, and the half-time of Na-DHA in swine tissues was 9.07 days for kidney, 7.19 days for liver, 6.66 days for muscle, and 5.39 days for fat tissue. The accuracy of the HPLC method for Na-DHA determination ranged from 80.18% to 91.33% recovery, with coefficients of variation <6.4%, limit of detection of 0.08 mg kg-1 , and limit of quantification of 0.2 mg kg-1 . CONCLUSION: Na-DHA included at 200 mg kg-1 in a swine diet is a safe feed additive based on residue elimination and ADI values reported. © 2017 Society of Chemical Industry.

p-Hydroxybenzoate esters metabolism in MCF7 breast cancer cells.

Parabens are among the most frequently used preservatives to inhibit microbial growth and extend the shelf life of a range of consumer products. The objective of the present study was to gain insight into the metabolism of parabens in breast cancer cells (MCF7) since they have demonstrated estrogenic activity towards these cells and have been detected in breast cancer tissues. The toxicity of parabens to MCF7 cells was determined using MTT assays. Hydrolysis of methyl-, butyl and benzyl-paraben to p-hydroxybenzoic acid was analyzed in cultured MCF7 cells and in cellular homogenates. Glucuronidation and sulfoconjugation were studied in MCF7 homogenates, and parabens were analyzed by HPLC. Methyl-paraben was shown to be far less toxic than butyl and benzyl-paraben. Parabens were completely stable in MCF7 homogenates whereas p-nitrophenyl acetate, a substrate type, underwent hydrolysis. MCF7 cell homogenates did not express glucuronidation and sulfoconjugation activities toward parabens. The higher stability of parabens may explain their accumulation in breast cancer tissue as previously reported in the literature.

Investigations on the metabolism and potentially adverse effects of ethoxyquin dimer, a major metabolite of the synthetic antioxidant ethoxyquin in salmon muscle.

The feed additive ethoxyquin (EQ) is a commonly used synthetic antioxidant preservative in animal feeds. In farmed Atlantic salmon fillets, EQ residues are present, both as the parent compound and as EQ derivatives. One of the main EQ derivates in fish muscle is an ethoxyquin dimer (EQDM), and the potential toxicity of this metabolite is not known. The aim of this study was to evaluate the metabolism and potentially toxicological effects of EQDM. A 90-day subchronic exposure study with repeated dietary exposure to EQDM at 12.5 mg/kg of body weight per day was performed with male F344 rats. Hepatic Cyp1a1 mRNA was significantly reduced to <3% of the control in rats fed EQDM, and hepatic Cyp2b1 mRNA was increased to 192%. EQDM increased Gstpi1 mRNA expression to 144% that of the control, but the activity level of this phase II enzyme was reduced. Biomarkers of liver and kidney function did indicate adverse effects of EQDM when F344 rats were fed 12.5 mg/kg of body weight per day. The present study revealed that EQDM produces responses that are comparable to those produced by the parent compound (EQ) in terms of activating the same enzyme systems.

Inhibition of bladder cancer development by allyl isothiocyanate.

Bladder cancer is one of the common human cancers and also has a very high recurrence rate. There is a great need for agents capable of inhibiting bladder cancer development and recurrence. Here, we report that allyl isothiocyanate (AITC), an ingredient of many common cruciferous vegetables, potently inhibited the proliferation of bladder carcinoma cell lines in vitro [half maximal inhibitory concentration (IC(50)) of 2.7-3.3 microM], which was associated with profound G(2)/M arrest and apoptosis. In contrast, AITC was markedly less toxic to normal human bladder epithelial cells (IC(50) of 69.4 microM). AITC was then evaluated in two rat bladder cancer models in vivo (an orthotopic model and a subcutaneous model). The orthotopic model closely mimics human bladder cancer development and recurrence. We show that a low oral dose of AITC (1 mg/kg) significantly inhibited the development and muscle invasion of the orthotopic bladder cancers but was ineffective against the subcutaneous xenografts of the same cancer cells in the same animals. This differential effect was explained by our finding that urinary levels of AITC equivalent were two to three orders of magnitude higher than that in the plasma and that its levels in the orthotopic cancer tissues were also three orders of magnitude higher than that in the subcutaneous cancer tissues. Moreover, we show that AITC is a multi-targeted agent against bladder cancer. In conclusion, AITC is selectively delivered to bladder cancer tissue through urinary excretion and potently inhibits bladder cancer development and invasion.

Refining aggregate exposure: example using parabens.

The need to understand and estimate quantitatively the aggregate exposure to ingredients used broadly in a variety of product types continues to grow. Currently aggregate exposure is most commonly estimated by using a very simplistic approach of adding or summing the exposures from all the individual product types in which the chemical is used. However, the more broadly the ingredient is used in related consumer products, the more likely this summation will result in an unrealistic estimate of exposure because individuals in the population vary in their patterns of product use including co-use and non-use. Furthermore the ingredient may not be used in all products of a given type. An approach is described for refining this aggregate exposure using data on (1) co-use and non-use patterns of product use, (2) extent of products in which the ingredient is used and (3) dermal penetration and metabolism. This approach and the relative refinement in the aggregate exposure from incorporating these data is illustrated using methyl, n-propyl, n-butyl and ethyl parabens, the most widely used preservative system in personal care and cosmetic products. When these refining factors were used, the aggregate exposure compared to the simple addition approach was reduced by 51%, 58%, 90% and 92% for methyl, n-propyl, n-butyl and ethyl parabens, respectively. Since biomonitoring integrates all sources and routes of exposure, the estimates using this approach were compared to available paraben biomonitoring data. Comparison to the 95th percentile of these data showed that these refined estimates were still conservative by factors of 2-92. All of our refined estimates of aggregate exposure are less than the ADI of 10mg/kg/day for parabens.

Metabolism and pharmacokinetics of ethyl N(alpha)-lauroyl-L-arginate hydrochloride in human volunteers.

The human metabolism and pharmacokinetics of ethyl N(alpha)-lauroyl-L-arginate hydrochloride (LAE), a new antimicrobial agent for use in foods have been investigated using both in vitro and in vivo techniques with (14)C-LAE and (13)C-LAE respectively. LAE was readily hydrolysed to the corresponding lauroyl arginine (LAS) on incubation with human plasma samples to the extent of about 50% during 4h. LAE was stable in simulated gastric fluid but in simulated intestinal fluid it was rapidly hydrolysed to LAS and arginine with more than 90% conversion to arginine after 1h. Oral doses of (13)C-LAE in propylene glycol were administered to human volunteers at dose levels of 1.5mg/kg (4 subjects) and 2.5mg/kg (2 subjects). LAE was only detected in two plasma samples in one individual at the higher dose level close to the limit of quantification (1 ng/ml). Maximum plasma concentrations of LAS generally occurred at 2h with mean peak levels of 18.2 ng/ml (1.5mg/kg dose) and 23.9 ng/ml (2.5mg/kg dose). Maximum concentrations of (13)C-arginine occurred earlier (0.5 to 1h) and at much higher levels than LAS with mean peak levels of 124 ng/ml (1.5mg/kg dose) and 240 ng/ml (2.5mg/kg dose). The results showed that in humans LAE was rapidly metabolized to the naturally occurring dietary components lauric acid and arginine.

Chemoprotective and carcinogenic effects of tert-butylhydroquinone and its metabolites.

Tert-butylhydroquinone (tBHQ) has been commonly used as a synthetic food antioxidant to prevent oils and fats from oxidative deterioration and rancidity due to its potent anti-lipid peroxidation activity. In North America, the maximum level of tBHQ allowed in fat products is 0.02% with an acceptable daily intake of 0-0.7 mg/kg body weight. Extensive studies have demonstrated that tBHQ exhibit anti-carcinogenic effect. The ability of tBHQ to induce phase II xenobiotic metabolizing enzymes through an Nrf2-dependent pathway is thought to be responsible for the observed protective effect of tBHQ. It has been proposed that tBHQ enhances Nrf2-mediated transcription by promoting reactive oxygen species-mediated dissociation of Nrf2-Keap1, Nrf2 stabilization, phosphatidylinositol 3-kinase (PI3K)/Akt activity, and MAPK pathway activation. In contrast to the beneficial effects of tBHQ, a number of studies have shown that chronic exposure to tBHQ may induce carcinogenicity. However, the precise mechanisms of tBHQ carcinogenicity are not well understood. The toxicity or carcinogenicity of tBHQ has been attributed to the formation of reactive GSH-conjugates, generation of reactive species, CYP1A1 induction, caspase activation and reduced GSH/ATP levels. This review provides an account of recent mechanisms proposed for both chemoprotective and carcinogenic effect of tBHQ.

Safety assessment of aqueous olive pulp extract as an antioxidant or antimicrobial agent in foods.

The olive fruit, its oil and the leaves of the olive tree have a rich history of nutritional, medicinal and ceremonial uses. Olive oil, table olives and olive products are an important part of the Mediterranean diet, the greatest value of which may be due to olive polyphenols that contribute to the modulation of the oxidative balance in vivo. The objective of this review is to examine the available safety/toxicity literature on olive polyphenols, particularly hydroxytyrosol, to determine the safety-in-use of a standardized aqueous olive pulp extract (HIDROX). Among the polyphenols found in the extract, the major constituent of biological significance is hydroxytyrosol (50-70%). In oral bioavailability studies, urinary excretion of hydroxytyrosol and its glucuronide was found to be associated with the intake of hydroxytyrosol. Oral bioavailability of hydroxytyrosol in olive oil and in an aqueous solution was reported as 99% and 75%, respectively. In comparative studies, urinary excretion of hydroxytyrosol was greater in humans than in rats. The LD(50) of the extract and hydroxytyrosol was reported to be greater than 2000 mg/kg. In a subchronic study, the no observed adverse effect level (NOAEL) of the extract in rats was found to be 2000 mg/kg/day. In developmental and reproductive toxicity studies, HIDROX did not cause toxicity at levels up to 2000 mg/kg/day. In an in vivo micronucleus assay, oral exposure of rats to HIDROX at dose levels up to 5000 mg/kg/day for 29 days did not induce increases in polychromatic erythrocytes in bone marrow. Based on the available studies of the extract and polyphenols, and a history of exposure and use of components of the extract through table olives, olive products and olive oil, the consumption of HIDROX is considered safe at levels up to 20 mg/kg/day.

Safety assessment of propyl paraben: a review of the published literature.

Propyl paraben (CAS no. 94-13-3) is a stable, non-volatile compound used as an antimicrobial preservative in foods, drugs and cosmetics for over 50 years. It is an ester of p-hydroxybenzoate. Propyl paraben is readily absorbed via the gastrointestinal tract and dermis. It is hydrolyzed to p-hydroxybenzoic acid, conjugated and the conjugates are rapidly excreted in the urine. There is no evidence of accumulation. Acute toxicity studies in animals indicate that propyl paraben is relatively non-toxic by both oral and parenteral routes, although it is mildly irritating to the skin. Following chronic administration, no-observed-effect levels (NOEL) as high as 1200-4000 mg/kg have been reported and a no-observed-adverse-effect level (NOAEL) in the rat of 5500 mg/kg is posited. Propyl paraben is not carcinogenic, mutagenic or clastogenic. It is not cytogenic in vitro in the absence of carboxyesterase inhibitors. The mechanism of propyl paraben may be linked to mitochondrial failure dependent on induction of membrane permeability transition accompanied by the mitochondrial depolarization and depletion of cellular ATP through uncoupling of oxidative phosphorylation. Sensitization has occurred when medications containing parabens have been applied to damaged or broken skin. Parabens have been implicated in numerous cases of contact sensitivity associated with cutaneous exposure, but high concentrations of 5-15% in patch testing are needed to elicit reaction in susceptible individuals. Allergic reactions to ingested parabens have been reported, although rigorous evidence of the allergenicity of ingested paraben is lacking.

Malabsorption of modified food starch (acetylated distarch phosphate) in normal infants and in 8-24-month-old toddlers with non-specific diarrhea, as influenced by sorbitol and fructose.

UNLABELLED: Acetylated distarch phosphate (ADiSP) is a modified starch used in some baby foods. The bioavailability of ADiSP and a native (unmodified) starch was evaluated in 20 normal infants and 21 toddlers aged 8-24 mo with chronic non-specific diarrhea. Formulae contained 8% native or 8% modified waxy maize starch. No infant or toddler consuming Formula N (native starch) had elevated peak breath hydrogen levels (20 ppm or greater), stools clinically positive for reducing substances (0.75% or greater) or loose stools. Fourteen infants received formula M (modified starch): 2 had elevated breath hydrogen, 1 had positive stools and another had loose stools. Of the 21 toddlers fed formula M, 2 had elevated breath hydrogen, but none had positive stools or loose stools. Formula NS (native starch with 2% sorbitol) had little effect on breath hydrogen in the infants but significantly increased it in the toddlers. Formula NS produced loose stools in 2 toddlers but no clinically positive stools in any infant or toddler. Formula MS (modified starch with 2% sorbitol) elevated breath hydrogen in 3 infants and 8 toddlers, and produced positive stools in 2 infants and 2 toddlers, and loose stools in 4 infants and 7 toddlers. Formula MSF (modified starch with 2% sorbitol and 5% fructose) elevated breath hydrogen in 7 infants and 10 toddlers, positive stools in 7 infants and 6 toddlers, and loose stools or diarrhea in 7 infants and 11 toddlers. CONCLUSION: ADiSP modified starch can increase breath hydrogen and produce loose stools. Sorbitol and fructose aggravate the malabsorption, in some cases leading to frank diarrhea.

Chemopreventive effects of S-methylcysteine on rat hepatocarcinogenesis induced by concurrent administration of sodium nitrite and morpholine.

The aim of the present study was to examine the chemopreventive efficacy of S-methylcysteine (SMC) on rat hepatocarcinogenesis induced by concurrent administration of sodium nitrite (NaNO(2)) and morpholine (Mor) using a medium-term rat liver carcinogenesis bioassay (Ito test). Administration of SMC caused significant reduction in the areas of glutathione S-transferase placental form positive foci along with a significant decrease of hepatocyte 5-bromo-2'-deoxyuridine (BrdU) labeling indices. These results demonstrated potent chemopreventive effects of SMC against hepatocarcinogenesis due to concurrent administration of Mor and NaNO(2). SMC could thus be an effective chemopreventive agent for decreasing the risk of carcinogenicity from environmental precursors of N-nitroso compounds.

The disposition of allyl isothiocyanate in the rat and mouse.

The urine was the major route of excretion of radioactivity (50-80% of dose) following the oral administration (2.5 and 25 mg/kg body weight) of allyl[14C]isothiocyanate (AITC) to male and female Fischer 344 rats and B6C3F1 mice. Smaller amounts were found in the faeces (6-12%) and expired air (3-7%). The major difference between the two species was the greater retention of radioactivity after 4 days within rats (18-24% of dose) when compared with mice (2-5% of dose). Three radioactive components were found in the urine of mice and two in rats. The three components were inorganic thiocyanate, allylthiocarbamoylmercapturic acid and allylthiocarbamoylcysteine in mice, but no cysteine conjugate was found in rat urine. In the mouse, approximately 80% of the 14C was present in the urine as the thiocyanate ion whereas in the rat some 75% was as the mercapturate. This indicates that in the mouse, hydrolysis of AITC was the major metabolic pathway whereas in the rat glutathione conjugation was the major route. A species difference was seen in the amount of [14C]AITC-derived radioactivity present in the whole blood of rats and mice; measurable levels of radioactivity remained within rat blood for a longer time period (up to 240 hr) when compared with mice (96 hr). Examination of the urinary bladders of male and female rats following oral dosing with [14C]AITC showed a sex difference with greater amounts of [14C]AITC and/or its metabolites within the bladder tissue of male rats. This data is discussed in terms of the known species- and sex-specificity of the urinary bladder tumours, which occurred after long-term administration to male rats, but not to female rats or mice of either sex, in a carcinogenicity study conducted by the National Toxicology Program in the USA.

Use of nuclear magnetic resonance spectroscopy and selective C-labeling for pharmacokinetic research in man: detection of benzoic acid conversion to hippuric acid.

This paper demonstrates that the stable isotope tracer technique using NMR spectroscopy and the selective 13C labeling of protonated carbons can provide a relatively sensitive method to investigate pharmacokinetic problems in man. The urinary excreted [1,3,5-13C3]hippuric acid ([13C]HA) formed from orally administered [1,3,5-13C3] benzoic acid ([13C]BA) as a model substrate was successfully quantitated without any separation procedures by proton-decoupled 13C-NMR spectroscopy of 10-fold concentrated urine in a 10 min accumulation time. In spite of the low dosage (10mg BA), the C3,5 resonances of [13C]HA were detected with favorable signal-to-noise ratios to quantitate [13C]HA concentration. The administered [13C]BA was found to be quantitatively biotransformed to HA and excreted in urine within 4h. The lower limit of detection was estimated to be 50 nmol in an NMR tube, which was improved about one order of magnitude over that of BA labeled in the quaternary carbon (C7). The potential of an inverse detection experiment using heteronuclear multiple quantum coherence was also investigated in order to detect [13C]HA in urine, with a higher sensitivity. The inverse experiment improved the sensitivity by a factor of 2--3 over 13C¿1H¿-NMR, although the specificity of detection was relatively poor.