No abstract

No abstract

No abstract

No abstract

A probiotic supplement reduces the biologically effective dose of aflatoxin exposure and may thereby offer an effective dietary approach to decrease the risk of liver cancer.

The prolyl oligopeptidase (POP) family of serine proteases includes prolyl oligopeptidase, dipeptidyl peptidase IV, acylaminoacyl peptidase and oligopeptidase B. The enzymes of this family specifically hydrolyze oligopeptides with less than 30 amino acids. Many of the POP family enzymes have evoked pharmaceutical interest as they have roles in the regulation of peptide hormones and are involved in a variety of diseases such as dementia, trypanosomiasis and type 2 diabetes. In this study we have clarified the evolutionary relationships of these four POP family enzymes and analyzed POP sequences from different sources. The phylogenetic trees indicate that the four enzymes were present in the last common ancestor of all life forms and that the b-propeller domain has been part of the family for billions of years. There are striking differences in the mutation rates between the enzymes and POP was found to be the most conserved enzyme of this family. However, the localization of this enzyme has changed throughout evolution, as three archaeal POPs seem to be membrane bound and one third of the bacterial as well as two eukaryotic POPs were found to be secreted out of the cell. There are also considerable distinctions between the mutation rates of the different substrate binding subsites of POP. This information may help in the development of species-specific POP inhibitors.

In this study, the modulation of aflatoxin B 1 (AFB 1 ) uptake in rats by administration of the probiotic Lactobacillus rhamnosus GG was demonstrated. Fecal AFB 1 excretion in GG-treated rats was increased via bacterial AFB 1 binding. Furthermore, AFB 1 -associated growth faltering and liver injury were alleviated with GG treatment.Aflatoxins are toxic and carcinogenic fungal metabolites that frequently contaminate staple crops (1). Interventions in aflatoxin exposure focus either on improving crop quality and storage (20) or on altering aflatoxin bioavailability on the individual level, using various adsorbents (5, 18). However, no safe means are presently available to completely protect humans from aflatoxin exposure. Our previous work with lactic acid bacteria revealed that the probiotic strain Lactobacillus rhamnosus strain GG (ATCC 53013) efficiently binds several mycotoxins, including aflatoxin B 1 (AFB 1 ) and aflatoxin M 1 (AFM 1 ), its hydroxylated metabolite, in vitro (6,7,9,10,19). Even though heat-killed bacteria have the highest binding capacities, most studies use viable bacteria, which are more relevant in probiotic products intended for human consumption. Acid, intestinal enzymes, and intestinal mucus were shown not to interfere with AFB 1 binding of this probiotic (6,7,11,12). GG was subsequently demonstrated to bind AFB 1 and to reduce its uptake into intestinal tissue in the ligated duodenal loops of 1-week-old chicks (8). However, this ex vivo method has its limitations in simulating intestinal conditions, and we therefore conducted an in vivo single-dose experiment in rats, investigating the effects of GG on AFB 1 absorption and toxic effects. Five-week-old Han-Wistar rats were kept individually in metabolic cages on standard powdered feed and water ad libitum. After acclimatization, the rats (n ϭ 12/group) received either vehicle (phosphate-buffered saline [PBS]) or probiotic suspension (5 ϫ 10 10 CFU GG/0.5 ml PBS, prepared by directly suspending lyophilized bacteria in PBS) by oral gavage daily for 3 days before and 3 days after a single oral dose of AFB 1 (1.5 mg or 4.8 mol/kg of body weight in dimethyl sulfoxide [Sigma-Aldrich, St. Louis, Mo.]). Four additional rats served as untreated controls. Body weight was recorded at the beginning of the study (prior to GG treatment), on the day of AFB 1 dosing, and at the end of the study. Urine and fecal samples were collected daily, weighed, and stored at Ϫ20°C. At the end of the study, blood samples were taken by cardiac puncture and centrifuged, and the plasma was stored at Ϫ20°C. The experiments were approved by the University of Kuopio animal ethics committee.Extraction of fecal samples was modified from a method developed in our laboratory (17). The samples were mixed with 2.5 volumes of 0.2 M sodium acetate in 10% NaCl, and aliquots (2 ml) were spiked with aflatoxin G 2 (AFG 2 ) (18.6 pmol/sample) as an internal standard and centrifuged (3,000 ϫ g; 15 min; 4°C). The pellets were suspended in 4 ml 80% methanol (in 10% NaCl [vol/vol]) and h...

Viable, heat-and acid-killed Lactobacillus rhamnosus strain GG (LGG) has shown high binding properties with zearalenone (ZEN). To identify the type of chemical moieties and interactions involved in binding with the ZEN, LGG was subjected to different chemical and enzymatical treatments, prior to the binding experiments. Pretreating the viable, heat- and acid-killed bacteria with m-periodate significantly decreased ZEN binding, suggesting that ZEN binds predominantly to carbohydrate components. Pretreatment with Pronase E had no effect on the ability of viable cells to bind ZEN, however, a reduction in the binding of ZEN by heat- and acid-killed cells, suggesting that the new binding sites exposed by heat or acid are proteins in nature. Pretreatment with urea also decreased binding, suggesting that hydrophobic interactions play a role in ZEN binding. The binding of ZEN in concentrations ranging from 0.79 to 62.82 microM and its subsequent dissociation by repetitive aqueous washes was also studied. The binding sites of the bacteria were not saturated by the maximum ZEN concentration studied.