A randomized, double-blind study of larazotide acetate to prevent the activation of celiac disease during gluten challenge.

OBJECTIVES: In patients with celiac disease, enteropathy is caused by the entry of gluten peptides into the lamina propria of the intestine, in which their immunogenicity is potentiated by tissue transglutaminase (tTG) and T-helper type 1-mediated immune responses are triggered. Tight junction disassembly and paracellular permeability are believed to have an important role in the transport of gluten peptides to the lamina propria. Larazotide acetate is a tight-junction regulator peptide that, in vitro, prevents the opening of intestinal epithelial tight junctions. The aim of this study was to evaluate the efficacy and tolerability of larazotide acetate in protecting against gluten-induced intestinal permeability and gastrointestinal symptom severity in patients with celiac disease. METHODS: In this dose-ranging, placebo-controlled study, 86 patients with celiac disease controlled through diet were randomly assigned to larazotide acetate (0.25, 1, 4, or 8 mg) or placebo three times per day with or without gluten challenge (2.4 g/day) for 14 days. The primary efficacy outcome was the urinary lactulose/mannitol (LAMA) fractional excretion ratio. Secondary endpoints included gastrointestinal symptom severity, quality-of-life measures, and antibodies to tTG. RESULTS: LAMA measurements were highly variable in the outpatient setting. The increase in LAMA ratio associated with the gluten challenge was not statistically significantly greater than the increase in the gluten-free control. Among patients receiving the gluten challenge, the difference in the LAMA ratios for the larazotide acetate and placebo groups was not statistically significant. However, larazotide acetate appeared to limit gluten-induced worsening of gastrointestinal symptom severity as measured by the Gastrointestinal Symptom Rating Scale at some lower doses but not at the higher dose. Symptoms worsened significantly in the gluten challenge-placebo arm compared with the placebo-placebo arm, suggesting that 2.4 g of gluten per day is sufficient to induce reproducible gluten toxicity. Larazotide acetate was generally well tolerated. No serious adverse events were observed. The most common adverse events were headache and urinary tract infection. CONCLUSIONS: LAMA variability in the outpatient setting precluded accurate assessment of the effect of larazotide acetate on intestinal permeability. However, some lower doses of larazotide acetate appeared to prevent the increase in gastrointestinal symptom severity induced by gluten challenge.

[Determination of glucosinolates in rapeseed by reversed-phase ion-pair liquid chromatography].

This paper presents a method for the direct determination of intact glucosinolates in rapeseed by reversed-phase ion-pair liquid chromatography (RILC). The catechin, mercaptoethanol and phytic acid was adopted respectively in sample pretreatment to prevent indole glucosinolates being oxidized. Among them, the effect of mercaptoethanol was the most obvious. The effects of composition and concentration of the mobile phase, the pH of the mobile phase and the column temperature on the retention and the capacity factor of the glucosinolates were studied. The condition of this method by RILC has been set up: column, YWG-C18 H37(10 microns, 250 mm x 4 mm i.d.); mobile phase, 0.02 mol/L KH2PO4 buffer(pH 6) containing 4.5 mmol/L (C4H8)4NBr and CH3CN(90/10, V/V); flow rate, 1 mL/min; detector, UV-226 nm; column temperation, 30 degrees C. In this condition, six glucosinolates were separated completely. The relative correction factors were determined by using sinigrin or benzoic acid as the internal standard. The characteristics of glucosinolates in different kinds of Chinese rapeseed can be determined by this RILC method.

Matrix-assisted laser desorption/ionization mass spectrometry of deoxynucleotides labeled with an IMI dye.

The four major deoxynucleotides of DNA, and adduct mixtures resulting from separate reactions of 5'-dAMP and 5'-dGMP with benzo[a]pyrene diolepoxide (BPDE), were analyzed by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) after labeling of their phosphate group with an IMI dye. The latter reagent comprises an imidazole functional group attached to a BODIPY (4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene+ ++) fluorophore. Good sensitivity was observed in the detection of the IMI-labeled products by MALDI-MS: 300-500 fmol in the laser spot (1% of the 30-50 pmol sample on the target) gave a signal-to-noise (S/N) of >/=30 from 20-30 superimposed laser shots. The BPDE reaction products, after the IMI labeling, were also subjected to capillary electrophoresis with laser-induced fluorescence detection, which revealed a complex mixture of products. Overall the results encourage the further development of this 'IMI-postlabeling' methodology as an alternative to (32)P-postlabeling for the detection of DNA adducts.

Preparation of an IMI dye (imidazole functional group) containing a 4-(N,N-dimethylaminosulfonyl)-2,1,3-benzoxadiazole fluorophore for labeling of phosphomonoesters.

We are studying dye-imidazole conjugates ("IMI dyes") as reagents for labeling phosphomonoesters such as nucleotides. Previously we have employed a BODIPY dye in our IMI reagents, and analyzed the labeled products by capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) involving an argon ion laser. (The BODIPY fluorophore is a 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene). Here we broaden the technology by preparing a DBD-IMI dye [DBD = 4-(N,N-dimethylaminosulfonyl)-2,1,3-benzoxadiazole], and using a helium-cadmium laser. While DBD-IMI (IMI3) is about 50x more stable photolytically than a BODIPY-IMI dye (IMI2, a conjugate of a BODIPY dye with histamine, was tested), the detection limit for IMI2 (5.10(-11) M; S/N = 5, CE-LIF with an argon ion laser) is tenfold better than that for IMI3 (5.10(-10) M, S/N = 5, helium-cadmium laser). IMI3 conjugates of the four major DNA nucleotides were prepared and detected by CE-LIF.