Porous polymer monoliths with complementary retention mechanisms for online solid-phase extraction liquid chromatography to determine lysozyme in egg white

This work demonstrates the determination of lysozyme in egg-white samples after enrichment and cleanup by weak cation exchange (WCX) following separation by reversed-phase liquid chromatography (RPLC). The WCX column was prepared from glycidyl methacrylate (GMA) and ethylene glycol dimethacrylate (EDMA) and functionalized with iminodiacetate (IDA). Reversed-phase columns were prepared using butyl methacrylate (BMA) and EDMA. Photopolymerization formed the poly(GMA-co-EDMA) column inside vinylized polypropylene tubes whereas poly(BMA-co-EDMA) used thermal polymerization inside functionalized Silcosteel® tubes. The preparation of poly(GMA-co-EDMA) was fast (about 2 h), from preparing the polypropylene tube to washing the formed monolith with acetonitrile (ACN), but functionalization demanded an overnight period of pumping IDA through the column immersed in a water bath thermostated at 80 °C. Preparation of the poly(BMA-co-EDMA) also demanded overnight heating at 60 °C, with subsequent washing of the formed monolith with ACN. Egg-white samples diluted at a 1:10 m v−1 ratio in phosphate buffer (pH 7.0) were injected first through IDA@poly(GMA-co-EDMA) to retain lysozyme (pI 11.4) and remove the proteins with a pI < 7.0. Elution of the lysozyme from the cation exchange column was made with 5% (v v−1) acetonitrile in 0.1% (v v−1) TFA. RPLC then analyzed the eluate with a gradient from 5 to 50% ACN in 0.1% TFA. The limits of detection and quantification were 0.07 and 0.23 mg mL−1, respectively. Egg-white lysozyme concentrations varied between 2.26 ± 0.06 and 4.41 ± 0.08 mg g−1, and spiking/recovery experiments at two concentration levels (0.25 and 0.50 mg mL−1) resulted in recoveries from 94 to 115%, thus demonstrating the columns working with orthogonal selectivity provided enrichment of less abundant lysozyme and accurate results, provided by an efficient cleanup of the sample matrix

Chemical constituents and antileukemic activity of <i>Eugenia dysenterica</i>

<p>The study about <i>Eugenia dysenterica</i> led to the isolation of 3<i>-</i>acetyl<i>-</i>urs<i>-</i>12-en-28<i>-</i>oic (<b>1</b>), 3-acetyl-olean-12-en-28-oic acid (<b>2</b>) and isoquercetin (<b>3</b>) from the stem barks, and of 3-<i>O</i>-β-glucopyranosyl-β-sitosterol (<b>4</b>), methyl 3-hydroxy-4-methoxybenzoate (<b>5</b>), methyl 4-hydroxyphenyl propionate (<b>6</b>), <i>E</i>-methyl-4-hydroxycinnamate (<b>7</b>), quercetin-3-<i>O</i>-(6ꞌꞌ-<i>O</i>-galloyl)-β-d-glucopyranoside (<b>8</b>) and quercetin-3-<i>O</i>-β-d-galactopyranoside (<b>9</b>) from the leaves. The structures <b>1</b>–<b>9</b> were set through the analysis of their NMR spectroscopic data. Compounds <b>2</b>, <b>3</b> and <b>5</b>–<b>8</b> were reported for the first time in the <i>Eugenia</i> genus. Compound <b>8</b> reduced cell viability and presented IC₅₀ values 40.3 and 36.7 μM, for the CCRF-CEM and the Kasumi-1 cells, respectively.</p