Quercetin Inhibits Advanced Glycation End Product Formation by Trapping Methylglyoxal and Glyoxal

Methylglyoxal (MGO) and glyoxal (GO) not only are endogenous metabolites but also exist in exogenous resources, such as foods, beverages, urban atmosphere, and cigarette smoke. They have been identified as reactive dicarbonyl precursors of advanced glycation end products (AGEs), which have been associated with diabetes-related long-term complications. In this study, quercetin, a natural flavonol found in fruits, vegetables, leaves, and grains, could effectively inhibit the formation of AGEs in a dose-dependent manner via trapping reactive dicarbonyl compounds. More than 50.5% of GO and 80.1% of MGO were trapped at the same time by quercetin within 1 h under physiological conditions. Quercetin and MGO (or GO) were combined at different ratios, and the products generated from this reaction were analyzed with LC-MS. Both mono-MGO and di-MGO adducts of quercetin were detected in this assay using LC-MS, but only tiny amounts of mono-GO adducts of quercetin were found. Additionally, di-MGO adducts were observed as the dominant product with prolonged incubation time. In the bovine serum albumin (BSA)–MGO/GO system, quercetin traps MGO and GO directly and then significantly inhibits the formation of AGEs

Influence of Quercetin and Its Methylglyoxal Adducts on the Formation of α‑Dicarbonyl Compounds in a Lysine/Glucose Model System

Increasing evidence has identified α-dicarbonyl compounds, the reactive intermediates generated during Maillard reaction, as the potential factors to cause protein glycation and the development of chronic diseases. Therefore, there is an urgent need to decrease the levels of reactive dicarbonyl compounds in foods. In this study, we investigated the inhibitory effect of quercetin, a major dietary flavonoid, and its major mono- and di-MGO adducts on the formation of dicarbonyl compounds, such as methylglyoxal (MGO) and glyoxal (GO), in a lysine/glucose aqueous system, a model system to reflect the Maillard reaction in food process. Our result indicated that quercetin could efficiently inhibit the formation of MGO and GO in a time-dependent manner. Further mechanistic study was conducted by monitoring the formation of quercetin oxidation and conjugation products using LC-MS/MS. Quercetin MGO adducts, quercetin quinones, and the quinones of quercetin MGO adducts were detected in the system, indicating quercetin plays a dual role in inhibiting the formation of MGO and GO by scavenging free radicals generated in the system and trapping of MGO and GO to form MGO adducts. In addition, we prepared the mono- and di-MGO quercetin adducts and investigated their antioxidant activity and trapping capacity of MGO and GO. Our results indicated that both mono- and di-MGO quercetin adducts could scavenge the DPPH radical in a dose-dependent manner with >40% DPPH scavenged by the MGO adducts at 10 μM, and the di-MGO quercetin adduct could further trap MGO to generate tri-MGO adducts. Therefore, we demonstrate for the first time that quercetin MGO adducts retain the antioxidant activity and trapping capacity of reactive dicarbonyl species

Elimination of Acrolein by Disodium 5′-Guanylate or Disodium 5′-Inosinate at High Temperature and Its Application in Roasted Pork Patty

Acrolein (ACR) is a highly active, simple unsaturated aldehyde found in various high-temperature processed foods. Its long-term accumulation in the human body increases the risk of chronic diseases. Animal and plant foodstuffs are rich in disodium 5′-guanylate (GMP) and disodium 5′-inosinate (IMP), which are authorized flavor enhancers. Herein, we used liquid chromatography with tandem mass spectrometry to explore the reaction-active kinetics and pathway of the interaction between GMP/IMP and ACR and validated it in roasted pork patties. Our results suggested that GMP and IMP could efficiently eliminate ACR by forming ACR adducts (GMP–ACR, IMP–ACR). In addition, IMP exhibited a higher reaction rate, whereas GMP had a good trapping capacity at a later stage. As carriers of GMP and IMP, dried mushrooms and shrimp exhibited an effective ACR-trapping ability in the ACR model and roasted pork patty individually and in combination. Adding 10% of dried mushroom or shrimp alone or 5% of dried mushroom and shrimp in combination eliminated up to 53.9%, 55.8%, and 55.2% ACR in a roasted pork patty, respectively. This study proposed a novel strategy to eliminate the generation of ACR in roasted pork patties by adding foodstuffs rich in GMP and IMP

Over-expression of <i>bmp2b</i> specifically localized in the CHT area at 79 hpf upon ginger or 10-G exposure in normal and in anemic zebrafish embryos.

<p>Whole-mount in situ hybridization of <i>bmp2b.</i> (A–C, left) Normal non-anemic control embryos or embryos treated with ginger/10-G. (D–F, right) Anemic control embryos or anemic embryos treated with ginger/10-G. Anemic groups were treated with 0.5 µM PHZ from 33 to 48 hpf. Embryos express <i>bmp2b</i> in the CHT region (arrows) following exposure to ginger (B, E) or 10-G (C, F). (G) A table shows the percentage of embryos with <i>bmp2b</i> expression in the CHT area at 79 hpf. Scale bars = 420 µm.</p

Ginger/10-G treatment after gastrulation promotes <i>bmp2b/7a</i> in the developing caudal hematopoietic tissue.

<p>(A–B) Zebrafish embryos were treated with ginger (5 µg/ml) or 10-G (2 µg/ml) from 10 to 48 hpf, followed by whole-mount in situ hybridization of <i>bmp2b</i> (A) and <i>bmp7a</i> (B). Both <i>bmp2b</i> and <i>bmp7a</i> were up-regulated locally in the CHT (and underlying fin) upon ginger or 10-G exposure (whereas they are not expressed in the CHT of control embryos at 48 hpf). Scale bars = 700 µm.</p

Ginger extract and its purified phenolic compounds promote <i>Tg(gata1:dsRed)</i> fluorescence and <i>gata1</i> mRNA expression.

<p>(A) Bright field (top left) and <i>Tg(gata1:dsRed)</i> fluorescence of zebrafish embryos at about 22 hpf, before the onset of circulation (anterior to the left). Exposure to ginger extract or its compounds 8-gingerol (8-G), 10-gingerol (10-G), 8-shogaol (8-S) and 10-shogaol (10-S) promoted <i>Tg(gata1:dsRed)</i> fluorescent erythroid cell development in the ICM and PBI (arrows), as compared to control embryos. N = 35 embryos per group. In this panel, we show an embryo treated with a lower concentration of 6-S (2 µg/ml) as this compound was toxic at higher doses. Scale bar = 400 µm. (B) Whole-mount in situ hybridization of ginger or 10-G treated embryos (8 hpf to 21 hpf exposure) revealed increased expression of <i>gata1</i> transcript at 22 hpf. N = 50 embryos per group. Scale bar = 350 µm. (C) At 48 hpf, control embryos at the top; ginger or 10-G treated embryos at the bottom. Scale bar = 500 µm. Fluorescent erythrocytes circulating in the axial vasculature (arrows) and in the pericardial space (arrow heads).</p

Over-expression of <i>bmp7a</i> specifically localized in the CHT region at 79 hpf upon ginger or 10-G exposure in normal and in anemic zebrafish embryos.

<p>Whole-mount in situ hybridization of <i>bmp7a</i>. (A–C, left) Normal non-anemic control embryos or embryos treated with ginger/10-G. (D–F, right) Anemic control embryos or anemic embryos treated with ginger/10-G. Anemic group were treated with 0.5 µM PHZ from 33 to 48 hpf. Embryos express <i>bmp7a</i> in the CHT area (arrows) following exposure to ginger (B, E) or 10-G (C, F). (G) A table shows the percentage of embryos with <i>bmp7a</i> expression in the CHT region at 79 hpf. Scale bars = 420 µm.</p

Ginger/10-G treatment during gastrulation promotes <i>bmp2b/7a</i> and Bmp target gene expression in zebrafish embryos.

<p>(A) Treatment of late gastrulae with ginger at 15 or 20 µg/ml induces the <i>mercedes</i> mutant-like phenotype (partial duplication of the tail fin) at 1 dpf in 8% or 10% of the treated embryos, respectively. Thus, the zebrafish embryos exposed to ginger extract mimic the phenotype of the <i>ogon</i> mutant, which has a mutation in <i>sizzled</i>, a <i>bmp</i> suppressor gene, at 1 dpf. (B) <i>bmp7a</i> expression was strongly increased and extended to the entire blastoderm at 60% epiboly, following short-term exposure to ginger (5 µg/ml) or 10-G (1 µg/ml) from sphere (4 hpf) to 60% epiboly (7 hpf) stages. (C) Up-regulation and extension of the expression domain were observed for <i>bmp2b</i> at 60% epiboly. (D–E) Accordingly, BMP target genes were up-regulated after ginger/10G treatment from the sphere stage (4 hpf) to 7 hpf, as illustrated by enhanced <i>eve1</i> extended towards the dorsal side (arrow heads), a ventral mesoderm marker (D), and <i>gata2,</i> a non-neural ectoderm marker (E), in zebrafish embryos at 60% epiboly. Pictures on left panels show gastrulae, dorsal side to the right (B–E) and statistics tables (right panels) are representative of three independent experiments. N = number of embryos per group. Scale bars = 250 µm.</p

Ginger/10-G treatment increases hematopoietic progenitor markers expression.

<p>Zebrafish embryos were treated with ginger or 10-G from 9 to 21 hpf. (A) <i>Tg(gata1:dsRed)</i> for erythrocyte and <i>Tg(flk1:GFP)</i> for blood vessels, double-fluorescent overlay pictures of embryos at 22 hpf after exposure to ginger or 10-G. Hypertrophy of the PBI vascular plexus in <i>Tg(flk1:GFP)</i> after ginger or 10-G treatment, with <i>Tg(gata1:dsRed)</i> red fluorescent erythrocytes accumulated inside the honeycomb-like vasculature (arrows). Scale bars = 500 µm. Whole-mount in situ hybridization of <i>c-myb</i> (B) and <i>scl</i> (C) in zebrafish embryos at 22 hpf. Both hematopoietic progenitor markers were up-regulated in primitive hematopoietic tissues (ICM+PBI) upon ginger (arrow head) or 10-G (arrow) exposure. Scale bar = 350 µm.</p

Ginger or 10-G exposure promotes erythrocyte recovery from anemia via a Bmp/Smad signal-dependent mechanism.

<p>Bmp/Smad inhibition abolishes the hematopoiesis promoting effect of ginger and 10-G. (A–B) The effect of ginger on hematopoiesis was quantitated in zebrafish embryos after phenylhydrazine (PHZ) induced acute hemolytic anemia, followed by extensive washes and treatments with ginger extract (A) or 10-G (B) with or without dorsomorphin (DMP; 0.1 µM). Ginger and 10-G promote hematopoietic recovery in PHZ treated embryos. Videos of circulating erythrocytes were analyzed and erythrocyte numbers for “PHZ+ginger” and “PHZ+ginger+DMP” assays were calculated and normalized with blood flow (velocity) using the PHZ control value as a reference. Tables summarize the results of one representative experiment. Experiments were repeated 3 times. n = number of embryos analyzed per group. <i>p</i> values were determined by using the Student’s t-test. (C) Regions of erythropoiesis promoted by ginger and 10-G are indicated on cartoons of zebrafish embryos at 22 hpf (primitive wave; before circulation), and at 5–6 dpf during the definitive wave of hematopoiesis. DMP-mediated inhibition of Bmp/Smad signal refers to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone.0039327.s005" target="_blank">Figures S5</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone.0039327.s006" target="_blank">S6</a>, 6 and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone.0039327.s007" target="_blank">S7</a> data. DMH1-mediated inhibition of Bmp/Smad signaling refers to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone.0039327.s005" target="_blank">Figures S5</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone.0039327.s006" target="_blank">S6</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone.0039327.s008" target="_blank">S8</a> data. During the primitive wave of hematopoiesis, expression of <i>gata1</i> and <i>Tg(gata1:dsRed)</i> were increased in the ICM and PBI, as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone-0039327-g001" target="_blank">Figure 1</a>, and the hematopoietic progenitor markers <i>cmyb</i> and <i>scl</i> were up-regulated in the same hematopoietic tissues, as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone-0039327-g002" target="_blank">Figure 2</a>. During the definitive wave, <i>Tg(gata1:dsRed)</i> circulating cells were promoted at 5/6 dpf upon ginger/10-G exposure (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone-0039327-g005" target="_blank">Figures 5</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone-0039327-g006" target="_blank">6</a>), and the hematopoietic progenitor markers <i>cmyb</i>, <i>scl</i> and <i>lmo2</i> were up-regulated in the CHT/hemogenic endothelium at 6 dpf (<i>cmyb, </i><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone-0039327-g007" target="_blank">Figure 7</a>) or in the CHT only at 5 dpf (<i>scl</i> and <i>lmo2, </i><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039327#pone.0039327.s009" target="_blank">Figure S9</a>).</p