Novel Method to Quantify β‑Glucan in Processed Foods: Sodium Hypochlorite Extracting and Enzymatic Digesting (SEED) Assay

Some β-glucans have attracted attention due to their functionality as an immunostimulant and have been used in processed foods. However, accurately measuring the β-glucan content of processed foods using existing methods is difficult. We demonstrate a new method, the Sodium hypochlorite Extracting and Enzymatic Digesting (SEED) assay, in which β-glucan is extracted using sodium hypochlorite, dimethyl sulfoxide, and 5 mol/L sodium hydroxide and then digested into β-glucan fragments using Westase which is an enzyme having β-1,6- and β-1,3 glucanase activity. The β-glucan fragments are further digested into glucose using exo-1,3-β-d-glucanase and β-glucosidase. We measured β-glucan comprising β-1,3-, -1,6-, and -1,(3),4- bonds in various polysaccharide reagents and processed foods using our novel method. The SEED assay was able to quantify β-glucan with good reproducibility, and the recovery rate was >90% for food containing β-glucan. Therefore, the SEED assay is capable of accurately measuring the β-glucan content of processed foods

Toddaculin, Isolated from of <i>Toddalia asiatica</i> (L.) Lam., Inhibited Osteoclastogenesis in RAW 264 Cells and Enhanced Osteoblastogenesis in MC3T3-E1 Cells

<div><p>Osteoporosis with bone loss is widely recognized as a major health problem. Bone homeostasis is maintained by balancing bone formation and bone resorption. The imbalance caused by increased bone resorption over bone formation can lead to various bone-related diseases such as osteoporosis and rheumatoid arthritis. Osteoclasts are the principal cells responsible for bone resorption and the main targets of anti-resorptive therapies. However, excessive inhibition of osteoclast differentiation may lead to inhibition of osteoblast differentiation. Therefore, it is important to screen for new compounds capable of inhibiting bone resorption and enhancing bone formation. <i>Toddalia asiatica</i> (L.) Lam. has been utilized traditionally for medicinal purposes such as the treatment of rheumatism. Currently, the extract is considered to be a good source of pharmacological agents for the treatment of bone-related diseases, but the active compounds have yet to be identified. We investigated whether toddaculin, derived from <i>Toddalia asiatica</i> (L.) Lam., affects both processes by inhibiting bone resorption and enhancing bone formation. Towards this end, we used pre-osteoclastic RAW 264 cells and pre-osteoblastic MC3T3-E1 cells. We found that toddaculin not only inhibited the differentiation of osteoclasts via activation of the NF-κB, ERK 1/2, and p38 MAPK signaling pathways, but it also induced differentiation and mineralization of osteoblasts by regulating differentiation factors. Thus, toddaculin might be beneficial for the prevention and treatment of osteoporosis.</p></div

Effect of toddaculin on ALP activity (A) and osteoblast-related genes (<i>Alpl</i> and <i>Bglap</i>) (B) in ascorbic acid-treated MC3T3-E1 cells.

<p>Values are expressed as means ± SEM (n = 3). *, <i>P</i> < 0.05 compared with control.</p

Effect of toddaculin on RANKL-induced NF-κB activation (A) and MAPKs phosphorylation (B) in RAW 264 cells.

<p>Values are expressed as means ± SEM (n = 3 for NF-κB activation and n = 4 for MAPKs phosphorylation). *, <i>P</i> < 0.05 compared with control.</p

Primer pairs used for RT-PCR.

<p>Primer pairs used for RT-PCR.</p

Effect of toddaculin on osteoblast mineralization in ascorbic acid and β-glycerophosphate-treated MC3T3-E1 cells that were evaluated by Alizarin red S staining (microplate reader analysis (A) and light microscopic evaluation (B)).

<p>Values are expressed as means ± SEM (n = 3). *, <i>P</i> < 0.05 compared with control. Each picture is representative of at least triplicate analyses.</p

Effect of toddaculin on osteoclastogenesis.

<p>Chemical structure of toddaculin (A) and its effect on TRAP activity in RANKL-treated RAW 264 cells (B). Values are expressed as means ± SEM (n = 3). *, <i>P</i> < 0.05 compared with control. Light microscopic analysis of osteoclast formation is shown in (C). Arrows show multi-nucleated cells. Each picture is representative of at least triplicate analyses.</p

A proposed mechanism of the inhibitory effects of toddaculin on osteoclastogenesis.

<p>A proposed mechanism of the inhibitory effects of toddaculin on osteoclastogenesis.</p

Correlation between hemodialysis duration and rSO₂.

<p>Correlation between hemodialysis duration and rSO₂.</p

Patients Characteristics and the correlation between cerebral rSO₂ and clinical parameters.

<p>Patients Characteristics and the correlation between cerebral rSO₂ and clinical parameters.</p