α-glucosidase

α-葡萄糖苷酶抑制活性[2]
几十年来，研究人员已经证明，桑叶提取物对大鼠和人α-葡萄糖苷酶有很强的抑制作用（Anno et al., 2004, Miyahara et al., 2004, Oku et al., 2006）。α-葡萄糖苷酶位于肠细胞的刷状缘表面膜上，被认为是淀粉和其他碳水化合物消化过程中最重要的酶（Herscovics, 1999）。通过膳食食物和药物改变碳水化合物代谢可能具有治疗价值。1-Deoxynojirimycin (Duvoglustat)/DNJ结合到α-葡萄糖苷酶的活性中心，在小肠中是该酶的有效抑制剂（Junge, Matzke, & Stoltefuss, 1996）。 对于营养保健品的商业开发，应该知道目标化合物及其在产品中的浓度，以达到最佳的治疗效果。在桑树干茶中，我们认为DNJ是关键化合物，因为它能强烈抑制α-葡萄糖苷酶，桑叶中含有高浓度的DNJ（占总亚氨基糖的50%）（Asano et al., 2001）。α-葡萄糖苷酶抑制与纯DNJ (r = 0.96)（图4B）和桑叶DNJ含量（r = 0.84）（图4A）高度相关。在1-Deoxynojirimycin (Duvoglustat)/DNJ浓度下，桑叶提取物对α-葡萄糖苷酶的抑制活性高于DNJ标准：如在5 μg DNJ/ml时，桑叶提取物对α-葡萄糖苷酶的抑制作用为27%，纯DNJ对α-葡萄糖苷酶的抑制作用为23%。这种额外的抑制作用可以解释为桑树提取物中存在其他亚氨基糖（即n -甲基- dnj， 2-O-α-d-半乳糖酰氨基- dnj和fagomine）和其他成分，如异槲皮素，槲皮素和芦丁。

Duvoglustat 盐酸盐（1-Deoxynojirimycin 盐酸盐）（20-80 mg/kg；静脉注射；每天一次，持续四个星期）显示出抗肥胖作用 [3]。盐酸度伏鲁司他通过激活 db/db 小鼠骨骼肌中的胰岛素信号 PI3K/AKT 通路，显着提高胰岛素敏感性[3]。

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1-Deoxynojirimycin (Duvoglustat)/DNJ作为肠道α-葡萄糖苷酶抑制剂广泛用于糖尿病的治疗。然而，关于其对改善胰岛素敏感性的作用的报道很少。本研究的目的是探讨DNJ是否通过改善胰岛素敏感性来降低高血糖。建立了一种制备大量DNJ的经济方法。然后，db/db小鼠静脉注射DNJ(20、40和80 mg·kg（-1）·d(-1)) 4周。通过血糖和生化分析来评价其对高血糖的治疗效果，并探讨骨骼肌的相关分子机制。DNJ显著降低体重、血糖和血清胰岛素水平。DNJ治疗也改善了葡萄糖耐量和胰岛素耐量。此外，虽然骨骼肌中总蛋白激酶B （AKT）、磷脂酰肌醇3激酶（PI3K）、胰岛素受体β (IR-β)、胰岛素受体底物-1 （IRS1）和葡萄糖转运蛋白4 （GLUT4）的表达不受影响，但DNJ处理显著增加了Ser473-AKT、p85-PI3K、Tyr1361-IR-β和Tyr612-IRS1的GLUT4易位和磷酸化。这些结果表明，DNJ通过激活db/db小鼠骨骼肌中胰岛素信号通路PI3K/AKT显著改善胰岛素敏感性。[3]

α-葡萄糖苷酶抑制试验[1]
α-葡萄糖苷酶抑制活性是通过Ma， Hattori， Daneshtalab和Wang（2008）描述的程序的修改来测量的。简单地说，将大鼠肠丙酮粉（1 g）悬浮在100 mM磷酸钾缓冲液（pH 7.0）中，超声振荡20 min。在3000 rpm下离心30 min后，将上清液作为α-葡萄糖苷酶的来源。底物（2 mM 4-硝基苯-α-d-葡萄糖吡喃苷）在100 mM磷酸钾缓冲液（pH 7.0）中移液至96孔板（40 μl/孔）。加入5 μl桑树样品或对照溶液（乙醇与蒸馏水的比例为50:50）混合。加入酶（5 μl）后，37℃孵育20 min，测定405 nm的紫外吸光度。计算桑树样品和标准1-Deoxynojirimycin (Duvoglustat)/DNJ α-葡萄糖苷酶抑制活性的百分比为：（ΔAcontrol-ΔAsample） × 100/ΔAcontrol，其中ΔA为405 nm吸光度。

蛋白质印迹[3]
为了研究1-脱氧野尻霉素/1-Deoxynojirimycin (Duvoglustat)对胰岛素信号通路的影响，如前所述进行了蛋白质印迹分析。简而言之，骨骼肌组织（0.1 g）在裂解缓冲液（50 mM Tris（pH 7.4）、150 mM NaCl、0.1%SDS、0.5%脱氧胆酸钠、1%NP40、10μL磷酸酶抑制剂、1μL蛋白酶抑制剂和5μL 100 mM PMSF）中裂解，在4°C下以16000×g离心15分钟，并通过双辛可宁酸蛋白测定法定量蛋白质浓度。将等量的蛋白质（70μg）装载在10%SDS-PAGE上，并转移到PVDF膜上。膜被阻断后，它们与抗IR-β、对-Tyr1361-IR-β，IRS1、对-Tur612-IRS1、PI3K、对-p85-PI3K、AKT、对-Ser473-AKT、GLUT4、β-actin或Na+K+-ATP酶α1的一抗在4°C下孵育过夜，然后在室温下与HRP偶联的二抗孵育2小时。使用ECL检测试剂盒观察蛋白质条带。以β-actin为对照进行总蛋白表达的正常化。以Na+K+-ATP酶α1为对照进行m-GLUT4表达的正常化。

db/db mice
20, 40, 80 mg/kg
Intravenously; daily for four weeks

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At the end of ten weeks, wild-type C57BLKS mice, which received intravenously normal saline, served as a normal control (N control) (n = 6). The db/db mice were divided into four groups (n = 6): Group I served as a diabetic control and received intravenously normal saline (D control). Group II, III, and IV were treated intravenously with 1-Deoxynojirimycin (Duvoglustat)/DNJ 20, 40, and 80 mg·kg−1·day−1, respectively. An intravenous injection was selected to avoid the function of DNJ as an α-Glycosidase inhibitor inthe gastrointestinal tract. For DNJ doses selection, in our previous study, we screened a large number of Chinese traditional medicines including mulberry leaves by glucose tolerance test of ICR mice. We found the alkaloids (DNJ 40 mg·kg−1) isolated from mulberry leaves could improve the glucose tolerance test of ICR mice (Figure A1). We then tested doses of 10, 20, and 40 mg·kg−1, but both 10 and 20 mg·kg−1 did not have any effect (Figure A2). Therefore, we selected the 1-Deoxynojirimycin (Duvoglustat)/DNJ doses as 20, 40, and 80 mg·kg−1·day−1. All these doses were given for 4 weeks. The blood glucose, body weight and average food intake, water intake, and urine output were measured every week. At the end of the experimental period, the mice were anesthetized with chloral hydrate after withholding food for 12 h, and blood samples were taken to determine the serum insulin levels. Besides, skeletal muscle were removed after the blood was collected, then rinsed with a physiological saline solution, and immediately stored at −80 °C [3].

rat LD50 oral >5 gm/kg BEHAVIORAL: SOMNOLENCE (GENERAL DEPRESSED ACTIVITY); KIDNEY, URETER, AND BLADDER: URINE VOLUME INCREASED; SKIN AND APPENDAGES (SKIN): HAIR: OTHER International Journal of Toxicology., 16(Suppl

[1]. 1-Deoxynojirimycin: Occurrence, Extraction, Chemistry, Oral Pharmacokinetics, Biological Activities and In Silico Target Fishing. Molecules. 2016 Nov 23;21(11). pii: E1600.

[2]. Development of high 1-deoxynojirimycin (DNJ) content mulberry tea and use of response surface methodology to optimize tea-making conditions for highest DNJ extraction. LWT - Food Science and Technology. Volume 45, Issue 2, March 2012, Pages 226-232.

[3]. 1-Deoxynojirimycin Alleviates Insulin Resistance via Activation of Insulin Signaling PI3K/AKT Pathway in Skeletal Muscle of db/db Mice. Molecules. 2015 Dec 4;20(12):21700-14.

AT2220 is an experimental, oral therapy for the treatment of Pompe disease and belongs to a class of molecules known as pharmacological chaperones. It is a small molecule designed to act as a pharmacological chaperone that specifically binds, stabilizes, and facilitates the proper folding and trafficking of α-glucosidase (GAA). GAA to the lysosome, where it can perform its normal function. AT2220 has been shown to increase GAA activity in cell lines derived from Pompe patients and in transfected cells expressing misfolded forms of GAA.
An alpha-glucosidase inhibitor with antiviral action. Derivatives of deoxynojirimycin may have anti-HIV activity.

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Drug Indication
Pompe disease, also known as glycogen storage disease type II or acid maltase deficiency, is a relatively rare neuromuscular and lysosomal storage disorder caused by inherited genetic mutations in a key enzyme called α-glucosidase (Gaa).

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Mechanism of Action
AT2220 is designed to act as a pharmacological chaperone by selectively binding to the misfolded enzyme responsible for Pompe disease, Gaa. After binding to the enzyme, it is thought that AT2220 promotes the proper folding, processing, and trafficking of the enzyme from the endoplasmic reticulum to its final destination, the lysosome, the area of the cell where the enzyme does its work. Once it reaches the lysosome, the pharmacological chaperone is displaced, and the enzyme can perform its normal function, which is the breakdown of its natural substrate, glycogen.

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Duvoglustat is an optically active form of 2-(hydroxymethyl)piperidine-3,4,5-triol having 2R,3R,4R,5S-configuration. It has a role as an EC 3.2.1.20 (alpha-glucosidase) inhibitor, an anti-HIV agent, an anti-obesity agent, a bacterial metabolite, a hypoglycemic agent, a hepatoprotective agent and a plant metabolite. It is a 2-(hydroxymethyl)piperidine-3,4,5-triol and a piperidine alkaloid.
An alpha-glucosidase inhibitor with antiviral action. Derivatives of deoxynojirimycin may have anti-HIV activity.
1-Deoxynojirimycin has been reported in Parmotrema austrosinense, Parmotrema praesorediosum, and other organisms with data available.
An alpha-glucosidase inhibitor with antiviral action. Derivatives of deoxynojirimycin may have anti-HIV activity.
See also: Fagomine (annotation moved to).

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1-Deoxynojirimycin (DNJ, C₆H13NO₄, 163.17 g/mol), an alkaloid azasugar or iminosugar, is a biologically active natural compound that exists in mulberry leaves and Commelina communis (dayflower) as well as from several bacterial strains such as Bacillus and Streptomyces species. Deoxynojirimycin possesses antihyperglycemic, anti-obesity, and antiviral features. Therefore, the aim of this detailed review article is to summarize the existing knowledge on occurrence, extraction, purification, determination, chemistry, and bioactivities of DNJ, so that researchers may use it to explore future perspectives of research on DNJ. Moreover, possible molecular targets of DNJ will also be investigated using suitable in silico approach.[1]

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Mulberry 1-deoxynojirimycin (DNJ), a potent α-glucosidase inhibitor, suppresses postprandial blood glucose, thereby possibly preventing diabetes mellitus. At present, mulberry dry teas are commercially supplied as functional foods in many countries, but these products may not provide an effective dose (6 mg DNJ/60 kg human wt) due to their low DNJ content (about 100 mg/100 g of dry wt). Therefore, development of tea with higher DNJ content is desirable. To do this, we investigated distribution of DNJ content and α-glucosidase inhibitory activity in 35 Thai mulberry varieties. DNJ content in young leaves varied among mulberry varieties from 30 to 170 mg/100 g of dry leaves. Varieties having highest DNJ content were Kam, Burirum 60 and Burirum 51. Leaf position affected DNJ content: shoots > young leaves > mature leaves. DNJ concentration and α-glucosidase inhibitory activity were highly correlated (r = 0.84), suggesting that α-glucosidase inhibitory activity of mulberry leaves is mainly due to DNJ. Consequently, high DNJ content mulberry tea was produced from shoots of varieties such as Burirum 60, which contains 300 mg/100 g of dry wt. Tea-making conditions were optimized for highest DNJ extraction using response surface methodology. Approximate 95% of total DNJ in high DNJ content dry tea was extracted when temperature was maintained at 98 °C for 400 s; these conditions could be applicable for preparation of commercial products with high DNJ content. One cup (230 ml, a normal serving) of DNJ-enriched mulberry tea contained enough DNJ (6.5 mg) to effectively suppress postprandial blood glucose.[2]

C6H14CLNO4

199.631

199.061

C, 36.10; H, 7.07; Cl, 17.76; N, 7.02; O, 32.06

73285-50-4

1-Deoxynojirimycin;19130-96-2

13018787

White to off-white solid powder

1.456 g/cm3

361.1ºC at 760 mmHg

195-196ºC

197.3ºC

92.95

6

5

1

12

132

4

Cl.OC[C@H]1NC[C@H](O)[C@@H](O)[C@@H]1O

ZJIHMALTJRDNQI-VFQQELCFSA-N

InChI=1S/C6H13NO4.ClH/c8-2-3-5(10)6(11)4(9)1-7-3/h3-11H,1-2H21H/t3-,4+,5-,6-/m1./s1

(2R,3R,4R,5S)-2-(hydroxymethyl)piperidine-3,4,5-triol hydrochloride

Duvoglustat; BAY-h 5595; AT2220; AT-2220; AT 2220; DNJ; 1DNJ; NOJ; Moranoline; Moranolin; Duvoglustat hydrochloride; deoxynojirimycin; 1-Deoxynojirimycin; 1-Deoxy-Nojirimycin 1-Deoxynojirimycin hydrochloride; 73285-50-4; Duvoglustat HCl; Moranoline hydrochloride; Moranoline HCl; (2R,3R,4R,5S)-2-(Hydroxymethyl)piperidine-3,4,5-triol hydrochloride

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 请将本产品存放在密封且受保护的环境中（例如氮气保护），避免吸湿/受潮和光照。

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

H2O : ~250 mg/mL (~1252.32 mM)

注意: 如下所列的是一些常用的体内动物实验溶解配方，主要用于溶解难溶或不溶于水的产品（水溶度<1 mg/mL）。 建议您先取少量样品进行尝试，如该配方可行，再根据实验需求增加样品量。

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注射用配方1: DMSO : Tween 80： Saline = 10 : 5 : 85 (如： 100 μL DMSO → 50 μL Tween 80 → 850 μL Saline)
*生理盐水/Saline的制备：将0.9g氯化钠/NaCl溶解在100 mL ddH ₂ O中，得到澄清溶液。
注射用配方 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (如： 100 μL DMSO → 400 μL PEG300 → 50 μL Tween 80 → 450 μL Saline)
注射用配方 3: DMSO : Corn oil = 10 : 90 (如： 100 μL DMSO → 900 μL Corn oil)
示例: 以注射用配方 3 (DMSO : Corn oil = 10 : 90) 为例说明, 如果要配制 1 mL 2.5 mg/mL的工作液, 您可以取 100 μL 25 mg/mL 澄清的 DMSO 储备液，加到 900 μL Corn oil/玉米油中, 混合均匀。

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注射用配方 4: DMSO : 20% SBE-β-CD in Saline = 10 : 90 [如：100 μL DMSO → 900 μL (20% SBE-β-CD in Saline)]
*20% SBE-β-CD in Saline的制备（4°C，储存1周）：将2g SBE-β-CD (磺丁基-β-环糊精) 溶解于10mL生理盐水中，得到澄清溶液。
注射用配方 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (如： 500 μL 2-Hydroxypropyl-β-cyclodextrin (羟丙基环胡精)→ 500 μL Saline)
注射用配方 6: DMSO : PEG300 : Castor oil : Saline = 5 : 10 : 20 : 65 (如： 50 μL DMSO → 100 μL PEG300 → 200 μL Castor oil → 650 μL Saline)
注射用配方 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (如： 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
注射用配方 8: 溶解于Cremophor/Ethanol (50 : 50), 然后用生理盐水稀释。
注射用配方 9: EtOH : Corn oil = 10 : 90 (如： 100 μL EtOH → 900 μL Corn oil)
注射用配方 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (如： 100 μL EtOH → 400 μL PEG300 → 50 μL Tween 80 → 450 μL Saline)

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口服配方 1: 悬浮于0.5% CMC Na (羧甲基纤维素钠)
口服配方 2: 悬浮于0.5% Carboxymethyl cellulose (羧甲基纤维素)
示例: 以口服配方 1 (悬浮于 0.5% CMC Na)为例说明, 如果要配制 100 mL 2.5 mg/mL 的工作液, 您可以先取0.5g CMC Na并将其溶解于100mL ddH2O中，得到0.5%CMC-Na澄清溶液；然后将250 mg待测化合物加到100 mL前述 0.5%CMC Na溶液中，得到悬浮液。

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口服配方 3: 溶解于 PEG400 (聚乙二醇400)
口服配方 4: 悬浮于0.2% Carboxymethyl cellulose (羧甲基纤维素)
口服配方 5: 溶解于0.25% Tween 80 and 0.5% Carboxymethyl cellulose (羧甲基纤维素)
口服配方 6: 做成粉末与食物混合

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注意: 以上为较为常见方法，仅供参考， InvivoChem并未独立验证这些配方的准确性。具体溶剂的选择首先应参照文献已报道溶解方法、配方或剂型，对于某些尚未有文献报道溶解方法的化合物，需通过前期实验来确定（建议先取少量样品进行尝试），包括产品的溶解情况、梯度设置、动物的耐受性等。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
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