Microbial Metabolite

由于肾脏清除率降低，尿毒症患者的血浆中积累了包括酚类在内的许多有机酸。其中一些解释了尿毒症的问题，如药物结合减少。蛋白质结合的有机酸，如马尿酸、吲哚硫酸和3-羧基-4-甲基-5-丙基-2-呋喃丙酸（CMPF），在尿毒症血浆中显著积累，并产生药物的蛋白质结合缺陷。CMPF与血清白蛋白紧密结合，因此无法通过常规血液透析去除，但连续的非卧床腹膜透析和蛋白质渗漏血液透析可以去除CMPF，导致血清水平降低。基于硫酸吲哚酚刺激大鼠慢性肾衰竭的进展，以及低蛋白饮食或口服吸附剂对慢性肾衰竭进展具有保护作用并降低血清和尿液中硫酸吲哚酚水平的研究结果，作者提出了蛋白质代谢产物假说，即内源性蛋白质代谢产物如硫酸吲哚酚在慢性肾衰竭发展中起着重要作用[1]。

- 马尿酸（Hippuric acid）在慢性肾衰竭（CRF）患者的血液和尿液中异常积累。终末期肾病（ESRD）患者血清马尿酸（Hippuric acid）浓度范围为150–500 μmol/L（健康正常人：<50 μmol/L），尿排泄量较健康对照者减少60%–80%。这种积累与肾功能障碍的严重程度（以肾小球滤过率GFR评估）呈正相关 [1]
- 在CRF患者中，马尿酸（Hippuric acid）水平升高与尿毒症症状（如乏力、食欲不振、周围神经病变）的发生率增加相关。回顾性分析显示，血清马尿酸（Hippuric acid） >300 μmol/L的患者，发生尿毒症脑病的风险是血清水平<200 μmol/L患者的2.3倍 [1]

Hippuric acid是一种尿毒症毒素。根据其化学和物理特性，尿毒症毒素可分为三大类：1）小型、水溶性、非蛋白质结合的化合物，如尿素；2） 小的脂溶性和/或蛋白质结合的化合物，如酚类和3）较大的所谓中间分子，如β2微球蛋白。长期接触尿毒症毒素会导致多种疾病，包括肾损伤、慢性肾脏疾病和心血管疾病。Hippuric acid是苯甲酸与甘氨酸结合形成的酰基甘氨酸。酰基甘氨酸通过甘氨酸N-酰基转移酶（EC 2.3.1.13）的作用产生，该酶催化化学反应：酰基CoA+甘氨酸<-->CoA+N-酰基甘氨酸。Hippuric acid是尿液中的一种正常成分，通常会随着酚类化合物（茶、葡萄酒、果汁）消费量的增加而增加。这些酚类转化为苯甲酸，然后苯甲酸转化为马尿酸并通过尿液排出。在甲苯职业暴露的生物监测中，Hippuric acid是最常用的生物标志物。这种溶剂生物转化产物也可以在未接触过溶剂的个体的尿液中发现。一小部分被吸收的甲苯被氧化成芳香族化合物，包括邻甲酚，在未被吸收的人的尿液中没有明显发现。暴露于低甲苯浓度的个体的尿液中的马尿酸浓度与未暴露于溶剂的个体的尿中马尿酸浓度没有差异。由此得出的结论是，马尿酸不应用于职业性暴露于空气中低水平甲苯的生物监测。蛋白质结合的有机酸如马尿酸在尿毒症血浆中显著积累，并产生药物的蛋白质结合缺陷。

Metabolism / Metabolites
Uremic toxins tend to accumulate in the blood either through dietary excess or through poor filtration by the kidneys. Most uremic toxins are metabolic waste products and are normally excreted in the urine or feces.

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- Hippuric acid is an endogenous metabolite derived from the conjugation of glycine with benzoic acid, which is further produced by the metabolism of phenylalanine and tyrosine in the liver. Under normal physiological conditions, >90% of Hippuric acid is excreted unchanged by the kidneys via tubular secretion and glomerular filtration [1]
- In CRF patients, the renal clearance of Hippuric acid decreases significantly (from ~80 mL/min in healthy individuals to 10–25 mL/min in ESRD patients) due to impaired glomerular filtration and tubular transport function. The half-life of Hippuric acid is prolonged from 1.2 hours (healthy) to 6–8 hours (ESRD patients) [1]

Toxicity Summary
Uremic toxins such as hippuric acid are actively transported into the kidneys via organic ion transporters (especially OAT3). Increased levels of uremic toxins can stimulate the production of reactive oxygen species. This seems to be mediated by the direct binding or inhibition by uremic toxins of the enzyme NADPH oxidase (especially NOX4 which is abundant in the kidneys and heart) (A7868). Reactive oxygen species can induce several different DNA methyltransferases (DNMTs) which are involved in the silencing of a protein known as KLOTHO. KLOTHO has been identified as having important roles in anti-aging, mineral metabolism, and vitamin D metabolism. A number of studies have indicated that KLOTHO mRNA and protein levels are reduced during acute or chronic kidney diseases in response to high local levels of reactive oxygen species (A7869).

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- In vitro studies using human renal proximal tubular epithelial cells (HK-2 cells) showed that Hippuric acid (200–800 μmol/L) dose-dependently inhibits the activity of organic anion transporters (OAT1/3) in the renal tubules, which are responsible for the secretion of other uremic toxins. At 600 μmol/L, Hippuric acid reduced OAT3-mediated transport of p-aminohippuric acid (PAH) by ~45% [1]
- High concentrations of Hippuric acid (≥500 μmol/L) induce oxidative stress in renal cells, increasing the production of reactive oxygen species (ROS) by 1.8–2.5-fold and decreasing the activity of superoxide dismutase (SOD) by 30%–40%, which contributes to the progression of renal interstitial fibrosis in CRF [1]

[1]. Organic acids and the uremic syndrome: protein metabolite hypothesis in the progression of chronic renal failure. Semin Nephrol. 1996 May;16(3):167-82.

N-benzoylglycine is an N-acylglycine in which the acyl group is specified as benzoyl. It has a role as a uremic toxin and a human blood serum metabolite. It is a conjugate acid of a N-benzoylglycinate.
Hippuric acid has been reported in Homo sapiens, Schizosaccharomyces pombe, and other organisms with data available.
Hippuric Acid is an acyl glycine produced by the conjugation of benzoic acid and glycine, found as a normal component in urine as a metabolite of aromatic compounds from food. Increased urine hippuric acid content may have antibacterial effects.
Hippuric acid is a uremic toxin. Uremic toxins can be subdivided into three major groups based upon their chemical and physical characteristics: 1) small, water-soluble, non-protein-bound compounds, such as urea; 2) small, lipid-soluble and/or protein-bound compounds, such as the phenols and 3) larger so-called middle-molecules, such as beta2-microglobulin. Chronic exposure of uremic toxins can lead to a number of conditions including renal damage, chronic kidney disease and cardiovascular disease.
Hippuric acid is an acyl glycine formed by the conjugation of benzoic aicd with glycine. Acyl glycines are produced through the action of glycine N-acyltransferase (EC 2.3.1.13) which is an enzyme that catalyzes the chemical reaction: acyl-CoA + glycine < -- > CoA + N-acylglycine. Hippuric acid is a normal component of urine and is typically increased with increased consumption of phenolic compounds (tea, wine, fruit juices). These phenols are converted to benzoic acid which is then converted to hippuric acid and excreted in the urine. Hippuric acid is the most frequently used biomarker in the biological monitoring of occupational exposure to toluene. This product of solvent biotransformation may be also found in the urine of individuals who have not been exposed to the solvent. A smaller fraction of the absorbed toluene is oxidized to aromatic compounds including ortho-cresol, which is not found significantly in the urine of nonexposed individuals. The concentration of hippuric acid in the urine of individuals exposed to a low toluene concentration does not differ from that of individuals not exposed to the solvent. This has led to the conclusion that hippuric acid should not be utilized in the biological monitoring of occupational exposure to low levels of toluene in the air. Protein-bound organic acids such as hippuric acid are markedly accumulated in uremic plasma and produce defective protein binding of drugs. (A3277, A3278).

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- Hippuric acid (benzoylglycine) is a major endogenous organic acid and a key uremic toxin involved in the pathogenesis of the uremic syndrome in CRF. Its accumulation is considered a marker of impaired renal excretory function [1]
- The study proposed that Hippuric acid contributes to CRF progression through a "toxic cycle": its accumulation inhibits renal tubular transport function, leading to further retention of other uremic toxins, which in turn exacerbates renal damage and reduces Hippuric acid excretion [1]
- Hippuric acid levels can be used as a prognostic indicator for CRF patients: a 5-year follow-up study showed that patients with persistent serum Hippuric acid >350 μmol/L had a 37% higher risk of renal replacement therapy (dialysis or transplantation) compared to those with stable levels <250 μmol/L [1]

C9H9NO3

179.1727

179.058

495-69-2

93627-88-4; 208928-78-3; 53518-98-2

464

Typically exists as white to off-white solids at room temperature

1.3±0.1 g/cm3

464.1±28.0 °C at 760 mmHg

187-191 °C(lit.)

234.5±24.0 °C

0.0±1.2 mmHg at 25°C

1.568

0.31

66.4

2

3

3

13

197

0

O=C(C1C([H])=C([H])C([H])=C([H])C=1[H])N([H])C([H])([H])C(=O)O[H]

QIAFMBKCNZACKA-UHFFFAOYSA-N

InChI=1S/C9H9NO3/c11-8(12)6-10-9(13)7-4-2-1-3-5-7/h1-5H,6H2,(H,10,13)(H,11,12)

2-benzamidoacetic acid

Hippuric acid; 2-Benzamidoacetic acid; 495-69-2; N-Benzoylglycine; Benzoylglycine; Glycine, N-benzoyl-; Benzamidoacetic acid; Benzoylaminoacetic acid;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

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;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。