Endogenous Metabolite; Microbial Metabolite

高羊茅(Festuca arundinacea Schreb)是一种典型的冷季草，广泛用于草坪和牧场。然而，高温作为一种非生物胁迫严重影响了其利用。本研究旨在探讨亚精胺(Spd)对高羊茅热应激反应的影响。在22℃(常温)和44℃(热胁迫)条件下处理4 h。结果表明，外源Spd在常温条件下部分改善了高羊茅叶片的质量。然而，经过热胁迫处理后，外源Spd显著降低了高羊茅叶片的电解质泄漏。Spd还能显著降低H2O2和O2⋅-含量，提高抗氧化酶活性。此外，PAs还能调节SOD、POD、APX等抗氧化酶的活性，有助于清除ROS。此外，施用Spd还能显著提高叶绿素含量，并对高温下叶绿素α荧光瞬态有积极影响。通过jip测试发现，Spd试剂增强了光系统II (PSII)的性能。在热应力作用下，Spd显著提高了能量分岔阶段的部分势(PIABS和PItotal)、量子产率和效率(φP0、δR0、φR0和γRC)。外源Spd还能降低QA-还原PSII反应中心(RC)的比能通量(TP0/RC和ET0/RC)。此外，外源Spd提高了编码PSII核心反应中心复合物蛋白的psbA和psbB的表达水平。我们推断PAs可以稳定核酸的结构，保护RNA免受核糖核酸酶的降解。总之，我们的研究表明，外源Spd通过维持细胞膜稳定性、增加抗氧化酶活性、改善PSII及相关基因表达等途径增强高羊茅的耐热性[1]。

粗酶萃取[1]
酶提物取0.2 g叶粉，用液氮浸泡在4℃预冷的4ml磷酸盐缓冲液(150 mM, pH 7.0)中，用0.2 M Na2HPO4和0.2 M NaH2PO4均质。然后，匀浆在4℃下15000 × g离心30 min，最后收集上清，4℃保存，测定酶活性[1]。

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抗氧化酶活性[1]
在2.9 mL溶液中加入50 mM磷酸缓冲液(pH 7.8)、1.125 mM硝基蓝四氮唑(NBT)、60 μM核黄素、195 mM蛋氨酸和3 μM乙二胺四乙酸(EDTA)，取0.1 mL酶提取物。然后，在4000 lx光照下孵育30 min。以3ml不含酶提取物的溶液为对照，记录560nm处吸光度的变化。一个单位的SOD活性被定义为抑制NBT降低50%。[1]
根据Fan et al.(2014)描述的方法测量POD活性。简单地说，将50 μL酶提取物加入2.95 mL含有0.075% H2O2, 0.1 M醋酸钠-乙酸缓冲液(pH 5.0)， 0.25 mL愈创木酚(溶于50%乙醇溶液)的溶液中。然后我们记录在460 nm / min下吸光度的变化，持续3 min。单位POD活性定义为每分钟吸光度的增加。[1]
APX活性测定采用Plant APX Elisa Kit。[1]

最佳Spd浓度评价[1]
为了确定适当的亚精胺(Spd)浓度对缓解热应激的有效作用，我们进行了不同浓度Spd的初步实验。根据水稻上的Mostofa实验(Mostofa et al.， 2014)初步选择Spd浓度(0、0.5、1、2 mM)。随后，我们通过比较热应激4 h后的荧光瞬态，选择最佳浓度(0.5 mM)(图11)。图11显示了热胁迫下不同浓度Spd处理后叶绿素荧光瞬态的差异变化。0.5 mM Spd通过提高FJ、FI和FP对光合作用有积极影响。

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.

Toxicity Summary
Uremic toxins such as spermidine 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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Toxicity Summary
Uremic toxins such as spermidine 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). A7868: Schulz AM, Terne C, Jankowski V, Cohen G, Schaefer M, Boehringer F, Tepel M, Kunkel D, Zidek W, Jankowski J: Modulation of NADPH oxidase activity by known uraemic retention solutes. Eur J Clin Invest. 2014 Aug;44(8):802-11. doi: 10.1111/eci.12297. PMID:25041433

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Health Effects
Chronic exposure to uremic toxins can lead to a number of conditions including renal damage, chronic kidney disease and cardiovascular disease.

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Exposure Routes: Endogenous, Ingestion, Dermal (contact)

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Symptoms
As a uremic toxin, this compound can cause uremic syndrome. Uremic syndrome may affect any part of the body and can cause nausea, vomiting, loss of appetite, and weight loss. It can also cause changes in mental status, such as confusion, reduced awareness, agitation, psychosis, seizures, and coma. Abnormal bleeding, such as bleeding spontaneously or profusely from a very minor injury can also occur. Heart problems, such as an irregular heartbeat, inflammation in the sac that surrounds the heart (pericarditis), and increased pressure on the heart can be seen in patients with uremic syndrome. Shortness of breath from fluid buildup in the space between the lungs and the chest wall (pleural effusion) can also be present.

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Treatment
Kidney dialysis is usually needed to relieve the symptoms of uremic syndrome until normal kidney function can be restored.

[1]. Front Plant Sci. 2017 Oct 12:8:1747.

Spermidine is a triamine that is the 1,5,10-triaza derivative of decane. It has a role as a fundamental metabolite, a geroprotector and an autophagy inducer. It is a triamine and a polyazaalkane. It is a conjugate base of a spermidine(3+).
Spermidine is a polyamine formed from putrescine. It is found in almost all tissues in association with nucleic acids. It is found as a cation at all pH values, and is thought to help stabilize some membranes and nucleic acid structures. It is a precursor of spermine.
Spermidine is a metabolite found in or produced by Escherichia coli (strain K12, MG1655).
Spermidine has been reported in Camellia sinensis, Pseudomonas hydrogenovora, and other organisms with data available.
Spermidine is a polyamine derived from putrescine that is involved in many biological processes, including the regulation of membrane potential, the inhibition of nitric oxide synthase (NOS) and the induction of autophagy.
Spermidine 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.
Spermidine is a polyamine formed from putrescine. It is found in almost all tissues in association with nucleic acids. It is found as a cation at all pH values, and is thought to help stabilize some membranes and nucleic acid structures. It is a precursor of spermine.
Spermidine is a metabolite found in or produced by Saccharomyces cerevisiae.
A polyamine formed from putrescine. It is found in almost all tissues in association with nucleic acids. It is found as a cation at all pH values, and is thought to help stabilize some membranes and nucleic acid structures. It is a precursor of spermine.
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C7H19N3

145.25

145.157

C, 57.88; H, 13.19; N, 28.93

124-20-9

124-20-9; 334-50-9 (3HCl)

1102

Colorless to light yellow solid if <23°C, and liquid if >25°C

0.9±0.1 g/cm3

246.6±8.0 °C at 760 mmHg

23-25 °C

118.1±22.0 °C

0.0±0.5 mmHg at 25°C

1.475

-0.84

64.07

3

3

7

10

56.8

0

NCCCCNCCCN

ATHGHQPFGPMSJY-UHFFFAOYSA-N

InChI=1S/C7H19N3/c8-4-1-2-6-10-7-3-5-9/h10H,1-9H2

spermidine; 1,5,10-Triazadecane; 4-Azaoctamethylenediamine; N1-(3-Aminopropyl)butane-1,4-diamine

spermidine; 124-20-9; 1,5,10-Triazadecane; 4-Azaoctamethylenediamine; N1-(3-Aminopropyl)butane-1,4-diamine; Spermidin; 4-Azaoctane-1,8-diamine; N-(3-aminopropyl)butane-1,4-diamine;

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)

H2O:~100 mg/mL (688.5 mM)
DMSO: ~16.7 mg/mL (114.8 mM; with ultrasonic and warming as well as adjusting pH to 7 with HCl)

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。