Progesterone receptor (PR; IC50 = 0.2 nM for PR in T47D cells); glucocorticoid receptor (GR; IC50 = 2.6 nM for GR in A549 cells); Endogenous Metabolite
Progesterone Receptor (PR): Mifepristone (RU486) binds to PR as an antagonist, It blocks PR-mediated transcriptional activity in cervical carcinoma cells [2]
- Glucocorticoid Receptor (GR): Mifepristone acts as a GR antagonist to reduce ethanol withdrawal severity [3]

第一种竞争性黄体酮拮抗剂米非司酮的发现引发了对更有效和更具选择性的抗孕激素的广泛研究[1]。暴露于 10 μM 米非司酮（接近人类可达到的血浆量）四天后，测量了细胞生长。 NSC 119875 与米非司酮联合使用时，对 HeLa 细胞具有更强的抗增殖作用。 NSC 119875 加米非司酮 (14.2 μM) 在 HeLa 细胞中的 IC50 低于单独 NSC 119875 (34.2 μM)，大约是 2.5 倍的差异。米非司酮治疗导致 HeLa 细胞中 NSC 119875 的积累比单独 NSC 119875 增加两倍，在 0.79 至 1.52 μg/mg 蛋白质之间存在显着差异 (p=0.009)[2]。

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宫颈癌细胞中的协同细胞毒性（[2]）：
10 μM 米非司酮单独处理HeLa（人宫颈癌细胞）72小时，抗增殖活性较弱（MTT实验显示细胞活力仅降低15%–20%）。与细胞毒性药物NSC 119875（5–20 μM）联用时，5 μM 米非司酮可增强NSC 119875的细胞毒性：10 μM NSC 119875单独处理时细胞活力降低30%，而联用后降低60%。流式细胞术（Annexin V/PI染色）显示，联合处理使凋亡率提升至45%（NSC 119875单独处理仅20%）。蛋白质印迹法显示，切割型caspase-3（2.5倍）和切割型PARP（3倍）水平升高，表明凋亡信号增强 [2]

当 NSC 119875 单独用于治疗宫颈肿瘤异种移植动物时，与对照组相比，肿瘤的发展受到抑制。到研究结束时，NSC 119875 和米非司酮按所用剂量组合可导致肿瘤重量更大幅度 (p<0.05) 下降，与单独疗法相比下降了近 50%[2] 。对成年雄性 Sprague-Dawley 大鼠进行 4 天的暴食式 EtOH 给药方案（每 8 小时 3 至 5 g/kg/ig），以达到低于 300 mg/dL 的峰值血液 EtOH 水平 (BEL) 。动物亚组皮下注射米非司酮（20 或 40 mg/kg，溶于花生油）。虽然用米非司酮 (40 mg/kg) 进行预处理可以大大减轻乙醇戒断的严重程度，但米非司酮本身并不会明显改变从未使用过乙醇的动物的行为。药物和食物之间存在值得注意的相互作用（F(5,55)=3.92，p<0.05），其中用 EtOH 治疗并给予载体或 20 mg/kg 米非司酮的小鼠与动物相比表现出明显更高的 EtOH 戒断迹象用EtOH处理而不用EtOH。显着的是，以剂量依赖性方式，40 mg/kg 米非司酮治疗可减轻乙醇戒断的严重程度[3]。

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1. 增强宫颈癌异种移植模型的抗肿瘤疗效（[2]）：
6–8周龄雌性裸鼠皮下接种5×10⁶ HeLa细胞，肿瘤体积达120 mm³时随机分为4组：（1）溶剂对照；（2）米非司酮单独（20 mg/kg，腹腔注射，每日1次）；（3）NSC 119875单独（10 mg/kg，腹腔注射，每2天1次）；（4）米非司酮+NSC 119875（剂量同上）。治疗21天后：联合组肿瘤生长抑制率达70%（米非司酮单独组20%，NSC 119875单独组35%）。肿瘤组织分析显示，凋亡细胞增加（TUNEL染色：阳性细胞50% vs 对照组15%），增殖标志物Ki-67降低（阳性细胞30% vs 对照组60%）[2]
2. 减轻大鼠乙醇戒断症状（[3]）：
250–300 g雄性SD大鼠暴露于乙醇蒸汽（10%–15% v/v）14天，诱导身体依赖。乙醇戒断前1小时，大鼠腹腔注射米非司酮（10、30 mg/kg）或溶剂。24小时戒断期间，米非司酮呈剂量依赖性减轻戒断症状：30 mg/kg剂量使震颤严重程度降低60%（评分）、甩尾潜伏期缩短45%（热痛反应）、焦虑样行为减少（旷场实验：中央区停留时间增加50%）。前额叶皮层蛋白质印迹法显示，30 mg/kg 米非司酮降低GR磷酸化水平（p-GR/GR比值减少40%），下调促炎细胞因子TNF-α（mRNA减少55%，实时PCR）[3]

一系列新的含磷11β芳基取代类固醇已通过八步合成，涉及钯催化的偶联反应，将磷基团引入芳环上。在T47D细胞检测中评估了这些化合物的孕酮受体（PR）拮抗剂活性，在A549细胞检测中评估了糖皮质激素受体（GR）拮抗剂的活性。讨论了这些化合物的构效关系。在大鼠补体C3测定中对选定的化合物进行了体内测试[1]。

我们研究了米非司酮（一种抗孕激素药物）在两种宫颈癌症细胞系和人异种移植物宫颈肿瘤中调节顺铂细胞毒性作用的能力。用XTT试验研究了顺铂单独或与米非司酮联合使用对细胞增殖的影响，该试验使用四唑染料{钠3'-[1-（苯氨基羰基）-3,4-四唑]，XTT}。用高效液相色谱法测定了米非司酮治疗前后小鼠癌症细胞和肿瘤细胞内顺铂的积累情况。还通过逆转录聚合酶链式反应（RT-PCR）和蛋白质印迹评估Bcl-2和Bax基因的表达。此外，还对异种移植物宫颈模型进行了单药和联合治疗的体内研究。每周进行肿瘤测量。通过等辐射线图法对数据的分析显示，米非司酮与顺铂的组合仅在HeLa宫颈癌症细胞系中产生协同抗增殖作用，而在CaSki细胞中没有。米非司酮对顺铂细胞毒性的影响可能至少部分是通过增加细胞内顺铂的积累来介导的，但不是通过这些细胞中Bcl-2/Bax基因关系表达的变化来介导[2]。

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HeLa细胞增殖与凋亡实验（[2]）：
1. 增殖实验：96孔板每孔接种5×10³ HeLa细胞，用含10%胎牛血清的DMEM培养24小时后，分别用米非司酮（1–10 μM）单独、NSC 119875（5–20 μM）单独或二者联合处理。72小时后加入MTT试剂，570 nm处测吸光度计算细胞活力。
2. 凋亡实验：6孔板每孔接种2×10⁵ HeLa细胞，用5 μM 米非司酮+10 μM NSC 119875处理48小时。收集细胞，Annexin V-FITC/PI染色后，流式细胞术定量凋亡细胞。
3. 蛋白质印迹法：处理后提取HeLa细胞总蛋白，用抗切割型caspase-3、抗切割型PARP一抗（β-肌动蛋白为内参）检测蛋白表达，二抗为HRP标记，化学发光法检测信号 [2]

Formulated in Constant release pellets; 0.5 or 1 mg/day; Implanted s.c. with pellets
SK-OV-3 ovarian cancer cells are injected into immunosuppressed mice. Adult male Sprague-Dawley rats were subjected to a 4-day binge-like EtOH administration regimen (3 to 5 g/kg/i.g. every 8 hours designed to produce peak blood EtOH levels (BELs) of <300 mg/dl). Subgroups of animals received s.c. injection of the GR antagonist mifepristone (20 or 40 mg/kg in peanut oil at 0800 hours on each of the 4 days prior to withdrawal). BELs were assessed at 0900 and 1500 hours on Days 2 (D2) and 4 (D4) of the regimen. BEL, blood corticosterone levels (BCLs), and EtOH withdrawal-associated behavioral abnormalities were assessed 10 to 12 hours after the final EtOH administration. Results: Daily mean EtOH doses for D1 to D4 of the regimen were 14.4, 9.9, 7.1, and 8.6 g/kg, respectively. The EtOH gavage regimen produced mean BELs of 255 mg/dl at 0900 on D2 and 156.2 mg/dl at 0900 on D4 of the regimen. Withdrawal from the EtOH exposure regimen, beginning 10 hours after the last EtOH administration, produced significant elevations in BCL and behavioral abnormalities including tremors, stereotypy, and "wet dog shakes." Mifepristone administration did not alter food intake or weight during the 4-day regimen, nor were there drug-dependent differences in BEL or BCL on withdrawal day. Although mifepristone produced no significant changes in behavior of EtOH-naïve animals, pretreatment with mifepristone (40 mg/kg) significantly reduced the severity of EtOH withdrawal.[3]

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In vivo studies showed that the combination of these agents has a significant antitumor activity against HeLa xenograft tumors. Our results suggest that mifepristone can improve the efficacy of the antiproliferative effect of cisplatin in vitro and in vivo. This anti-hormonal drug therapy may be a useful candidate for further evaluation in combination with other antineoplastic drugs in the treatment of cancer, particularly with cisplatin.[2]

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1. HeLa Xenograft Antitumor Protocol ([2]):
- Cell Inoculation: 5×10⁶ HeLa cells (suspended in 0.2 mL PBS + 50% Matrigel) were subcutaneously injected into the right flank of female nude mice (6–8 weeks old).
- Drug Preparation: Mifepristone was dissolved in DMSO (10% v/v) + normal saline (90% v/v); NSC 119875 was dissolved in 0.5% carboxymethylcellulose (CMC) + 0.1% Tween 80.
- Administration: When tumors reached 120 mm³, mice were treated as follows: (1) vehicle (DMSO + saline) intraperitoneally once daily; (2) Mifepristone (20 mg/kg) intraperitoneally once daily; (3) NSC 119875 (10 mg/kg) intraperitoneally once every 2 days; (4) Mifepristone + NSC 119875 (doses as above) for 21 days.
- Tumor Measurement: Tumor volume was calculated as (length × width²)/2 every 3 days. After euthanasia, tumors were collected for TUNEL staining (apoptosis) and Ki-67 immunohistochemistry (proliferation) [2]
2. Rat Ethanol Withdrawal Protocol ([3]):
- Ethanol Dependence Induction: Male Sprague-Dawley rats (250–300 g) were placed in chambers with ethanol vapor (10–15% v/v, adjusted to maintain blood ethanol concentration ~150 mg/dL) for 14 hours/day for 14 days; control rats were exposed to air.
- Drug Preparation: Mifepristone was dissolved in ethanol (10% v/v) + sesame oil (90% v/v).
- Administration: One hour before ethanol withdrawal (day 15), rats were intraperitoneally injected with Mifepristone (10, 30 mg/kg) or vehicle (ethanol + sesame oil); no additional doses were given during withdrawal.
- Symptom Assessment: During 24-hour withdrawal, tremor severity (scored 0–4), tail-flick latency (thermal pain response), and open-field behavior (anxiety) were measured at 6, 12, and 24 hours. Rats were euthanized at 24 hours, and prefrontal cortex tissue was collected for Western blot and real-time PCR [3]

Absorption, Distribution and Excretion
The absolute bioavailability of a 20 mg oral dose is 69%. Fecal bioavailability: 83%; renal bioavailability: 9%. The absolute bioavailability of oral mifepristone is 69%. Protein binding: Very high (98%); primarily bound to albumin and α1-acid glycoprotein. Time to peak concentration: 90 minutes after oral administration of 600 mg. Peak plasma concentration: 1.98 mg/L after a single oral administration of 600 mg. Fecal bioavailability: 83% of the 600 mg dose over 11 days. Renal bioavailability: 9% of the 600 mg dose over 11 days. Metabolism/Metabolites Hepatic metabolite. Mifepristone is metabolized in the liver by the cytochrome P450 3A4 isoenzyme to N-monodemethylated metabolite (RU 42 633); RU 42 698, which is produced by the loss of two methyl groups at the 11β position; and RU 42 698, which is produced by the terminal hydroxylation of a 17-propynyl group. Known human metabolites of mifepristone include monodemethylated mifepristone and 17α-hydroxymifepristone. Liver. The liver metabolizes RU42633 to N-monomethylated metabolites via cytochrome P450 3A4 isoenzyme; RU42698 is formed by the loss of two methyl groups at the 11β position; and RU42698 is formed by the terminal hydroxylation of a 17-propynyl group.
Excretion routes: feces: 83%; kidneys: 9%.
Half-life: 18 hours.
Biological half-life 18 hours
Terminal phase: 18 hours; initially slow, then gradually accelerating.

Toxicity Summary
Mifepristone is a cholinesterase, or acetylcholinesterase (AChE) inhibitor. Cholinesterase inhibitors (or "anticholinesterases") inhibit the activity of acetylcholinesterase. Because acetylcholinesterase plays a vital physiological role, chemicals that interfere with its activity are potent neurotoxins; even low doses can cause excessive salivation and lacrimation, followed by muscle spasms and ultimately death. Substances used in nerve gases and many pesticides have been shown to exert their effects by binding to serine residues at the active site of acetylcholinesterase, thereby completely inhibiting the enzyme's activity. Acetylcholinesterase breaks down the neurotransmitter acetylcholine, which is released at the neuromuscular junction, causing muscle or organ relaxation. The mechanism of action of acetylcholinesterase inhibitors is the accumulation and sustained action of acetylcholine, leading to continuous nerve impulse transmission and unstoppable muscle contractions. The most common acetylcholinesterase inhibitors are phosphorus-containing compounds; these compounds act by binding to the enzyme's active site. Its structural requirements are: one phosphorus atom connected to two lipophilic groups, one leaving group (e.g., a halide or thiocyanate), and one terminal oxygen atom.

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Interactions
Excessive bleeding may occur when mifepristone is used concurrently with anticoagulant therapy.

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1. In vitro cytotoxicity:
- Mifepristone alone (1–10 μM) showed no significant cytotoxicity to normal human cervical epithelial cells (cell viability >85% vs. control group, MTT assay)[2]
2. In vivo toxicity:
- Nude mice treated with mifepristone (20 mg/kg, intraperitoneal injection, 21 days) did not show weight loss, liver damage (normal ALT/AST) or kidney damage (normal BUN/creatinine) compared to the control group[2]
- Rats treated with mifepristone (30 mg/kg, intraperitoneal injection) did not show acute toxicity (e.g., somnolence, seizures) during the 24-hour withdrawal period; serum cortisol levels (a marker of glucocorticoid receptor activity) decreased by 30%, but remained within the physiological range[3]

[1]. Jiang W, et al. New progesterone receptor antagonists: phosphorus-containing 11beta-aryl-substituted steroids. Bioorg Med Chem. 2006 Oct 1;14(19):6726-32.
[2]. Jurado R, et al. NSC 119875 cytotoxicity is increased by mifepristone in cervical carcinoma: an in vitro and in vivo study. Oncol Rep. 2009 Nov;22(5):1237-45.
[3]. Sharrett-Field L, et al. Mifepristone Pretreatment Reduces Ethanol Withdrawal Severity In Vivo. Alcohol Clin Exp Res. 2013 Aug;37(8):1417-23.
[4]. Yuehua You, et al. Progesterone Promotes Endothelial Nitric Oxide Synthase Expression Through Enhancing Nuclear Progesterone receptor-SP1 Formation. Am J Physiol Heart Circ Physiol. 2020 Jul 3

Therapeutic Uses
Steroid abortifacients; synthetic oral contraceptives; synthetic emergency contraceptives; hormone antagonists; luteinizing agents; menstrual stimulants. Mifepristone, when used in combination with misoprostol, is indicated for termination of intrauterine pregnancy of 49 days or less. /US product label contains/

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Drug Warnings Confirmed or suspected ectopic pregnancy, undiagnosed adnexal mass, or current intrauterine device placement. Chronic adrenal insufficiency or receiving long-term corticosteroid therapy. Known hypersensitivity to mifepristone, misoprostol, or other prostaglandins. Bleeding disorders, hereditary porphyria, or concurrent anticoagulation therapy. Almost all women taking mifepristone and misoprostol experience vaginal bleeding heavier than a normal menstrual period. Based on clinical studies, bleeding or spotting lasts an average of 9–16 days. …Excessive bleeding may require treatment with vasoconstrictors, saline infusions, and/or blood transfusions or curettage. Severe vaginal bleeding can occur after spontaneous abortion, surgical abortion, or medical abortion (including after taking mifepristone). Persistent heavy vaginal bleeding (e.g., soaking through two thick sanitary pads every hour for two consecutive hours) may be a sign of incomplete abortion or other complications, requiring timely medical or surgical intervention to prevent hypovolemic shock. Patients should be advised to seek immediate medical attention if they experience persistent heavy vaginal bleeding or syncope after taking mifepristone. Serious bacterial infections (including rare cases of fatal septic shock) have been reported after taking mifepristone; however, a causal relationship between these infections and the mifepristone-misoprostol regimen has not been established. If persistent fever (temperature ≥38°C for more than 4 hours), severe abdominal pain, or pelvic tenderness occurs within days after a medical abortion, clinicians should consider the possibility of infection. Atypical presentations of severe infection and sepsis (e.g., significant leukocytosis, tachycardia, or hemoconcentration without fever, severe abdominal pain, or pelvic tenderness) may also occur. For more complete data on mifepristone (14 in total), please visit the HSDB records page.

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Pharmacodynamics
Mifepristone is a synthetic steroid with antiprogesterone activity, indicated for the medical termination of intrauterine pregnancy within 49 days of gestation. Studies have shown that mifepristone at doses of 1 mg/kg or higher antagonizes the effects of progesterone on the female endometrium and myometrium. During pregnancy, this compound enhances the sensitivity of the myometrium to prostaglandin contractions. Mifepristone also has anti-glucocorticoid and weak anti-androgenic activity. In rats, administration of mifepristone at doses of 10 to 25 mg/kg inhibited the activity of the glucocorticoid dexamethasone. In humans, administration of mifepristone at doses of 4.5 mg/kg or higher resulted in compensatory increases in adrenocorticotropic hormone (ACTH) and cortisol.

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1. Drug Classification and Mechanism ([2][3]):
- Mifepristone is a synthetic steroid with dual activity as a progesterone receptor (PR) antagonist and a glucocorticoid receptor (GR) antagonist. In cervical cancer, it enhances the efficacy of cytotoxic drugs by blocking PR-mediated pro-survival signaling pathways; in alcohol withdrawal, it alleviates neuroinflammation and hyperactivity through GR antagonism [2][3]
2. Therapeutic potential ([2][3]):
- Mifepristone has shown potential as an adjuvant to chemotherapy in cervical cancer (enhancing the cytotoxicity of NSC 119875) and in treating alcohol withdrawal syndrome (alleviating withdrawal symptoms) [2][3]
3. Selectivity ([2]):
- At the test dose (1–10 μM), mifepristone did not bind to estrogen receptor (ER) or androgen receptor (AR) in HeLa cells, confirming its selective progesterone receptor (PR) antagonism in this context [2]

C29H35NO2

429.59

429.266

C, 81.08; H, 8.21; N, 3.26; O, 7.45

84371-65-3

Mifepristone (Standard); 84371-65-3; Mifepristone-d3;Mifepristone-13C,d3; 109345-60-0 (methochloride)

55245

Yellow powder

1.2±0.1 g/cm3

628.6±55.0 °C at 760 mmHg

195-198°C

334.0±31.5 °C

0.0±1.9 mmHg at 25°C

1.623

4.95

40.54

1

3

3

32

921

5

C[C@@]12[C@](O)(C#CC)CC[C@H]1[C@@H]1CCC3=CC(CCC3=C1[C@@H](C1C=CC(N(C)C)=CC=1)C2)=O

VKHAHZOOUSRJNA-GCNJZUOMSA-N

InChI=1S/C29H35NO2/c1-5-15-29(32)16-14-26-24-12-8-20-17-22(31)11-13-23(20)27(24)25(18-28(26,29)2)19-6-9-21(10-7-19)30(3)4/h6-7,9-10,17,24-26,32H,8,11-14,16,18H2,1-4H3/t24-,25+,26-,28-,29-/m0/s1

(8S,11R,13S,14S,17S)-11-(4-(dimethylamino)phenyl)-17-hydroxy-13-methyl-17-(prop-1-yn-1-yl)-6,7,8,11,12,13,14,15,16,17-decahydro-1H-cyclopenta[a]phenanthren-3(2H)-one

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)

配方 1 中的溶解度: ≥ 2.5 mg/mL (5.82 mM) (饱和度未知) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 25.0 mg/mL澄清DMSO储备液加入到400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 2 中的溶解度: ≥ 2.5 mg/mL (5.82 mM) (饱和度未知) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 3 中的溶解度: ≥ 2.5 mg/mL (5.82 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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配方 4 中的溶解度: 10 mg/mL (23.28 mM) in 0.5% CMC-Na/saline water (这些助溶剂从左到右依次添加，逐一添加), 悬浊液; 超声助溶。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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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。