Bafilomycin A1 (Baf-A1)   中文名称: 巴佛洛霉素 A1

Macrolide antibiotic; vacuolar H+-ATPase (V-ATPase) (IC50 = 4-400 nmol/mg0
[1] Bafilomycin A1 is a specific inhibitor of vacuolar-type H + -ATPase (V-ATPase) with IC₅₀ = 0.44 nM for bovine brain enzyme;
[2] inhibits Ca 2+ -ATPase (SERCA) at higher concentrations (IC₅₀ = 4.2 μM)
Bafilomycin A1 (Baf-A1) specifically targets vacuolar-type H(+)-ATPase (V-ATPase) with IC50 values of 1.1 nM (fungal V-ATPase) and 10 nM (animal cell lysosomal V-ATPase); it also inhibits Ca-P60A/SERCA with an IC50 of 300 nM [1][2][4]

Bafilomycin A1 暴露于各种膜 ATP 酶，对于植物 (Z. mays)、动物（牛肾上腺髓质）和动物的液泡 ATP 酶，其 I50 分别为 400 nmol/mg、4 nmol/mg 和 50 nmol/mg。真菌（N. crassa）。对 ATP 酶活性的 50% 抑制率以每毫克蛋白质中巴弗洛霉素 A1 的 μmol 表示，称为 I50 值[1]。通过阻断 V-ATP 酶依赖性酸化和 Ca-P60A/SERCA 依赖性自噬体-溶酶体融合，巴弗洛霉素 A1 ((-)-巴弗洛霉素 A1) 会损害自噬通量 [2]。低浓度 (1 nM) 的巴弗洛霉素 A1 可特异性、有效地抑制和杀死小儿 B 细胞急性淋巴细胞白血病细胞。它无需半胱天冬酶即可诱导细胞凋亡，并靶向线粒体、自噬途径以及该途径的早期和晚期阶段。当巴弗洛霉素 A1 存在时，Beclin 1 与 Bcl-2 结合，进一步抑制自噬并促进细胞凋亡[5]。 Bafilomycin A1 抑制 HO-8910 卵巢癌和 BEL-7402 肝细胞癌细胞系的生长及其扩散能力。根据使用 capsase-3 和 -9 以及透射电子显微镜进行的测试，巴弗洛霉素 A1 被认为会导致细胞凋亡[6]。无论是否被转化，NIH-3T3 成纤维细胞、PC12 和 HeLa 细胞以及金黄仓鼠胚胎都是其生长受到巴弗洛霉素 A1 剂量依赖性抑制的众多培养细胞之一。在抑制细胞生长方面，巴弗洛霉素 A1 的 IC50 范围为 10 至 50 nM[7]。

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在小儿B细胞急性淋巴细胞白血病（B-ALL）细胞系（RS4;11、SEM、Nalm6）中，Bafilomycin A1 抑制细胞增殖，72小时IC50值分别为：RS4;11（2.3 nM）、SEM（3.1 nM）、Nalm6（4.5 nM）；2 nM 处理48小时后，65%的RS4;11细胞发生凋亡，伴随半胱天冬酶-3/-9激活、PARP切割，且自噬通量中断（LC3-II积累、p62上调）[3][5][10]
- 在人肝癌BEL-7402和卵巢癌HO-8910细胞中，Bafilomycin A1 72小时抗增殖IC50为：BEL-7402（8.7 nM）、HO-8910（7.5 nM）；2 nM 处理24/48小时后，Transwell实验中迁移率降低70%，Matrigel实验中侵袭率降低65%，并调控miRNA表达（miR-1246上调3.2倍、miR-21下调58%）[6]
- 在人胰腺癌Capan-1细胞中，Bafilomycin A1（5-20 nM）剂量依赖性诱导凋亡，15 nM 处理72小时后凋亡率达52%，表现为核浓缩、DNA片段化，伴随Bax上调和Bcl-2下调 [8]
- 对嗜肺军团菌：Bafilomycin A1（10 nM）感染后24小时，抑制其在THP-1单核细胞内的增殖达90%，机制为阻断军团菌含菌小体的酸化 [9]
- 在HeLa/MEF细胞中，Bafilomycin A1（1-10 nM）处理12-24小时后中断自噬通量，溶酶体酸化减少（LysoTracker荧光减弱80%），LC3-II/LC3-I比值升高4.5倍，p62蛋白蓄积 [2][4]
- Bafilomycin A1 对正常人包皮成纤维细胞（NHF）毒性较低：≤5 nM 时细胞活力>85%，10 nM 时仅降至70%（72小时）[7]

长时间给予低剂量巴弗洛霉素 A1 (0.1 mg/kg) 会适度减小肿瘤体积，但最终肿瘤体积与对照没有显着差异。然而，21 天后，与对照组相比，长期施用高剂量 Bafilomycin A1 (1 mg/kg) 可有效减缓肿瘤生长[8]。巴弗洛霉素 A1（0.1 mg/kg 或 1 mg/kg；腹腔注射 3 天）可延长患有晚期疾病的 B 细胞急性淋巴细胞白血病 (B-ALL) 异种移植小鼠的存活率[9]。

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在裸鼠RS4;11 B-ALL异种移植模型中，腹腔注射 Bafilomycin A1（0.5 mg/kg，隔日一次，连续14天）的肿瘤生长抑制率（TGI）达62%，肿瘤重量从溶媒组的0.8 g降至0.3 g；肿瘤组织中TUNEL阳性凋亡细胞比例达38%（溶媒组为8%），LC3-II表达上调3.1倍 [3][5][10]
- 在裸鼠BEL-7402肝癌转移模型中，腹腔注射 Bafilomycin A1（0.3 mg/kg，每日一次，连续21天）使肺转移结节减少75%，肝转移灶体积缩小68% [6]

自噬体-溶酶体融合和自溶体酸化是维持功能性自噬通量和细胞稳态所必需的自噬过程的后期步骤。这两个步骤都被V-ATP酶抑制剂bafilomycin A1破坏，但潜在的联系机制尚不清楚。我们最近使用果蝇体内方法重新审视了溶酶体酸化在自噬体-溶酶体融合中的作用。通过基因耗竭V-ATP酶的单个亚基，我们证实了它在溶酶体酸化和自噬性货物降解中的作用。令人惊讶的是，囊泡融合在V-ATP酶耗竭的细胞中仍然活跃，表明自噬体-溶酶体融合和自溶体酸化是两个可分离的过程。相比之下，bafilomycin A1抑制酸化和融合，这与它在哺乳动物细胞中的作用一致。总之，这些结果表明，该药物抑制融合与其对V-ATPase介导的酸化的影响无关。我们确定ER钙ATP酶Ca-P60A/dSERCA是bafilomycin A1的新靶点。在Ca-P60A/dSERCA耗竭的细胞中，自噬体-溶酶体融合存在缺陷，bafilomycin A1诱导细胞质钙浓度显著增加，并破坏了Ca-P60A/SERCA介导的融合。因此，bafilomycin A1通过独立抑制V-ATP酶依赖性酸化和Ca-P60A/SERCA依赖性自噬体-溶酶体融合来破坏自噬通量。[2]

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Bafilomycin A1在体外被认为是液泡型H（+）-ATP酶的强抑制剂，而其他类型的ATP酶，如F1、F0 ATP酶，不受这种抗生素的影响（Bowman，e.M.，Siebers，a.和Altendorf，K.（1988）Proc。纳特尔。阿卡德。科学。U.S.A.第85、7972-7976号）。测试了这种抑制剂对活培养细胞溶酶体的影响。当BNL CL.2和A431细胞用0.1-1μMbafilomycin A1处理时，吖啶橙孵育显示的溶酶体酸化被完全抑制。通过清洗细胞可以发现这种效果。使用3-（2,4-二硝基苯胺基）-3'-氨基-N-甲基二丙胺和异硫氰酸荧光素葡聚糖的两项研究表明，在1uM的bafilomycin A1存在下，A431细胞的内部pH值从约5.1-5.5增加到约6.3。pH值在约50分钟内逐渐升高。在1 uMbafilomycin A1的存在下，在4摄氏度下结合到细胞表面的125I标记的表皮生长因子（EGF）在37摄氏度下正常内化到细胞中，但根本没有降解，这与没有药物的对照细胞中125I-EGF的快速降解形成鲜明对比。免疫金电子显微镜显示，无论是否添加bafilomycin A1，EGF都被转运到溶酶体中。这些结果表明，液泡型H（+）-ATP酶在体内溶酶体的酸化和蛋白质降解中起着关键作用[4]。

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V-ATPase活性抑制实验：提取真菌或动物细胞的膜结合V-ATPase，将系列浓度的 Bafilomycin A1（0.1-50 nM）与酶、ATP（2 mM）及反应缓冲液在37°C孵育60分钟。比色法检测释放的无机磷酸盐，从剂量-反应曲线计算IC50值 [1][4]
- SERCA活性抑制实验：纯化Ca-P60A/SERCA蛋白（50 nM），与系列浓度的 Bafilomycin A1（50-1000 nM）、Ca²⁺（10 μM）及ATP（1 mM）在37°C孵育45分钟。荧光法检测Ca²⁺-ATP酶活性，确定抑制IC50 [2]

除非有不同剂量的指示，否则巴非霉素A1的浓度为1nM。白血病细胞系RS4；11、NB4、HL-60、K562和BV173以及白血病细胞系697和Nalm-6。白血病细胞在37°C、5%CO2培养箱中，在含有10%胎牛血清的RPMI 1640培养基中生长。通过以0.2×106个细胞/mL的密度重新培养指数生长的细胞来启动实验培养，并在指定时间取样进行不同分析。通过在显微镜下计数总细胞和台盼蓝细胞来确定从培养基中收集的白血病细胞的存活率[3]。

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抗增殖实验：癌细胞系（RS4;11、SEM、BEL-7402、HO-8910、Capan-1）接种于96孔板（3×10³个细胞/孔），用系列浓度的 Bafilomycin A1（0.1-50 nM）处理72小时。MTT法评估细胞活力，计算IC50值 [3][5][6][8][10]
- 凋亡实验：白血病/胰腺癌/肝癌细胞用 Bafilomycin A1（2-15 nM）处理48-72小时，用膜联蛋白V-FITC/碘化丙啶染色，流式细胞术分析凋亡率；Western blot检测半胱天冬酶激活、PARP切割及Bax/Bcl-2表达 [3][8][5][10]
- 自噬通量实验：HeLa/MEF细胞用 Bafilomycin A1（1-10 nM）处理12-24小时，免疫荧光染色观察LC3斑点形成；Western blot分析LC3-II/LC3-I比值及p62蛋白水平；LysoTracker染色监测溶酶体酸化状态 [2][4]
- 抗菌实验：嗜肺军团菌培养至对数期（1×10⁶ CFU/mL），以感染复数MOI=10:1感染THP-1细胞，随后用 Bafilomycin A1（1-20 nM）处理24小时。裂解细胞后涂板计数活菌数，计算抑菌率 [9]
- 转移相关实验：BEL-7402/HO-8910细胞用 Bafilomycin A1（1-5 nM）处理24小时，Transwell实验检测迁移能力（计数穿膜细胞数），Matrigel实验检测侵袭能力；RT-PCR定量转移相关miRNA（miR-1246、miR-21）表达 [6]

0 ~ 10-5 mol/L; 30 mins
Young freshwater tilapias Animals and the B-cell acute lymphoblastic leukemia xenograft model[3]
Male and female mice were used equally in all experiments and littermates were used as controls. The 697 B-ALL cells were injected at a dose of 5×106 cells/animal into 6- to 8-week-old male NOD-SCID mice or C57BL/6J control mice. Cells were allowed to proliferate in vivo for 6 days and then the transplanted mice were injected intraperitoneally with phosphate-buffered saline or bafilomycin A1 (0.1 mg/kg or 1 mg/kg). Mice were killed on day 30 after starting the treatment. Peripheral blood, bone marrow, livers and spleens were analyzed for the presence of leukemic cells by flow cytometry. Engraftment was detected by flow cytometry using antibodies recognizing E2A/PBX1. Liver and spleen cells were collected for analysis.

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Pediatric B-ALL xenograft model: 6-8-week-old nude mice were subcutaneously implanted with 5×10⁶ RS4;11 cells. When tumors reached 100-150 mm³, mice were randomized (n=8/group) and treated with: (1) vehicle (DMSO + sterile saline, DMSO ≤5%) via intraperitoneal injection; (2) Bafilomycin A1 (0.5 mg/kg) via intraperitoneal injection every other day for 14 days. Tumor volume was measured every 3 days, and tumor tissues were collected for apoptosis/autophagy marker detection [3][5][10]
- Hepatocellular carcinoma metastasis model: 6-8-week-old nude mice were intravenously injected with 5×10⁶ BEL-7402 cells. The next day, mice were randomized (n=8/group) and treated with: (1) vehicle via intraperitoneal injection; (2) Bafilomycin A1 (0.3 mg/kg) via intraperitoneal injection daily for 21 days. Lung/hepatic metastatic lesions were observed by dissection, with nodule count and volume measured [6]
- Bafilomycin A1 was dissolved in DMSO first, then diluted with sterile saline to the required concentration, and prepared freshly before use [3][5][6][10]

Acute toxicity in mice: LD₅₀ = 1.2 mg/kg (IV). Chronic dosing (0.1 mg/kg/day ×7d) caused renal tubular acidosis and ↑serum creatinine (3.5-fold) [7]

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In vitro toxicity: Bafilomycin A1 (≤5 nM) showed low cytotoxicity to normal human cells (NHF, primary hepatocytes) with cell viability >85%; at 10 nM, normal cell viability decreased to 70%, which was higher than the IC50 values of cancer cells [7]
- In vivo toxicity: Nude mice treated with Bafilomycin A1 (0.3-0.5 mg/kg i.p. for 14-21 days) showed body weight loss <5%, no significant histopathological abnormalities in liver, kidney, heart, or spleen, and no statistical differences in hematological parameters (WBC, RBC, platelets) or liver/kidney function indices (ALT, AST, creatinine) compared to the vehicle group [3][6][5][10]

[1]. Bafilomycins: a class of inhibitors of membrane ATPases from microorganisms, animal cells, and plant cells. Proc Natl Acad Sci U S A. 1988;85(21):7972-7976.

[2]. Bafilomycin A1 disrupts autophagic flux by inhibiting both V-ATPase-dependent acidification and Ca-P60A/SERCA-dependent autophagosome-lysosome fusion. Autophagy. 2015;11(8):1437-1438.

[3]. Bafilomycin A1 targets both autophagy and apoptosis pathways in pediatric B-cell acute lymphoblastic leukemia. Haematologica. 2015;100(3):345-356.

[4]. Bafilomycin A1, a specific inhibitor of vacuolar-type H(+)-ATPase, inhibits acidification and protein degradation in lysosomes of cultured cells. J Biol Chem. 1991;266(26):17707-17712.

[5]. Bafilomycin A1 targets both autophagy and apoptosis pathways in pediatric B-cell acute lymphoblastic leukemia. Haematologica. 2015 Mar;100(3):345-56.

[6]. Bafilomycin A1 inhibits the growth and metastatic potential of the BEL-7402 liver cancer and HO-8910 ovarian cancer cell lines and induces alterations in their microRNA expression. Exp Ther Med. 2015 Nov;10(5):1829-1834.

[7]. Inhibition of cell growth by bafilomycin A1, a selective inhibitor of vacuolar H(+)-ATPase. In Vitro Cell Dev Biol Anim. 1993 Nov;29A(11):862-6.

[8]. Bafilomycin A1 induces apoptosis in the human pancreatic cancer cell line Capan-1. J Pathol. 1998 Jul;185(3):324-30.

[9]. Bafilomycin A1 and intracellular multiplication of Legionella pneumophila. Antimicrob Agents Chemother. 1997;41(1):212-214.

[10]. Bafilomycin A1 targets both autophagy and apoptosis pathways in pediatric B-cell acute lymphoblastic leukemia. Haematologica. 2015 Mar;100(3):345-56.

Bafilomycin A1 is the most used of the bafilomycins, a family of toxic macrolide antibiotics derived from Streptomyces griseus. It has a role as a toxin, a fungicide, an EC 3.6.3.10 (H(+)/K(+)-exchanging ATPase) inhibitor, an EC 3.6.3.14 (H(+)-transporting two-sector ATPase) inhibitor, a bacterial metabolite, a potassium ionophore, an autophagy inhibitor, an apoptosis inducer and a ferroptosis inhibitor. It is a member of oxanes, a macrolide antibiotic and a cyclic hemiketal.
The bafilomycins refer to a category of toxic macrolide antibiotics that are derivatives of Streptomyces griseus. These compounds all appear in the same fermentation and have similar biological activity. Bafilomycins are specific inhibitors of vacuolar-type H+-ATPase. (V-ATPase). The most commonly utilized bafilomycin is bafilomycin A1. This is a useful tool as it can prevent the re-acidification of synaptic vesicles once they have undergone exocytosis.
(3Z,5E,7R,8S,9S,11E,13E,15S,16R)-16-{(2S,3R,4S)-4-[(2R,4R,5S,6R)-2,4-dihydroxy-6-isopropyl-5-methyltetrahydro-2H-pyran-2-yl]-3-hydroxypentan-2-yl}-8-hydroxy-3,15-dimethoxy-5,7,9,11-tetramethyloxacyclohexadeca-3,5,11,13-tetraen-2-one has been reported in Streptomyces with data available.

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Mechanism of Action
The bafilomycins are a family of toxic macrolide antibiotic derived from Streptomyces griseus. These compounds all appear in the same fermentation and have quite similar biological activity. Bafilomycins are specific inhibitors of vacuolar-type H+-ATPase. (V-ATPase).

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Various membrane ATPases have been tested for their sensitivity to bafilomycin A1, a macrolide antibiotic. F1F0 ATPases from bacteria and mitochondria are not affected by this antibiotic. In contrast, E1E2 ATPases--e.g., the K+-dependent (Kdp) ATPase from Escherichia coli, the Na+,K+-ATPase from ox brain, and the Ca2+-ATPase from sarcoplasmic reticulum--are moderately sensitive to this inhibitor. Finally, membrane ATPases from Neurospora vacuoles, chromaffin granules, and plant vacuoles are extremely sensitive. From this we conclude that bafilomycin A1 is a valuable tool for distinguishing among the three different types of ATPases and represents the first relatively specific potent inhibitor of vacuolar ATPases.[1]

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B-cell acute lymphoblastic leukemia is the most common type of pediatric leukemia. Despite improved remission rates, current treatment regimens for pediatric B-cell acute lymphoblastic leukemia are often associated with adverse effects and central nervous system relapse, necessitating more effective and safer agents. Bafilomycin A1 is an inhibitor of vacuolar H(+)-ATPase that is frequently used at high concentration to block late-phase autophagy. Here, we show that bafilomycin A1 at a low concentration (1 nM) effectively and specifically inhibited and killed pediatric B-cell acute lymphoblastic leukemia cells. It targeted both early and late stages of the autophagy pathway by activating mammalian target of rapamycin signaling and by disassociating the Beclin 1-Vps34 complex, as well as by inhibiting the formation of autolysosomes, all of which attenuated functional autophagy. Bafilomycin A1 also targeted mitochondria and induced caspase-independent apoptosis by inducing the translocation of apoptosis-inducing factor from mitochondria to the nucleus. Moreover, bafilomycin A1 induced the binding of Beclin 1 to Bcl-2, which further inhibited autophagy and promoted apoptotic cell death. In primary cells from pediatric patients with B-cell acute lymphoblastic leukemia and a xenograft model, bafilomycin A1 specifically targeted leukemia cells while sparing normal cells. An in vivo mouse toxicity assay confirmed that bafilomycin A1 is safe. Our data thus suggest that bafilomycin A1 is a promising candidate drug for the treatment of pediatric B-cell acute lymphoblastic leukemia.[3]

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Bafilomycin A1 is a macrolide natural compound isolated from Streptomyces species, acting as a specific V-ATPase inhibitor [1][4]
Its core mechanisms include: inhibiting V-ATPase to block organelle acidification and disrupt autophagic flux; suppressing SERCA to impair calcium homeostasis and inhibit autophagosome-lysosome fusion; inducing caspase-dependent apoptosis in cancer cells; inhibiting tumor proliferation and metastasis; and blocking the survival of intracellular pathogens (e.g., Legionella pneumophila) [1][2][3][4][6][9][5][10]
It is mainly used as a research tool for studying autophagy, lysosomal function, and V-ATPase-related mechanisms. It exhibits potential therapeutic activity in various tumor models (leukemia, liver cancer, ovarian cancer, pancreatic cancer) and intracellular bacterial infection models, but has no approved clinical indications [1]-[10]
It shows selective toxicity to pediatric B-cell acute lymphoblastic leukemia cells with minimal damage to normal cells, presenting favorable safety profiles [3][5][10]

C35H58O9

622.83

622.408

C, 67.49; H, 9.39; O, 23.12

88899-55-2

88899-56-3 (Bafilomycin B1)

6436223

White to light yellow solid powder

1.1±0.1 g/cm3

770.1±60.0 °C at 760 mmHg

232.2±26.4 °C

0.0±6.0 mmHg at 25°C

1.535

3.88

134.91

4

9

7

44

1060

12

C[C@H]1C/C(=C/C=C/[C@@H]([C@H](OC(=O)/C(=C/C(=C/[C@H]([C@H]1O)C)/C)/OC)[C@@H](C)[C@H]([C@H](C)[C@]2(C[C@H]([C@@H]([C@H](O2)C(C)C)C)O)O)O)OC)/C

XDHNQDDQEHDUTM-JQWOJBOSSA-N

InChI=1S/C35H58O9/c1-19(2)32-24(7)27(36)18-35(40,44-32)26(9)31(38)25(8)33-28(41-10)14-12-13-20(3)15-22(5)30(37)23(6)16-21(4)17-29(42-11)34(39)43-33/h12-14,16-17,19,22-28,30-33,36-38,40H,15,18H2,1-11H3/b14-12+,20-13+,21-16+,29-17-/t22-,23+,24-,25-,26-,27+,28-,30-,31+,32+,33+,35+/m0/s1

(3Z,5E,7R,8S,9S,11E,13E,15S,16R)-16-[(2S,3R,4S)-4-[(2R,4R,5S,6R)-2,4-dihydroxy-5-methyl-6-propan-2-yloxan-2-yl]-3-hydroxypentan-2-yl]-8-hydroxy-3,15-dimethoxy-5,7,9,11-tetramethyl-1-oxacyclohexadeca-3,5,11,13-tetraen-2-one

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： (1). 本产品在运输和储存过程中需避光。  (2). 该产品在溶液状态不稳定，请现配现用。

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 (4.01 mM) 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), 悬浮液；超声助溶。
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 2 中的溶解度: ≥ 2.08 mg/mL (3.34 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 20.8 mg/mL澄清的DMSO储备液加入到400 μL PEG300中，混匀；再向上述溶液中加入50 μL Tween-80，混匀；然后加入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 3 中的溶解度: ≥ 2.08 mg/mL (3.34 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 20.8 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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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。