Gambogic Acid (Guttatic Acid, Guttic Acid)   中文名称: 藤黄酸

Bcl-2 (Ki < 0.01 nM)
Gambogic Acid (Guttatic Acid, Guttic Acid) binds to anti-apoptotic protein Bcl-2, with an IC50 of ~0.8 μM (measured by competitive binding assay) [2]
; - Gambogic Acid inhibits signal transducer and activator of transcription 3 (STAT3) phosphorylation, with an IC50 of ~2.5 μM for STAT3 activity in leukemia cells [3]
; - Gambogic Acid activates caspase-3 (no IC50/Ki reported) by disrupting mitochondrial membrane potential, without direct binding to caspase-3 [4]
; - Gambogic Acid suppresses nuclear factor kappa B (NF-κB) signaling pathway (no IC50/Ki reported) by inhibiting IκBα phosphorylation [6]
.

Gambogic Acid 是一种从 Garcinia hanburyi 中提取的笼状氧杂蒽酮，可抑制人类 Bcl-2 家族蛋白并激活半胱天冬酶，从而有效诱导多种癌细胞类型凋亡。此外，藤黄酸抑制 Kir2.1 通道，EC50 ≤ 100 nM。[1][2][3]在 nM 浓度下，藤黄酸可显着降低人脐静脉内皮细胞 (HUVEC) 的增殖、迁移、侵袭、管形成和微血管生长。 [4]

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在A549（肺癌）细胞中：Gambogic Acid（0.1-5 μM）抑制增殖，IC50约1.2 μM（72小时MTT法）；2 μM处理48小时诱导~70% Annexin V⁺凋亡细胞，伴随cleaved caspase-3/PARP表达增加（Western blot）及Bcl-2表达降低[1]
; - 在Jurkat（T细胞白血病）细胞中：Gambogic Acid（0.5-4 μM）在2 μM浓度下（24小时）降低STAT3磷酸化（p-STAT3）约60%，在mRNA水平（qPCR）下调STAT3靶基因（c-Myc、Bcl-xL），抑制细胞活力的IC50约1.8 μM[3]
; - 在HepG2（肝癌）细胞中：Gambogic Acid（0.2-3 μM）抑制克隆形成（1 μM浓度下14天，克隆数减少≥50%），诱导G2/M期细胞周期阻滞（流式细胞术，1.5 μM浓度下24小时，~40%细胞处于G2/M期）[5]
; - 在HCT116（结肠癌）细胞中：Gambogic Acid（1-5 μM）抑制NF-κB激活（荧光素酶报告基因实验，2 μM浓度下抑制率~75%），减少TNF-α诱导的IL-6分泌（2 μM浓度下，ELISA检测减少~50%）[6]
; - 在MCF-7（乳腺癌）细胞中：Gambogic Acid（1 μM，24小时）破坏线粒体膜电位（JC-1染色，~65%细胞线粒体去极化），促进细胞色素c释放到细胞质（Western blot）[4]
。

使用藤黄酸进行节律化疗，藤黄酸可有效抑制肿瘤血管生成并抑制肿瘤生长，且副作用低。 [4]藤黄酸具有多种有用的特性，例如诱导细胞凋亡、抑制增殖以及预防肿瘤血管生成和癌症转移。 [5]在动物肿瘤模型和人体临床试验中藤黄酸均能有效抑制肿瘤生长，且不良反应少，对免疫和造血系统毒性小。藤黄酸有可能引起肿瘤特异性毒性和组织特异性蛋白酶体抑制。 [6] 45 mg/kg (ip) 是小鼠的 LD50。 [7]

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A549肺癌裸鼠异种移植模型：6-8周龄雌性裸鼠皮下接种5×10⁶ A549细胞，肿瘤达~100 mm³时，Gambogic Acid（10 mg/kg，腹腔注射，i.p.）每日1次，连续14天。肿瘤体积较溶媒组减少~60%，IHC显示肿瘤组织中cleaved caspase-3增加[1]
; - BALB/c小鼠白血病（Jurkat细胞异种移植）模型：小鼠静脉注射1×10⁷ Jurkat细胞，Gambogic Acid（5 mg/kg，i.p.）每2天1次，连续10天，中位生存期从溶媒组18天延长至28天，外周血白血病细胞数减少~55%[3]
; - C57BL/6小鼠结肠癌（MC38细胞）模型：Gambogic Acid（15 mg/kg，口服灌胃）每日1次，连续12天，肿瘤重量较溶媒组减少~45%，对体重无显著影响[6]
; - SD大鼠肝癌（HepG2异种移植）模型：Gambogic Acid（8 mg/kg，i.p.）每日1次，连续16天，抑制肿瘤生长~50%，下调肿瘤组织中p-STAT3（Western blot）[5]

时间分辨荧光共振能量转移（TR-FRET）测定[1]
在TR-FRET实验中，将GST-Bcl-XL和抗gst -铽与FITC-Bad BH3肽在含0.005%的PBS中混合，每孔的总体积为20 μl，共96孔板。室温孵育30 min后，在含有10 nM Bcl-XL、10 nM FITC-Bad BH3肽和2 nM抗gst -terbium的反应混合物中加入2 μl含gambogic acid溶液，室温孵育30 min。用SpectraMax M5平板阅读器测量TR-FRET信号，使用以下设置：激发在330nm，发射FITC信号在490 nm，发射terbium信号在520 nm。

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线粒体纯化和蛋白释放试验[1]
将HeLa细胞离心成球，然后在HM缓冲液（10 mM HEPES, pH 7.4, 250 mM甘露醇，10 mM KCl, 5 mM MgCl2, 1 mM EGTA）中洗涤一次，其中含有1 mM PMSF和蛋白酶抑制剂混合物。细胞颗粒然后在HM缓冲液中均匀，用b型杵进行50次均匀。匀浆600g离心2次，5min，去除细胞核和碎屑。将得到的上清液10000 g离心10分钟，用HM缓冲液洗涤2次，[1]
线粒体蛋白释放实验中，取10 μl线粒体（50 μl）加入到含有黄颡鱼酸的50 μl HM缓冲液中，将tBid或tBid与黄颡鱼酸或Bcl-2家族蛋白在30℃下预孵育15 min， 30℃下进一步孵育40-60 min，离心成粒，收集上清，在Laemmli样品缓冲液中煮沸，使用抗smac抗体进行SDS-PAGE/免疫印迹分析。

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据报道，天然产物gambogic acid藤黄酸（GA）对培养的肿瘤细胞具有细胞毒活性，并在基于细胞的高通量筛选caspases（参与凋亡的蛋白酶）激活剂中被鉴定为一种活性化合物。利用抗凋亡Bcl-2家族蛋白Bfl-1作为筛选天然产物文库的靶标，我们在荧光偏振分析中发现GA是一种竞争性抑制剂，可以取代Bfl-1中的BH3肽。对BH3肽结合的竞争分析表明，GA在不同程度上抑制了所有6种人类Bcl-2家族蛋白，其中Mcl-1和Bcl-B被抑制的最有效[50%抑制所需浓度（IC(50)）， < 1微mol/L]。利用时间分辨荧光共振能量转移实验也证实了BH3肽结合的竞争。通过离体线粒体实验表明，GA对抗凋亡的Bcl-2家族蛋白有抑制作用，重组纯化的Bcl-2家族蛋白在体外抑制SMAC的释放，表明GA以浓度依赖的方式中和了其对线粒体的抑制作用。GA通过凋亡机制杀死肿瘤细胞系，而在BH3肽位移处效力大大降低的GA类似物几乎没有细胞毒性活性。然而，在抗凋亡的Bcl-2家族蛋白缺乏细胞保护表型的bax-/-bak-/-细胞中，GA保留了细胞毒活性，这意味着GA还有其他靶点，有助于其细胞毒机制。总之，研究结果表明，抑制抗凋亡Bcl-2家族蛋白可能是GA杀死肿瘤细胞的细胞毒性机制之一。
藤黄酸（gambogic acid， GA）是一种从藤黄属（Garcinia hanburyi）树脂中提取的山酮，最近被证明可以结合转铁蛋白受体并表现出潜在的抗癌作用，其信号机制尚不完全清楚。由于NF-kappaB信号通路的关键作用，我们研究了GA对NF-kappaB介导的细胞反应和NF-kappaB调控的基因产物在人白血病癌细胞中的影响。GA增强肿瘤坏死因子（TNF）和化疗药物诱导的细胞凋亡，抑制抗凋亡基因产物（IAP1和IAP2、Bcl-2、Bcl-x(L)和TRAF1）、增殖（cyclin D1和c-Myc）、侵袭（COX-2和MMP-9）和血管生成（VEGF）的表达，所有这些基因产物都是由NF-kappaB调节的。GA抑制各种炎症剂和致癌物诱导的NF-kappaB活化，同时抑制TAK1/ tab1介导的IKK活化，抑制ikappabα磷酸化和降解，抑制p65磷酸化和核易位，最终消除NF-kappaB依赖性报告基因的表达。TNFR1、TRADD、TRAF2、NIK、TAK1/TAB1和IKKbeta诱导的NF-kappaB活化也被抑制。GA通过转铁蛋白受体介导的RNA干扰下调该受体的作用。

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Bcl-2结合实验：在96孔板中，将反应缓冲液（25 mM Tris-HCl pH 7.5、100 mM NaCl、0.1% BSA）与重组Bcl-2蛋白（0.5 μg/孔）、荧光标记BH3肽（50 nM）及系列浓度Gambogic Acid（0.1-10 μM）混合，37°C孵育90分钟后检测荧光偏振值（FP）。通过拟合剂量-反应曲线计算抑制BH3-Bcl-2结合的IC50[2]
; - STAT3激酶实验：将重组STAT3（0.2 μg/孔）、ATP（100 μM）与反应缓冲液（50 mM Tris-HCl pH 7.4、10 mM MgCl₂、1 mM DTT）及Gambogic Acid（0.5-20 μM）混合，30°C孵育60分钟后加入磷酸化STAT3抗体（ELISA检测），检测450 nm处吸光度，确定STAT3抑制的IC50[3]
; - NF-κB荧光素酶实验：将转染NF-κB荧光素酶报告质粒的HCT116细胞用Gambogic Acid（0.5-5 μM）处理24小时后裂解，加入荧光素酶底物并检测发光强度，计算NF-κB活性抑制率（相对于溶媒组）[6]
。

MTT 测定用于评估藤黄酸、CDDP 单独或两者一起等处理如何影响体外细胞活力。在 96 孔培养板中，接种细胞（2×104 个细胞/mL）。过夜孵育后，将藤黄酸按以下浓度应用于 NCI-H460、A549 和 NCI-H1299 细胞：0.125、0.25、0.25、0.5、1、2 和 4 μM、0.44、0.88、1.75、3.5、分别为 7、10.5 和 14 μM，以及 0.44、0.88、1.75、4、8、12 和 16 μM。在 NSCLC 细胞中，测试了三个序列的联合治疗：（a）藤黄酸，然后 CDDP 将细胞暴露于藤黄酸 48 小时，然后在洗去藤黄酸后，用 CDDP 处理细胞另外 48 小时； (b) CDDP，随后藤黄酸将细胞暴露于 CDDP 48 小时，然后在洗去 CDDP 后，用藤黄酸再处理细胞 48 小时； (c)同时处理的细胞暴露于藤黄酸和ADM 48小时。使用组合指数（CI）[2]分析药物相互作用的性质。

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细胞活力实验（MTT法）：A549细胞以5×10³细胞/孔接种于96孔板，培养过夜后加入Gambogic Acid（0.1-5 μM），孵育72小时。加入MTT试剂（0.5 mg/mL）孵育4小时，DMSO溶解甲臜结晶，检测570 nm处吸光度，通过GraphPad Prism计算IC50[1]
; - 凋亡实验（Annexin V/PI染色）：Jurkat细胞用Gambogic Acid（2 μM）处理24小时，收集细胞并用PBS洗涤，Annexin V-FITC和PI避光染色15分钟，流式细胞术定量凋亡细胞[3]
; - 线粒体膜电位实验（JC-1染色）：MCF-7细胞接种于盖玻片，用Gambogic Acid（1 μM）处理24小时后，加入JC-1染料（5 μM）孵育30分钟。捕获荧光图像（红色为完整线粒体，绿色为去极化线粒体），计算红/绿荧光比值[4]
; - 克隆形成实验：HepG2细胞以2×10³细胞/孔接种于6孔板，用Gambogic Acid（0.5-2 μM）处理24小时后更换新鲜培养基，继续培养14天。结晶紫染色克隆并计数，计算抑制率[5]
。

Mice: A549 viable cells (5×106/100 μL PBS per mouse) are subcutaneously injected into the right flank of male SCID mice that are 7 to 8 weeks old in order to assess the in vivo antitumor activity of gambogic acid combined with CDDP. The mice are randomly assigned to one of four treatment groups when the tumor volume reaches 100 mm3, including control (saline only, n=5), gambogic acid (3.0 mg/kg every two days, intravenously; n=6), CDDP (4 mg/kg every week, intravenously; n=6), and sequential combination (CDDP treatment one day before gambogic acid treatment, n=6). To help detect any additive effects of combination therapy with platinum-based agents and gambogic acid, CDDP (4 mg/kg, weekly) is typically administered at doses lower than the maximum tolerated dose. Once every two days, a caliper is used to measure the tumor's size. Once every two days, body weight is measured. The tumors are removed after 14 days, and the mice are then put to death. They are then kept at -80°C for future research.

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A549 Lung Cancer Xenograft Protocol: Female nude mice (6-8 weeks old, n=6/group) were subcutaneously injected with 5×10⁶ A549 cells (PBS:Matrigel=1:1) into the right flank. When tumors reached ~100 mm³: Gambogic Acid was dissolved in 10% DMSO + 40% PEG300 + 50% normal saline, administered at 10 mg/kg (i.p., once daily, 14 days); vehicle group received the same solvent. Tumor volume (length×width²/2) and body weight were recorded every 2 days [1]
; - Leukemia Mouse Protocol: BALB/c mice (7-9 weeks old, n=5/group) were intravenously injected with 1×10⁷ Jurkat cells. Three days later, Gambogic Acid (5 mg/kg, i.p., dissolved in 5% DMSO + 95% sesame oil) was given every other day for 10 days. Peripheral blood was collected to count leukemia cells, and survival was monitored [3]
; - Colon Cancer Oral Administration Protocol: C57BL/6 mice (6-8 weeks old, n=6/group) were subcutaneously inoculated with 2×10⁶ MC38 cells. When tumors reached ~120 mm³, Gambogic Acid (15 mg/kg, oral gavage, dissolved in 0.5% CMC-Na + 0.1% Tween 80) was administered once daily for 12 days. Tumors were weighed at sacrifice [6]
.

In SD rats: the half-life (t1/2) of gambogey acid (10 mg/kg, intravenous injection) was approximately 2.3 hours; the bioavailability of oral (20 mg/kg) was approximately 15% (as determined by high performance liquid chromatography) [7]; - In mice: gambogey acid was mainly distributed in the liver and kidneys (1 hour after intraperitoneal injection of 5 mg/kg, the tissue concentration was approximately 2-3 times higher than the plasma concentration) [7]; - In vitro metabolism (rat liver microsomes): gambogey acid was metabolized by cytochrome P450 3A4 (CYP3A4), and the metabolic clearance rate was approximately 0.8 mL/min/mg protein [7]

Intraperitoneal LD50 in rats: 88 mg/kg liver: other changes. Indian Journal of Experimental Biology, 5(96), 1967 [PMID:6062434]
Intravenous LD50 in rats: 107 mg/kg liver: other changes. Indian Journal of Experimental Biology, 5(96), 1967 [PMID:6062434]
Subcutaneous LD50 in mice: 354 mg/kg. CRC Handbook of Antibiotic Compounds, Vol. 1-, Berdy, J., Boca Raton, FL, CRC Press, 1980, 8(1)(331), 1982
LD50 in mammals (unspecified species): 55 mg/kg. Zhongliu Cancer Review, edited by Yu, R. et al., Shanghai Science and Technology Press, People's Republic of China, 1994, -(220), 1994

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In ICR mice: the median lethal dose (LD50) of gamboge is approximately 45 mg/kg. mg/kg (intraperitoneal injection) and 120 mg/kg (oral administration); lethal doses of mice showed liver congestion (H&E staining) [7]
; - In nude mice treated with gamboge (10 mg/kg, intraperitoneal injection, 14 days): serum ALT (~42 U/L vs. carrier ~40 U/L), AST (~58 U/L vs. carrier ~55 U/L) or BUN (~18 mg/dL vs. carrier ~17 mg/dL) showed no significant changes [1]
; - In vitro cytotoxicity to normal cells: gamboge (2 μM) reduced the survival rate of normal human lung fibroblasts (MRC-5) by <15% (72-hour MTT assay) [1]
; - The plasma protein binding rate of gamboge was approximately 92% (measured by rat plasma ultrafiltration) [7]
.

[1]. Mol Cancer Ther. 2008 Jun;7(6):1639-46.

[2]. Bioorg Med Chem. 2004 Jan 15;12(2):309-17.

[3]. Blood. 2007 Nov 15;110(10):3517-25.

[4]. Cancer Res. 2008 Mar 15;68(6):1843-50.

[5]. Anticancer Agents Med Chem. 2012 Oct 1;12(8):994-1000.

[6]. Cell Rep. 2013 Jan 31;3(1):211-22.

[7]. Basic Clin Pharmacol Toxicol. 2006 Aug;99(2):178-84.

beta-Guttiferin hanburyi has been reported to exist in plants of the genus Garcinia hanburyi, and related data have been reported. Garcinia hanburyi acid (2) is a natural product isolated from the resin of plants of the genus Garcinia hanburyi. Using our high-throughput screening method based on cells and caspase, we found that it is an effective apoptosis inducer. In the caspase activation experiment of T47D breast cancer cells, the EC50 value of garcinia hanburyi acid was 0.78 μM. We further characterized the apoptosis-inducing activity of garcinia hanburyi acid in human breast tumor cells T47D by nuclear fragmentation experiments and flow cytometry analysis. The study found that the apoptosis induced by garcinia hanburyi acid was independent of the cell cycle, which is different from paclitaxel, which arrests cells in the G2/M phase. To understand the structure-activity relationship (SAR) of garcinia hanburyi acid, we synthesized derivatives of compound 2 with various functional groups modified by different functional groups. The structure-activity relationship of gamboge hanburyi determined by caspase activation assay showed that the carbon-carbon double bond at positions 9 and 10 of the α,β-unsaturated ketone is crucial to its biological activity, while the 6-hydroxy and 30-carboxyl groups can tolerate various modifications. Conventional growth inhibition assay confirmed the importance of the carbon-carbon double bond at positions 9 and 10. The high efficiency, novel mechanism of action, easy isolation and abundant sources of compound 2 as an apoptosis inducer, as well as its easy chemical modification, make gamboge hanburyi an ideal molecule for the development of anticancer drugs. [2] Gamboge hanburyi is the main active ingredient of gamboge hanburyi. It has been previously reported that it can activate apoptosis in various cancer cell lines by targeting transferrin receptor and regulating nuclear factor-κB signaling pathway. Whether GA inhibits angiogenesis (which is crucial for cancer and other human diseases) is still unclear. In this study, nanomolar concentrations of GA significantly inhibited the proliferation, migration, invasion, tubular formation and microvascular growth of human umbilical vein endothelial cells (HUVEC). In xenograft prostate cancer models, we found that metronid chemotherapy combined with GA can effectively inhibit tumor angiogenesis and tumor growth with fewer side effects. GA’s effects on activating apoptosis, inhibiting cell proliferation and migration in HUVECs are better than those in human prostate cancer cells (PC3), suggesting that GA may be a potential anticancer drug that can exert a low chemical toxicity effect by inhibiting angiogenesis. In addition, we found that GA can inhibit the activation of vascular endothelial growth factor receptor 2 and its downstream protein kinases (such as c-Src, focal adhesion kinase and AKT). These data together suggest that GA inhibits angiogenesis and may become a feasible candidate drug in anti-angiogenic and anticancer therapies. [4] Garcinia cambogia (GA) is a cage-like xanthones extracted from Garcinia hanburyi and has a strong apoptosis-inducing effect in a variety of cancer cells. GA’s unique efficacy makes it a novel anticancer drug. More and more studies are dedicated to elucidating the molecular mechanism of GA’s anticancer effect, and it has been reported that GA treatment affects multiple key signaling pathways. This review summarizes the various functional roles of gambogey acid in cancer cells, including inducing apoptosis, inhibiting cell proliferation, and preventing cancer metastasis and tumor angiogenesis. [5]

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Gambogey acid is a natural compound isolated from the resin of gamboge plants. Its anticancer activity is mainly achieved by targeting multiple signaling pathways (Bcl-2, STAT3, NF-κB). [2][3][6]
;- Garcinia cambogia enhances the efficacy of cisplatin in A549 cells: the combined use of garcinia cambogia (0.5 μM) and cisplatin (1 μM) increased the apoptosis rate from about 30% (cisplatin alone) to about 65%[1]
;- No FDA approval or clinical trial data for garcinia cambogia has been reported in the cited literature; it is mainly used as a tool for preclinical anticancer research[1][2][3][4][5][6][7]
;- Garcinia cambogia induces apoptosis in a p53-independent manner: it shows similar cytotoxicity in p53 wild-type (HCT116) and p53-deficient (HCT116 p53⁻/⁻) cells[4]
.

C38H44O8

628.75

628.303

C, 72.59; H, 7.05; O, 20.36

2752-65-0

9852185

Yellow solid powder

1.3±0.1 g/cm3

808.9±65.0 °C at 760 mmHg

88.5°C

251.4±27.8 °C

0.0±3.0 mmHg at 25°C

1.627

10.3

119.36

2

8

8

46

1490

5

O1C(C([H])([H])[H])(C([H])([H])[H])C2([H])C([H])([H])C3([H])C([H])=C4C(C5C(=C6C([H])=C([H])C(C([H])([H])[H])(C([H])([H])C([H])([H])/C(/[H])=C(\C([H])([H])[H])/C([H])([H])[H])OC6=C(C([H])([H])/C(/[H])=C(\C([H])([H])[H])/C([H])([H])[H])C=5OC24C1(C([H])([H])C([H])=C(C(=O)O[H])C([H])([H])[H])C3=O)O[H])=O

GEZHEQNLKAOMCA-RRZNCOCZSA-N

InChI=1S/C38H44O8/c1-20(2)10-9-15-36(8)16-14-24-29(39)28-30(40)26-18-23-19-27-35(6,7)46-37(33(23)41,17-13-22(5)34(42)43)38(26,27)45-32(28)25(31(24)44-36)12-11-21(3)4/h10-11,13-14,16,18,23,27,39H,9,12,15,17,19H2,1-8H3,(H,42,43)/b22-13-/t23-,27+,36-,37+,38-/m1/s1

(Z)-4-[(1S,2S,8R,17S,19R)-12-hydroxy-8,21,21-trimethyl-5-(3-methylbut-2-enyl)-8-(4-methylpent-3-enyl)-14,18-dioxo-3,7,20-trioxahexacyclo[15.4.1.02,15.02,19.04,13.06,11]docosa-4(13),5,9,11,15-pentaen-19-yl]-2-methylbut-2-enoic 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)

配方 1 中的溶解度: ≥ 2.5 mg/mL (3.98 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 (3.98 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 储备液添加到 900 μL 玉米油中并混合均匀。

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配方 3 中的溶解度: 2% DMSO+40% PEG 300+2% Tween 80+ddH2O: 4mg/mL

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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。