生物活性

Curcumin, a polyphenolic component ofturmeric (Curcuma longa), has promising therapeutic potential as anantimalarial, anticancer, antioxidant, and anti-inflammatory agent. Curcumin (antitumor, anti-inflammatory and antioxidant agent) is a yellow-orange dye obtained from tumeric, the powdered root of Curcuma longa; it is employed in the preparation of curcuma paper, the detection of boron, and possesses a vast number of biological targets. A study investigating the effects of curcumin on reported inflammatory mediators in inflammatory bowel disease on colonic myofibroblasts obtained ex vivo reported that a dose-dependent suppression of matrix metalloproteinase-3 (MMP-3) occured. Curcumin has been shown to inhibit NF-κB, possibly through inhibition of JNK (IC50=10 μM). Curcumin blocks amyloid peptide induced expression of: TNF-α, IL-1β, MCP-1, IL-8, and CCR5, regulates NOS2, inhibits IgE and Ag-induced degranulation of mast cells, inhibits 5-LO (5-lipoxygenase) (IC50=8 μM), Cox-2 (cyclooxygenase-2) (IC50=52 μM), the COP9 signalosome kinase (IC50=10μM), and p300/CREB Binding Protein (p300/CBP). Curcumin is an inhibitor of GST, IKK and NQO1.Also displays antimicrobial, antidiabetic neuro- and cardioprotective properties in vivo.

Curcumin is a potent inhibitor of CBR1 with anIC50-value of 382 ± 1.1 nM and a Ki-value of 223 ± 12 nM.[1] Curcuminexhibits DPPH radical scavenging activity with an IC50 value of 14.7 ± 1 μM.[2] Cellviability of curcumin[3]

Cell viability of curcumin

Anti-proliferation of curcumin

Anti-parasite activity of curcumin

Bindinginhibition of PKCα by curcumin[11]

体外研究

体内研究

激酶实验

Protein Purification[11]

The αC1A and αC1B domains in the pET28dvector were used for protein expression. The expressed proteins contain asix-His tag at the N-terminus. Recombinant plasmids were transformed into BL21Gold (DE3) cells. Protein expression was induced with IPTG (5mM) at an opticaldensity (OD) of 0.7, and then cells were allowed to grow overnight at 18 °C.Cells expressing αC1A and αC1B were resuspended in lysis buffer A [50mM Tris-HCl (pH8.0), 150mM NaCl, 50mM urea, and 10mM imidazole] and lysis buffer B [50mMTris-HCl (pH 8.0), 150mM NaCl, and 10mM imidazole], respectively. Cells were lysedby sonication (2min, 30% amplitude, 3s on and 3 s off cycle), and solubleproteins were obtained by centrifugation at 15000g for 15 min at 4 °C.Extracted lysates were incubated with Ni-NTA-bound agarose beads for 1 h andthen washed with respective lysis buffer to remove unbound proteins. Bound proteinswere eluted in lysis buffer containing 300mM imidazole. Eluted proteinfractions were pooled, concentrated, and further purified on a size exclusioncolumn using gel filtration buffer [50mM Tris-HCl (pH 8.0) and 15 mM NaCl]. Thepurity of the proteins was determined by SDS−PAGE (15%) and Coomassie bluestaining.

FluorescenceMeasurements

Fluorescence measurements were taken usinga PTI LPS 220B fluorimeter equipped with temperature and stirring controlsystems. For intrinsic fluorescence measurements, proteins (1μM) in buffer(50mM Tris, 150mM NaCl, pH 7. 2) were excited at 290 nm, and emission spectrawere recorded from 300 to 500 nm. For the binding assay, emission spectra ofcurcumin (5μM) were recorded in the presence of a varying concentration ofprotein (1−5μM) and TPA (1−10μM). Sample mixtures in buffer [50mM Tris and150mM NaCl (pH 7.2)] were incubated for 30 min at room temperature before eachmeasurement. Curcumin was excited at 425 nm, and emission spectra were recordedfrom 440 to 700nm. The maximum (Em_max) of the emission spectrum wasdetermined by a Gaussian function fit using IGOR Pro. The change in fluorescenceintensity at Emmax for each concentration of TPA was normalized using theequation (Fi − F0)/F0, where Fi and F0 are the fluorescence intensities ofcurcumin with and without TPA, respectively. IC50 was measured from the fittedcurve using the Hill equation in Sigma Plot 11

细胞实验

Cells and culture[4]

The breast cancer cell line MCF-7 was wellmaintained in culture growth media DMEM supplemented with 10% fetal bovineserum (FBS), 1% penicillin/streptomycin and incubated at 37°C, 5% CO₂,and 95% humidity.

Treatmentof MCF-7 breast cancer cell line with curcumin

The MCF-7 cells were grown until 60%–70% ofconfluence was reached. The cells were treated with curcumin at variousconcentrations, that is, 0, 1, 3, 5, 10, 20, 30, 50, and 100 μM for differenttime periods, that is, 24, 48, and72 h after that. The treated cells were usedfor further experiments. The cytotoxic effects of curcumin against MCF-7 weredetermined by MTT dye uptake method. The cells were incubated in triplicate ina 96-well plate in the presence or absence of curcumin in a final volume of 0.1mL for 24, 48, and 72 h at 37°C in a CO₂ incubator.

MTTassay

5000 exponentially growing cells per wellwere seeded in 96-well plates. After curcumin treatment and 4 h prior tocompletion of incubation period, 10 μL of MTT reagent was added to each well.After 4 h, MTT solution was removed, and the blue crystalline precipitate ineach well was dissolved in dimethyl sulfoxide. The optical density at awavelength 570 nm was measured using a 96-well multiscanner autoreader with thelysis buffer serving as blank. Cell viability was estimated using the followingformula: Percentage cell viability = absorbance values of test/absorbancevalues of control×100.

动物实验

Sepsis mice[3]

6–8 weeks old male C57BL/6 mice weighing 21± 2 g were maintained under specific pathogen-free conditions and 12 hlight/dark cycles. In the study’s animal experiments, the sepsis model wasbuilt by intraperitoneal injection of LPS (10 mg/kg). C57BL/6 mice were dividedinto three groups of 6 animals randomly as follows: Group a:carboxymethylcellulose solution; Group b: LPS (10mg/kg); Group c: LPS + curcumin(Cur) (20mg/kg). Mice were pretreated (3 days) with curcumin in 0.5%carboxymethylcellulose via oral gavage before an LPS intraperitoneal injectionand then sacrificed at 16 h.

Determiningbiochemical parameters in serum

Blood samples were obtained from mouse. Thelevels of serum aspartate transaminase (AST) and blood urea nitrogen (BUN) weredetermined using an automatic biochemical analyzer Beckman Coulter AU5800.

不同实验动物依据体表面积的等效剂量转换表（数据来源于FDA指南）

例如，已知某工具药用于小鼠的剂量为88 mg/kg , 则用于大鼠的剂量换算方法：将88 mg/kg 乘以小鼠的Km系数（3），再除以大鼠的Km系数（6），得到该药物用于大鼠的等效剂量44 mg/kg。

参考文献

[1] Hintzpeter J, Hornung J, Ebert B, Martin HJ, Maser E. Curcumin is a tight-binding inhibitor of the most efficient human daunorubicin reductase--Carbonyl reductase 1. Chem Biol Interact. 2015;234:162-168.
[2] Pany S, Majhi A, Das J. Selective Modulation of Protein Kinase C alpha over Protein Kinase C epsilon by Curcumin and Its Derivatives in CHO-K1 Cells. Biochemistry. 2016;55(14):2135-2143.
[more]

体内溶解度

Clinical Trial Information ( data from http://clinicaltrials.gov )

注：以上所有数据均来自公开文献，并不保证对所有实验均有效，数据仅供参考。

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