delivery of curcumin and medicinal effects of the copper(ii)-curcumin complexes

Send Orders of Reprints at [email protected]

2070 Current Pharmaceutical Design, 2013, 19, 2070-2083

Delivery of Curcumin and Medicinal Effects of the Copper(II)-Curcumin Complexes

Mandy H. M. Leung†, Takaaki Harada

† and Tak W. Kee*

Department of Chemistry, University of Adelaide, Adelaide, South Australia, 5005, Australia

Abstract: Curcumin, a yellow pigment extracted from the rhizome of Curcuma longa, commonly known as turmeric, is the most active agent of this herbal medicine. The therapeutic activities of curcumin are exemplified not only by its enhancement in wound healing but

also in the treatment of inflammation, cystic fibrosis, Alzheimer’s disease and cancer. There are two critical issues involving low aqueous stability and solubility that limit the bioavailability and application of curcumin as a therapeutic agent. To address these issues, delivery

systems of curcumin including surfactant micelles, liposomes, polymer nanoparticles, casein micelles, plasma proteins and cyclodextrins have been developed and characterized. From a biochemical perspective, the medicinal activities of curcumin are proposed to be related

to an elevated level of transition metals including copper, zinc and iron in many disease sites, especially those in cancer and Alzheimer’s disease. Previous studies have demonstrated the importance of copper(II)-curcumin complexes in DNA damage owing to the strong in-

teraction between curcumin and copper(II). Curcumin, as an anti-oxidant, possesses the abilities to scavenge radicals and maintain the levels of anti-oxidant enzymes in the presence of copper. On the other hand, copper(II)-curcumin complexes show pro-oxidant effects by

generating reactive oxygen species at a high free copper level in a reducing environment. This condition results in DNA damage and in-hibition of vital signaling pathways in cancer cells, leading to apoptosis. In short, curcumin has dual roles as an anti-oxidant and a pro-

oxidant in the presence of copper and these fascinating phenomena contribute greatly to its multiple medicinal effects.

Keywords: Curcumin, copper, drug delivery, anti-cancer, anti-oxidant, pro-oxidant, inflammation, Alzheimer’s disease.

HISTORY AND BIOSYNTHESIS OF CURCUMIN AND

CURCUMINOIDS

Turmeric powder is the dried ground rhizome of Curcuma longa, a member of the ginger family Zingiberaceae (Fig. 1) [1]. This intensely yellow powder has a long history of useful benefits and widespread applications in India and other Asian countries as turmeric is a key ingredient in Indian and Thai cuisines [2]. It was also introduced in a remedial theory called Ayurveda, the knowl-edge for long life, in which a system of traditional medicine and health maintenance was established in India over 2000 years ago [3]. The yellow color and therapeutic activities of turmeric powder are derived from a group of active molecules, called curcuminoids.

Curcuminoids are present at 3 5% in turmeric powder after liquid extraction and filtration [1,4]. In 1815, Vogel and Pelletier etal. were the first to isolate curcuminoids [5]. Curcuminoids consist of three major components, with curcumin being the predominant species (77%), which is followed by demethoxycurcumin (17%) and bisdemethoxycurcumin (3%) (Fig. 2). In addition to these three curcuminoids, a recent study showed that a trace amount of a fourth curcuminoid, cyclocurcumin, is also present [6]. In 1973, Roughly and Whiting et al. investigated the biosynthesis of the major cur-cuminoids using

14C-labeled precursors and proposed two possible

pathways (Scheme 1) [4]. Scheme 1a depicts the synthesis of cur-cumin using two portions of ferulic acid and a portion of malonic acid. In addition, Scheme 1b shows the reaction between cinnamic acid and multiple malonic acid units through a condensation reac-tion to produce 7-methoxy-8-hydroxy-curcumin (curcumin without the methoxy and hydroxyl groups on one side). This research was further investigated [7], in which

13C-nuclear magnetic resonance

(NMR) results were used to propose a more detailed biosynthesis of curcuminoids (Scheme 2) [7,8]. In the major pathway, the curcumin skeleton intermediate called bisdeshydroxybisdesmethoxycurcumin is first synthesized from two units of cinnamic acid and one unit of malonic acid, which is followed by modification at the phenyl

*Address correspondence to this author at the Department of Chemistry,

University of Adelaide, Adelaide, South Australia, 5005, Australia; Tel: +61 (0)8 8313 5039; Fax: +61 (0)8 8313 4358; E-mail: [email protected] †These authors contributed equally to this work.

Fig. (1). Picture of Curcuma longa and turmeric powder. This figure is

reprinted from ref. [1] and used with permission of Cell Division.

groups to form bisdemethoxycurcumin, demethoxycurcumin and then finally curcumin. Alternatively, cinnamic acid is modified to form p-coumaric acid and ferulic acid which randomly react with malonic acid to generate other curcuminoids in the minor pathway. This recently proposed biosynthesis scheme supports the earlier proposed pathway by Roughly and Whiting et al. (Scheme 1a) [4].

1873-4286/13 $58.00+.00 © 2013 Bentham Science Publishers

Effects of Copper(II)-Curcumin Complexes Current Pharmaceutical Design, 2013, Vol. 19, No. 11 2071

Scheme 1. Proposed mechanisms of biosynthesis of curcumin by Roughley et al. [4].

HO

O O

OHd

OH O

HO OH

O OCH3H3C

h

OH

O

HO

OH3C2

c

HO

O O

OHd

OH O

HO OH

OH3C

g

OH

O

HO

O

c

OH

O

HOb

HO

O O

OHd

OH O

HO OHf

OH

O

HO

2

b

HO

O O

OHd

OH O

e

OH

O

2

a

H3C

Scheme 2. Proposed major (solid arrows) and minor (dashed arrows) mechanisms of biosynthesis of curcumin and curcuminoids by Kita et al. [7]. The letters

represent the following molecules; (a) cinnamic acid, (b) p-coumaric acid, (c) ferulic acid, (d) malonic acid, (e) bisdeshydoxybisdesmethoxycurcumin, (f)

bisdemethoxycurcumin, (g) demethoxycurcumin and (h) curcumin.

PHYSICAL PROPERTIES OF CURCUMIN

Curcumin, (1E,6E)-1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione, is an orange-yellow solid at room tempera-ture (MW = 368.38 g/mol, mp = 183 °C), also known as E100 or

Natural Yellow 3. Curcumin exists in two tautomeric forms, the keto-enol and di-keto tautomers (Scheme 3). The predominant tautomer of curcumin is the keto-enol form when it is present in polar organic solvents such as methanol and DMSO [9]. This

S

O

OH3C

HO

2

HO

O

S

O

CoA

CoAO

HO

H3C

OH O

O

OH

CH3

Curcumin

a

b

S

OHO

O

S

O

CoA

CoA

5OH O

O

OH

CH3

7-methoxy-8-hydroxy-curcumin

2072 Current Pharmaceutical Design, 2013, Vol. 19, No. 11 Leung et al.

tautomer possesses intramolecular hydrogen bonding in the keto-enol moiety and -conjugation is maintained across the molecule, which results in an ultraviolet-visible (UV-Vis) absorption peak around 420 nm [10]. The strongly allowed – * transition gives rise to a high molar extinction coefficient of 30,000 – 70,000 M

-1cm

-1 at the absorption peak in water, methanol or ethanol (Fig.

3) [11]. Although curcumin is essentially non-fluorescent in water, it has a fluorescence quantum yield of 2 – 10% in polar organic solvents, lipid membranes, and hydrophobic environment including non-polar solvents [10]. Several groups have used time-resolved fluorescence spectroscopy to investigate the rapid intramolecular hydrogen atom transfer in the keto-enol moiety of curcumin in the excited state [12-15]. In addition, the keto-enol tautomer possesses three hydroxyl groups of which the enolic and phenolic hydrogens can be deprotonated under alkaline conditions. The pKa values of the enolic and phenolic hydrogens are 8.3 and ~10, respectively, in aqueous environment [16]. Hence, curcumin is expected to remain protonated at physiological pH (7.4).

Fig. (2). Structures of curcuminoids.

In contrast, the conjugation length of the minor di-keto tautomer is significantly shorter, extending over only half the length of curcumin. The shorter conjugation length is due to the presence of an sp

3 hybridized carbon between the two carbonyl groups in the

di-keto moiety of this tautomer (Scheme 3) [17,18], which results in an absorption maximum around 350 nm. The presence of the di-keto form is indicated by an absorption shoulder around 350 nm when curcumin is solubilized in water, as shown in Fig. 3.

SOLUBILITY AND STABILITY OF CURCUMIN

Although turmeric powder has a long history of medicinal ap-plications in Asia, two critical issues involving low aqueous solu-bility and stability must be addressed in order for curcumin to be utilized as an effective therapeutic agent. Because curcumin is a moderately hydrophobic molecule with polar moieties, it has a high solubility and stability in polar organic solvents such as methanol, ethanol, acetone and DMSO. Although these solvents can solubilize curcumin at a high concentration, it is crucial to consider its solubil-ity and stability in the aqueous environment for biological applica-tions. The water solubility of curcumin is only approximately 30 pM, which is equivalent to 11 g/mL [19,20]. In the presence of micelles, vesicles or polymer nanoparticles, curcumin partitions largely into the hydrophobic domains under either slightly acidic or neutral condition due to its hydrophobicity [21]. However, under

Scheme 3. Tautomerization of keto-enol and di-keto forms of curcumin.

Fig. (3). UV-Vis absorption spectra of curcumin in water (pH = 4), metha-

nol and ethanol. This figure is adapted with permission from ref. [11].

Copyright 2010 ACS.

alkaline condition, the aqueous solubility of curcumin is improved due to deprotonation of the enolic and phenolic groups such that it becomes negatively charged. Under this condition, curcumin de-composes rapidly in water by a retro-aldol hydrolysis reaction, which is catalyzed by hydroxide and followed by molecular frag-mentation [22,23]. The degradation of curcumin leads to formation of several metabolites, including trans-6-(4'-hydroxy-3'-methoxy-phenyl)-2,4-dioxo-5-hexenal (half-curcumin), vanillin, ferulic acid and feruloyl methane.

The poor aqueous stability of curcumin reduces an effective uptake in vivo and as such frequent doses may be required to main-tain a sufficient curcumin level for effective medicinal response [23]. Therefore, a number of research groups are focusing on en-capsulation of curcumin using delivery agents to improve the aque-ous solubility and stability, with the ultimate goal of enhancing bioavailability of curcumin in the treatment of diseases. Such deliv-ery agents require two key features to achieve a high solubility and stability of curcumin under physiological environment. First, en-capsulation of curcumin by delivery agents with either a hydropho-bic interior or moiety is essential to stabilize curcumin as well as segregate curcumin from water to prevent rapid hydrolysis. Second, a hydrophilic exterior is necessary to disperse the delivery agents in the aqueous environment. Recent work has demonstrated curcumin

O

HO

H3C

OH O

O

OH

CH3

Keto-enol tautomer

O

HO

H3C

O O

O

OH

CH3

Di-keto tautomer

O

HO

H3C

OH O

O

OH

CH3

O

HO

H3C

OH O

OH

HO

OH O

OHBisdemethoxycurcumin

Demethoxycurcumin

Curcumin


delivery using agents such as micelles, liposomes, polymer nanoparticles, plasma proteins and cyclodextrins.

CURCUMIN ENCAPSULATION BY MICELLES,

LIPOSOMES AND POLYMER NANOPARTICLES

Micelles and liposomes are often used as cell membrane mim-ics because of the similar microscopic environment produced by these self-assembled structures. Surfactant and lipid molecules consist of long alkyl tail(s) and a charged head group [24,25]. In the formation of micelles in the aqueous environment, the long alkyl tails of surfactants aggregate and form a hydrophobic core, while the polar head groups provide charges on the surface of the micellar particle to facilitate interaction with the surrounding water mole-cules. However, this aggregation can only occur above the critical micelle concentration [24]. Surfactant micelles are often used for solubilizing and stabilizing hydrophobic molecules. Curcumin is stable in anionic and neutral micelles including those of sodium dodecyl sulfate (SDS) and Triton-X 100 [26,27]. Cationic micelles such as cetyltrimethylammonium bromide (CTAB) and dodecyl-trimethylammonium bromide (DTAB) also show significant stabili-zation of curcumin, especially under alkaline condition as rapid hydrolysis is expected [11,18]. A measurable fluorescence quantum yield of approximately 4% indicates a strong interaction between curcumin and the hydrophobic domain of the micelles (as curcumin is non-fluorescent in water), resulting in segregation of curcumin from water molecules and hence inhibition of hydrolysis [18].

Similarly, lipid molecules aggregate to form vesicles with a lipid bilayer and they are also known as liposomes [28]. The core and the surrounding environment of the vesicle are composed of water with which the polar head groups interact, allowing the vesi-cles to remain suspended in an aqueous solution. Aggregation of the long alkyl tails of the lipid molecules provides a hydrophobic envi-ronment to accommodate hydrophobic species including drugs inside the bilayers [29-31]. Stabilization of curcumin using liposomes has been demonstrated in physiological environment [32,33]. In 1993, Tønnesen et al. first showed stabilization of cur-cumin using liposomes [32]. In a separate study, Barry et al. deter-mined the orientation of curcumin using 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) vesicles in order to infer the interaction between curcumin and the membrane [34]. At low con-centrations, curcumin binds to either side of DMPC bilayer through hydrogen bonding with the phosphate group of the lipids. At high concentrations of curcumin, however, linear dimers of curcumin that are held by hydrogen bonding between the phenol moieties bind across the lipid bilayer.

Synthetic polymers also form micellar aggregates to yield polymer nanoparticles, which are useful for curcumin delivery. Bisht et al. demonstrated the synthesis and characterization of 50-nm polymeric nanoparticles which are aggregates of cross-linked and random copolymers of N-isopropylacrylamide, with N-vinyl-2-pyrrolidone and poly(ethyleneglycol)monoacrylate [35]. The micel-lar nanoparticles solubilize curcumin in water at a concentration that curcumin alone would precipitate. In addition, Mohanty et al.achieved stabilization of curcumin using copolymeric micelles composed of methoxy(polyethyleneglycol) and poly- -caprolac-tone [36]. The curcumin-loaded micellar nanoparticles, which were present in 65% humidity for three months, were able to maintain a curcumin level of approximately 56% at the end of the study.

CURCUMIN BINDING WITH PROTEIN MICELLES AND PLASMA PROTEINS

In spite of the high stabilization effects on curcumin using mi-celles, liposomal vehicles and polymer nanoparticles, their potential incompatibility in vivo may become issues in clinical trials. While surfactants may have a low toxicity to humans, their degradation products may be harmful [37,38]. An alternative delivery method to

improve the bioavailability of curcumin is to use milk proteins and plasma proteins to resolve the potential toxicity issue.

Food proteins possess potential abilities not only to suppress curcumin degradation but also to exhibit biocompatibility. Casein is the major protein in mammalian milk and widely used in the food industry [39]. Cross-linking of caseins using glutaraldehyde enables formation of micellar microspheres [39,40]. Casein micelles exhibit a stabilization effect of curcumin with a strong binding constant on the order of 105

M-1

[39,41,42]. Cytotoxicity assays of curcumin in the presence and absence of casein micelles revealed that curcumin encapsulated in casein micelles is as effective as curcumin alone against cancer cells [39,41], indicating a potential application in clinical trials.

Plasma proteins play an important role as transporters in physiological functions [43,44]. Multiple binding sites of plasma proteins provide possible interaction with a variety of ligands in-cluding metal ions, fatty acids, amino acids and drugs [43,45]. Se-rum albumin is the most abundant plasma protein and its hydropho-bic pockets allow curcumin to form a complex in the aqueous envi-ronment [44]. Absorption and fluorescence quenching studies on the serum albumin-curcumin complex resulted in a binding constant of 10

4 10

5 M

-1 [44]. Furthermore, we have previously investi-

gated stabilization of curcumin using plasma proteins including human serum albumin (HSA), fibrinogen, transferrin and immuno-globulin G (IgG) under physiological conditions [46]. The spectro-scopic studies revealed that both HSA and fibrinogen suppress hy-drolysis of curcumin, whereas neither transferrin nor IgG stabilizes curcumin [46]. The stabilization effect of HSA and fibrinogen is enabled by the strong binding of curcumin to the hydrophobic do-mains of the proteins. The binding constants of curcumin to HSA and fibrinogen are (1.22 ± 0.35) 10

5 and (5.99 ± 1.75) 10

4 M

-1,

respectively [46]. In addition, denatured HSA at 50 °C shows a significantly weaker ability to stabilize curcumin than wild type, which is indicative of a weaker interaction between curcumin and the hydrophobic pockets [46]. In short, delivery of curcumin using plasma proteins, particularly HSA and fibrinogen, may be a practi-cal method in clinical trials. Curcumin can be delivered in vivo using the patient’s own plasma proteins through intravenous deliv-ery.

STABILIZATION OF CURCUMIN BY CYCLODEXTRINS AND DIAMIDE LINKED -CYCLODEXTRIN DIMERS

Oral dosage of curcumin may be achieved by using cyclodex-trins as delivery agents. Cyclodextrins are naturally occurring cyclic oligosaccharides, appearing as white crystalline powder at room temperature. The Food and Drug Administration (FDA) has ap-proved the clinical use of cyclodextrins due to their non-toxic prop-erties [47-49]. There are mainly three types of cyclodextrins; -, -and -cyclodextrins, consisting of six, seven and eight glu-copyranoside units, respectively, which are linked with -1,4 glyco-sidic bonds in a toroidal shape [50,51]. Cyclodextrins have a hy-drophilic exterior and a hydrophobic interior. The hydrophobic interior interacts with the hydrophobic moieties of the bound mole-cule, typically with the aromatic ring by van der Waals interactions [51,52]. In a drug carrier system, cyclodextrins act as hosts to cap-ture hydrophobic guest molecules [53,54]. The driving force for this self-assembly is the entropy increase due to exclusion of water molecules from the cyclodextrin cavity [51,55], which must over-come the corresponding decrease due to host-guest complexation to result in a successful binding event. A significant advantage of this delivery system is its high structural integrity. Micellar particles may lose their structural properties below critical micelle concen-trations, and plasma proteins may denature depending on pH and temperature. Cyclodextrins, however, retain their structural proper-ties as drug delivery systems under a wide range of physiological conditions, exhibiting more potential for drug delivery than mi-celles and plasma proteins.


The low toxicity of cyclodextrins and the structural characteris-tics lead to a possible delivery pathway for curcumin using cyclo-dextrin-curcumin complexation. However, neither - nor -cyclodextrin stabilizes curcumin even at 100:1 cyclodex-trin:curcumin molar ratio under physiological conditions, while -cyclodextrin at the same molar ratio shows some stabilization ef-fect, resulting in a short degradation half-life of curcumin of ap-proximately 4.46 ± 0.12 h [56]. The short half-lives of curcumin in the presence of -, - and -cyclodextrins imply a rather weak in-teraction between curcumin and cyclodextrins. Therefore, synthetic modifications of cyclodextrins are essential to improve the cyclo-dextrin-curcumin interaction and to increase the aqueous stability of curcumin. (2-hydroxypropyl)- -cyclodextrin, (2-hydroxypropyl)- -cyclodextrin and (2-hydroxypropyl)- -cyclodextrin are modified cyclodextrins with enhanced hydrophobicity. Suppression of cur-cumin degradation has been demonstrated in the presence of these modified cyclodextrins at 30 °C [57]. Baglole et al. and Singh et al.have demonstrated stronger interactions between the modified cy-clodextrins and curcumin, and the level of 1:1 host-guest complexa-tion is increased by 3 10-fold [58,59]. Another possible modifica-tion of cyclodextrin is to introduce a linker between two cyclodex-trins in order to induce cooperative interaction with a guest mole-cule. Pham et al. has established the synthesis of diamide linked cyclodextrin dimers for - and -cyclodextrins [60,61]. Our previ-ous study has demonstrated strong cooperative binding and effec-tive stabilization of curcumin using diamide linked -cyclodextrin dimers, N,N’-bis(6

A-deoxy- -cyclodextrin-6

A-yl)succinamide,

66 CD2su, and N,N’-bis(6A-deoxy- -cyclodextrin-6

A-yl)urea,

66 CD2ur (Fig. 4) [56]. Both 66 CD2su and 66 CD2ur stabilize curcumin at the 1:1 molar ratio and the half-life of curcumin is extended by at least 180 750 times, when compared to curcumin alone [56]. The binding constants of 66 CD2su-curcumin and 66 CD2ur-curcumin complexes are (8.7 ± 0.4) 10

6 and (2.0 ± 0.1)

106 M

-1, respectively [56]. Overall, the remarkable and effective

suppression of curcumin degradation using diamide linked -cyclodextrin dimers, 66 CD2su and 66 CD2ur, provide a potentially attractive method for oral dosage.

MEDICINAL ACTIVITIES OF CURCUMIN

Turmeric has been utilized not only as a dietary herbal medicine but also as a topical agent for wound healing enhancement for cen-turies [62]. Despite the limited bioavailability of curcumin as de-scribed before, the therapeutic activities of curcumin have been demonstrated for diseases including inflammation [63-70], cystic fibrosis [71-73], Alzheimer’s disease [74-77], and cancer [62,68,78-82] without significant side effects in the past decades. A recent clinical trial indicated that a large oral dose of curcumin shows insignificant toxicity [83].

Wound Healing and Anti-inflammatory Activities of Curcumin

Wound healing is a complex process of cell proliferation and migration involving different types of cells [84]. Inadequate treat-ment of the wound further causes inflammatory response at the trauma. The use of herbal medicine including turmeric as wound dressing has been documented in ancient Indian medical literature [62]. An in vitro study has revealed that curcumin is more effective in suppressing inflammation than demethoxycurcumin and bisde-methoxycurcumin [63]. In addition, recent in vivo studies have also demonstrated that curcumin possesses the ability to reduce inflam-mation and enhance wound healing [64-66]. The anti-inflammatory activity of curcumin has been related to the inhibition of the NF- Bactivation pathways [68]. Other proposed mechanisms are inhibi-tion of cyclooxygenase-II (COX-II) transcription and expression [63,68], activation of peroxisome proliferator-activated receptors (PPARs) and urokinase plasminogen mRNA [66], and scavenging of oxidative radicals [62,66,68].

Fig. (4). Structures of (a) -cyclodextrin, (b) 66 CD2su-curcumin and (c)

66 CD2ur-curcumin complexes. This figure is adapted with permission from

ref. [56]. Copyright 2011 ACS.

Anti-cystic Fibrosis Activities of Curcumin

Cystic fibrosis is a life threatening disease caused by mutations in the gene expressing the cystic fibrosis transmembrane conduc-tance regulator (CFTR). Misfolded CFTR proteins interact with calcium-dependant chaperones in the endoplasmic reticulum, result-ing in degradation by the proteasome in the cytoplasm before CFTR proteins activate the chloride ion channel across epithelial mem-branes [72]. Consequently, the elevated level of chloride ions in-duces influx of sodium ions [73], which causes dehydration of the tissue surface and production of viscous secretions typically in the lungs and nose. Egan et al. have demonstrated a potential use of curcumin to treat the cystic fibrosis defect [72]. The results of im-munoblot assay and surface density measurements are indicative of an enhanced level of CFTR proteins in the cytoplasm after doses of curcumin. It is possible that binding of curcumin corrects the CFTR protein expression and/or the tertiary structure of CFTR proteins


[72]. An alternative mechanism suggests that curcumin decreases the calcium level in the endoplasmic reticulum and inhibits cal-cium-dependent chaperones [73]. Hence, the reduced level of pro-teasomal degradation of CFTR proteins leads to efflux of chloride ions, even though the misfolded CFTR proteins can act as a chlo-ride channel [73]. Moreover, encapsulation of curcumin using poly(lactic-co-glycolic acid) (PLGA) nanoparticles enhances the activity of curcumin against the cystic fibrosis defect owing to a possible delivery of curcumin into the endoplasmic reticulum [71].

Alzheimer’s Disease and Activities of Curcumin

A proposed cause of Alzheimer’s disease is aggregation of amyloid -peptides and accumulation of -amyloid fibrils in the brain tissue [85]. The potential ability of curcumin to treat Alz-heimer’s disease has been studied both in vitro and in vivo [75-77]. Solutions of amyloid -peptides and -amyloid fibrils were pre-pared in the absence and presence of curcumin in order to study its effect on preventing aggregation of -amyloid fibrils. The results show that curcumin binds to the end of -amyloid fibril and/or amyloid -peptide, and hence it inhibits further aggregation of -amyloid fibrils [75,76]. Yanagisawa et al. have revealed molecular insight into the binding of curcumin to amyloid -peptides by dis-covering that the keto-enol and di-keto tautomerization of curcumin is essential to inhibit amyloid -peptide aggregation [86]. In an in vivo study, although the curcumin concentration in the mouse brain was as low as 1 M, curcumin was still able to either reduce aggre-gation or promote the destabilization of amyloid -peptides and -amyloid fibrils [76]. Furthermore, curcumin also reduces oxidative stress and inflammation of the brain due to Alzheimer’s disease [74]. Overall, curcumin exhibits a considerable promise to promote neuroprotective treatment for Alzheimer’s disease.

Anti-cancer Activities of Curcumin

Cancer (malignant neoplasm) is classified as a disease in which a group of cells exhibits uncontrolled growth, destruct adjacent tissues, and spread to other body parts by either the lymph nodes, blood stream, or both [87]. A statistical report has estimated that 10.9 million new cancer cases were found and 6.7 million deaths were recorded worldwide in 2002 [88]. The most common and lethal cancers are lung, stomach and liver cancers [88]. The anti-cancer activities of curcumin have been widely investigated using different types of cells and animal models [1,62,68,78-82]. The anti-proliferative effect of curcumin has been determined in previ-ous studies, using different delivery agents including PLGA nanoparticles [89], -cyclodextrins [90], glycerol manooleate based nanoparticles, copolymeric nanoparticles of Pluronic F-127 and polyvinyl alcohol [91], and O-carboxymethyl chitosan nanoparti-cles [92]. It is also known that curcumin induces cellular apoptosis [93-95]. Although the proposed mechanisms are complicated, there are two roles of curcumin in its anti-cancer activities. First, curcu-min activates and/or inhibits some of the regulators and receptors involving cellular proliferation, migration and transcription [89-92]. These indirect activities of curcumin seem to explain its efficacy against multiple types of cancerous cells. Second, curcumin inter-acts and directly damages deoxyribonucleic acid (DNA), which leads to apoptosis [93-95]. The ability of curcumin to cause DNA damage often involves transition metals including copper and iron.

THE ROLE OF COPPER IN CANCER AND OTHER DIS-

EASES

Transition metal ions including copper, iron and zinc form com-plexes with specific proteins and are well-regulated in the body. For instance, copper plays an essential role in the wound healing proc-ess. Jones et al. have determined a trace amount of copper in wound fluid [96]. Vascular endothelial growth factor, which is sensitive to the presence of copper, stimulates angiogenesis and extracellular matrix remodeling [97]. However, alteration in the ion-binding

protein conformation results in unregulated concentrations of the metals and potentially leads to development of diseases. A well known effect from accumulation of free ions is metal-induced oxi-dative stress. In particular, some diseases are related to the presence of high concentrations of copper which generates oxidative radicals through the Fenton reaction [98]. Free radicals have been estab-lished as mediators in many diseases [99-102]. Previous studies reported elevated concentrations of copper in tumors and cancer tissues [103-105]. The excess copper not only generates reactive oxygen radicals but also induces uncontrolled angiogenesis in can-cer tissues [104,105]. Moreover, Atwood et al. showed that cop-per(II) induces aggregation of amyloid -peptides in Alzheimer’s disease [106]. Studies have shown that a chelating agent limits the free copper concentration and leads to improvement in disease con-ditions [107-109].

BINDING OF CURCUMIN TO COPPER

The medicinal activities of curcumin such as anti- -amyloid fibril formation and anti-cancer effects have been related to its in-teraction with copper [110-112]. This relation arises from the ele-vated concentration of copper at the disease site and a high binding affinity of curcumin toward copper(II). The interaction between curcumin and copper(II) has been demonstrated with infrared (IR) absorption spectroscopy. Zhao et al. have shown that the presence of copper(II) shifts the carbonyl absorption band in the keto-enol moiety of curcumin to a lower vibrational frequency [113]. In addi-tion, the appearance of a new absorption band at 415 cm

-1 was ob-

served in the IR spectrum of curcumin [113]. The changes indicate a strong interaction between curcumin and copper(II). In particular, the appearance of the new band suggests direct interaction between the oxygen atoms of curcumin and copper(II) [113-115]. In the binding process, deprotonation of curcumin has been suggested to be important, as shown in Scheme 4 [111,116], which leads to the formation of a singly charged curcumin anion with a -diketonate moiety. It is well established that -diketonates form a strong, bi-dentate chelation with copper(II) [117]. Recently, the binding of curcumin with copper(II) has been under intense investigation to provide insight into the mode of action for the medicinal effects of curcumin.

Scheme 4. Proposed mechanism for formation of 1:1 copper(II)-curcumin

complex. This figure is adapted with permission from ref. [127]. Copyright

2012 CSIRO PUBLISHING.

O

HO

H3C

OH O

O

OH

CH3

O

HO

H3C

O O

O

OH

CH3

O

HO

H3C

O O

O

OH

CH3

+Cu2+

CuH2O OH2

-H


UV-Vis absorption and fluorescence emission spectroscopy are commonly used for studies of curcumin due to its desirable photo-chemical properties in various solvents, as mentioned earlier. The copper(II)-curcumin complexes exhibit a high molar absorptivity in the visible range [110-112]. The changes in the optical absorbance at 445 nm and 380 nm of curcumin in the presence of copper(II) indicate a strong interaction with copper(II) and the formation of a copper(II)-curcumin complex. In the work of Baum et al., the com-plex formed in a pH 7.4 buffer solution with 1% human serum and ascorbic acid was determined to be at a 1:2 copper(II):curcumin ratio with a binding constant for the first curcumin of about (1 1.7) 10

6 M

-1 and the second curcumin of about 7.7 10

5 M

-1 us-

ing the Scatchard plot analysis [112]. In addition to the UV-Vis absorption spectroscopy results, steady state fluorescence spectros-copy has been used to investigate the copper(II)-curcumin interac-tion in methanol and the SDS micellar solution [118]. These solu-tions were used for two reasons. First, the solubility and stability of curcumin are high in those media. Second, curcumin exhibits a relatively high fluorescence quantum yield in both solvent envi-ronments [10], hence, changes in the fluorescence spectrum signify the association of curcumin with copper(II) [10]. By using the effi-cient fluorescence quenching of curcumin by copper(II) in a con-centration dependent manner, as shown in Fig. 5 [118], the binding constants of the copper(II)-curcumin complexes were determined [118]. The binding constants for the formation of the 1:1 and 1:2 complexes are (1.33 ± 0.47) 10

8 M

-1 and (6.79 ± 1.77) 10

5 M

-1,

respectively, in methanol. The corresponding binding constants in the SDS micellar solution are (9.90 ± 1.68) 10

5 M

-1 and (1.70 ±

0.48) 106 M

-1, respectively. The results indicate that the 1:1 cop-

per(II)-curcumin complex formation is dominant in methanol while formation of both 1:1 and 1:2 copper(II)-curcumin complexes are almost equally favorable in the SDS micellar solution. This differ-ence in the binding affinity suggests that the interaction between curcumin and copper(II) is dependent on the solvent environment. Furthermore, minor changes in UV-Vis absorption and fluorescence emission spectra of curcumin in the presence of copper(I) indicate a weak interaction between curcumin and copper(I) [118]. This result suggests that the binding is also strongly dependent on the oxida-tion state of copper.

The 1:1 and 1:2 copper(II)-curcumin complexes have been synthesized by mixing a copper(II) salt with curcumin at either a 1:1 or 1:2 molar ratio [110,111,119]. A recent computational study has demonstrated that the 1:1 and 1:2 copper(II)-curcumin com-plexes have different geometries, which are shown in Fig. 6 [116]. Density functional theory shows that the lowest energy geometries of the 1:1 and 1:2 copper(II)-curcumin complexes are square planar around copper(II) with the 1:2 complex being the more stable of the two complexes [116]. In addition,

1H NMR spectroscopy, a combi-

nation of thermogravimetric analysis and differential thermal analy-sis (TG-DTA analysis) and mass spectrometry have indicated that water molecules interact with the copper(II) center in the 1:1 cop-per(II)-curcumin complex [113]. The square planar geometry of the complexes was further supported by electron paramagnetic reso-nance (EPR) spectroscopy results, which indicate a square planar orientation of oxygen atoms around the copper(II) center in both the 1:1 and 1:2 copper(II)-curcumin complexes [110,111].

As described earlier, an elevated level of copper(II) is found at the site of amyloid aggregates and tumors [103-106]. Therefore, formation of the copper(II)-curcumin complexes is highly likely at the disease site as curcumin exhibits a high binding affinity towards copper(II). The interaction between curcumin and copper(II) may be important in the medicinal effects of curcumin. A recent study has demonstrated that curcumin is capable of acting as an iono-phore to induce apoptosis in ovarian cancer cells [120]. The fluo-rescence images of ovarian cancer cells stained with a copper indi-cator, Phen Green™ FL probe, were recorded with a confocal

Fig. (5). Determination for binding constants by fluorescence quenching of

curcumin with copper(II). This figure is adapted with permission from ref.

[118]. Reproduced by permission of the PCCP Owner Societies.

Fig. (6). Calculated structures of the (a) 1:1 and (b) 1:2 copper(II)-curcumin

complexes. This figure is adapted with permission from ref. [116]. Copy-

right 2010 John Wiley and Sons.

microscope, as shown in Fig. 7. It is clear that the probe is present in the cytoplasm of the ovarian cancer cells without any addition of either copper(II) or curcumin. First, there was no change in the fluorescence intensity of cells due to the presence of curcumin. Similarly, the fluorescence intensity in cells exhibits a negligible change in the presence of copper(II). However, the fluorescence intensity decreases significantly in cells after treatment with both curcumin and copper(II) simultaneously. It is well established that binding of copper quenches the fluorescence of Phen Green™ FL, which indicates in this case that copper(II) is taken up by ovarian cancer cells in the presence of curcumin [120]. This significant increase in the intracellular copper(II) level suggests that curcumin facilitates the transport of copper(II) into cancer cells. This result further supports that the interaction between curcumin and cop-per(II) is important in the medicinal effects of curcumin.

400 500 600 700

Flu

ore

sc

en

ce

Inte

ns

ity

(a

.u.)

Wavelength (nm)

a

b

100

300

500

700

100

200

300

a b


ANTI-OXIDANT EFFECTS OF CURCUMIN IN THE PRES-

ENCE OF COPPER

The ability of curcumin to deactivate and neutralize reactive free radicals is well known [62,68]. Similar to curcumin, the cop-per(II)-curcumin complexes also exhibit anti-oxidant effects. Koiram et al. demonstrated that both curcumin and the 1:1 cop-per(II)-curcumin complex inhibit lipid peroxidation and reduce the radiation damage of cell membrane in Swiss Albino mouse liver homogenates using a thiobarbiturate acid assay [121]. Additionally, the 1:1 copper(II)-curcumin complex exhibits more effective inhibi-tion of radiation-induced lipid peroxidation than curcumin alone [121], implying that the anti-oxidant effects of curcumin are en-hanced in the presence of copper(II). Moreover, the reactions of curcumin and its copper(II) complexes with reactive oxygen species were investigated [121]. The rate constants for the reaction with singlet oxygen were determined to be (1.97 ± 0.30) 10

5 M

-1s

-1 and

4.6 102

M-1

s-1

for the 1:1 copper(II)-curcumin complex and cur-cumin, respectively. The significantly faster rate for radical scav-enging activity of the 1:1 complex than that of curcumin indicates that the complex is a more superior radical scavenger than curcu-min. The ability of the 1:1 copper(II)-curcumin complex to scav-enge superoxide radicals was also demonstrated using EPR spec-troscopy [110,111]. Superoxide is stabilized by complexation with crown ether in DMSO and its presence is detected using EPR spec-trometer. The results show that the level of superoxide is signifi-cantly reduced by the 1:1 and 1:2 copper(II)-curcumin complexes, indicating the ability of these complexes to scavenge superoxide [111]. Furthermore, the superoxide radical scavenging reaction by the 1:1 and 1:2 complexes were monitored with UV-Vis absorption spectroscopy, showing changes in the spectrum of the copper(II)-curcumin complexes over time after mixing with superoxide [110,111]. The rate constants for the reaction of the superoxide radical with the 1:1 and 1:2 copper(II)-curcumin complexes were determined to be (7.1 ± 0.1) 10

5 M

-1s

-1 and (1.04 ± 0.17) 10

5

M-1

s-1

, respectively. The faster rate of reaction of the 1:1 copper(II)-curcumin complex indicates that it has a more superior anti-oxidant ability than the 1:2 complex. Additionally, the IC50 values for the inhibition of cytochrome c (Fe

3+) by the 1:1 and 1:2 copper(II)-

curcumin complexes were determined to be 6.7 M and 68 M, respectively [110].

In addition to the enhanced anti-oxidant effects of the 1:1 cop-per(II)-curcumin complex, studies have shown the ability of the complex to maintain anti-oxidant enzyme levels inside mouse spleen cells [122]. It has been shown that there is an increase in the activity of anti-oxidant enzymes such as glutathione peroxidase and catalase in splenic lymphocytes after treatment with the 1:1 cop-

per(II)-curcumin complex. Furthermore, while radiation treatment suppresses the activities of these anti-oxidant enzymes, application of the complex after radiation reverses the radiation damage [122]. Specifically, there is evidence for the involvement of the cop-per(II)-curcumin complex in the protein kinase C (PKC ) and NF- B signaling pathway to result in inhibition of radiation-induced protein oxidation and improve cell viability. Similar pro-tective effects by the complex were observed in the liver of Swiss Albino mice after -radiation treatment [121]. Results have shown that the copper(II)-curcumin complex is capable of maintaining the levels of anti-oxidant enzymes including glutathione S-transferase, catalase and superoxide dismutase. The ability to maintain the lev-els of anti-oxidant enzymes as well as enhance radical scavenging activity may be important mechanisms for the anti-oxidant effects of curcumin in the presence of copper(II).

DNA DAMAGE IN THE PRESENCE OF COPPER – CUR-CUMIN AS A PRO-OXIDANT

In contrast to the anti-oxidant effects, curcumin and its cop-per(II) complexes have also shown pro-oxidant effects. Studies have demonstrated that curcumin has dual roles in its interaction with DNA. While curcumin and its copper(II) complexes exhibit anti-oxidant effects in cells and protect DNA from damages by radicals, curcumin also shows pro-oxidant effects and mediates DNA damage in the presence of copper(II). The interaction be-tween double stranded DNA (dsDNA) and curcumin with cop-per(II) has been investigated with differential pulse voltammetry, as shown in Fig. 8 [123]. First, curcumin exhibits an intense peak at 0.3 V and 0.6 V in the presence of dsDNA. At a 1:1 curcumin to copper(II) molar ratio, there is a decrease in the 0.3-V peak and this peak decreases significantly for 1:2 and 1:4 curcumin to copper(II) molar ratios. In addition, an increase was recorded for the peak at 0.6 V at the 1:1 ratio. At a higher ratio of copper(II) to curcumin, the 0.6-V peak increases as well as shifts to a higher potential. Fur-thermore, the peak current near 1 V is a characteristic oxidation peak of the guanine in the dsDNA and its decrease suggests DNA damage by the copper(II)-curcumin complex. These results suggest a strong interaction between the copper(II)-curcumin complex and the dsDNA. The presence of curcumin and copper(II) may be im-portant in the pro-oxidant effects.

Ahsan et al. have reported the damage of calf thymus and su-percoiled plasmid pBR322 DNA by curcumin in the presence of copper(II) [124]. Curcumin causes scission of the double stranded calf thymus DNA in the presence of copper(II) and this process is dependent on the concentrations of curcumin and copper(II). Cur-cumin also converts supercoiled plasmid pBR322 DNA into open circular DNA in the presence of copper(II) and iron(III), which is

Fig. (7). Fluorescence and confocal images of ovarian cancer cells demonstrate curcumin-mediated transport of copper(II). This figure is reprinted with per-

mission from ref. [120]. Copyright 2010 IIAR.


indicative of DNA damage. Similarly, damage of calf thymus DNA in the presence of copper(II) was observed after curcumin was treated with human cytochrome P450 isozyme [125]. While the detailed mechanism of curcumin-induced DNA damage in the pres-ence of copper(II) remains unclear, there is increasing evidence to suggest that the redox reaction of Cu(II)/Cu(I) couple may play an important role.

REDOX REACTION OF Cu(II)/Cu(I) COUPLE

Recently, a mechanism for the copper-mediated DNA damage of curcumin has been proposed (Scheme 5). The reduction of cop-per(II) to copper(I) has been suggested to be essential in the pro-oxidant effects of curcumin [125]. Copper(II) is essential to the curcumin-induced DNA damage and curcumin was found to be most effective in DNA damage among all curcuminoids [126]. It was suggested that the relative DNA cleavage activity is related to the efficiency of the curcuminoids to reduce copper(II) to copper(I). As mentioned earlier, curcumin converts supercoiled DNA to open circular in the presence of copper(II). However, this conversion is suppressed by the presence of a copper(I) chelating agent, such as neocuproine (Fig. 9) [124]. The suppression on the DNA damage is dependent on the concentration of neocuproine while a nearly com-plete inhibition was observed at 0.4 mM (Fig. 9, lane e to g). This result indicates that the presence of free copper(I), as a consequence of reduction of copper(II), is essential to the DNA cleavage ability of curcumin. As discussed earlier, curcumin forms stable 1:1 and 1:2 complexes with copper(II) in most solvents. However, our re-cent study has demonstrated the breakdown of curcumin in the presence of copper(II) under a reducing environment [127]. It was suggested that either the degradation products or the radicals gener-ated in the Cu(II)/Cu(I) redox process by the copper(II)-curcumin complex may be responsible for the medicinal effects of curcumin.

The reduction potential of the Cu(II)/Cu(I) couple in the cop-per-curcumin complex has been determined previously to be about 0.4 V, which is similar to other molecules that exhibit superoxide scavenging activity [111]. Furthermore, it is possible for an electron transfer reaction to occur between curcumin and copper(II) in the complex under a reducing environment [128,129], as shown in Scheme 6 [128,129]. As a result of electron transfer, curcumin radi-cals are formed and copper(II) is reduced to copper(I), which is then followed by a rapid decomposition [127]. Recent results from UV-Vis absorption spectroscopy, mass spectrometry and high per-formance liquid chromatography clearly show the generation of

Fig. (9). Electrophoresis gel demonstrating the effect of neocuproine on the

curcumin-induced DNA damage in the presence of copper(II) ion. (a)

pBR322 DNA, (b) as in (a) with addition of copper(II), (c) as in (b) with

addition of curcumin, (d) as in (b) with addition of 0.6 mM of neocuproine

and (e) (g) as in (c) with increasing concentration of neocuproine at 0.1,

0.2 and 0.4 mM, respectively. OC and SC stand for open circular and super-

coiled DNA, respectively. This figure is reprinted with permission from ref.

[124]. Copyright 1998 Elsevier.

metabolites of curcumin from copper-induced degradation in aceto-nitrile and in SDS micelles in the presence of ascorbic acid [127]. However, curcumin remains stable in the presence of copper(II) in methanol and SDS micellar solution, indicating that the degradation process is dependent on the redox environment. Additionally, the UV-Vis absorption and fluorescence emission spectra of curcumin in the presence of copper(I) are nearly identical to those of curcu-min, which implies a weak interaction between the two species. Therefore, the copper-induced degradation of curcumin is depend-ent on the redox environment as well as the oxidation state of cop-per.

GENERATION OF REACTIVE OXYGEN SPECIES BY COPPER(II) TO COPPER(I) REDUCTION

The ability to cause DNA damage by curcumin in the presence of copper(II) and cytochrome P450 isozymes has been demon-strated previously [125]. Results from mass spectrometry reveal the presence of O-demethyl curcumin, which suggests that curcumin is

Fig. (8). Differential pulse voltammogram of (a) dsDNA with curcumin, (b) (d) as in (a) with addition of 5, 10 and 20 M of copper(II). This figure is re-

printed with permission from ref. [123]. Copyright 2010 Elsevier.


Scheme 6. Proposed mechanism for the generation of the curcumin radical

by copper(II)/copper(I) redox reaction. This figure is adapted with permis-

sion from ref. [127]. Copyright 2012 CSIRO PUBLISHING.

first converted to this species by cytochrome P450 isozyme in the presence of nicotinamide adenine dinucleotide phosphotic acid (NADPH) [125]. Sakano et al. have demonstrated that O-demethyl curcumin undergoes a redox reaction in the presence of copper(II) and oxygen to result in the production of O-demethyl curcumin radical, singlet oxygen and copper(I), as illustrated in Scheme 5[125]. The resulting O-demethyl curcumin radical is then further oxidized to generate the o-quinone form of curcumin, singlet oxy-

gen and hydrogen peroxide. The generated reactive oxygen species through this proposed reaction cause oxidative damage to DNA [125]. In addition, Nair et al. have reported an increase in hepatic DNA adducts levels by curcumin and copper(II), which indicates DNA damage is resulted from oxidative stress [130]. Sahu et al.have also reported enhancement of the quercetin-induced DNA damage in the presence of curcumin and copper(II) [131]. Although the mechanism of mutagenicity of quercetin is not well understood, the potency of its mutagenicity under an aerobic environment sug-gests a possible involvement of reactive oxygen species. In addition to the copper-mediated DNA damage, stimulation of lipid peroxida-tion by curcumin was observed in the presence of the quercetin, which also indicates the involvement of reactive oxygen species [131]. As inhibition of DNA damage was observed in the presence of specific quenchers for radicals, the responsible reactive oxygen species for the damage can be identified, as shown in Fig. 10. Al-though a low level of inhibition of curcumin-mediated DNA dam-age was observed due to hydroxyl radical scavengers such as man-nitol, sodium formate and superoxide dismutase (SOD), the results clearly show inhibition of DNA damage by curcumin in the pres-ence of methional, which is also capable to scavenge hydroxyl radi-cals. In addition to methional, catalase, a scavenger for hydrogen peroxide, also exhibits ability to suppress DNA damage induced by curcumin, as shown in Fig. 10 [125]. Inhibition of curcumin-mediated DNA damage in the presence of quencher for active oxy-gen species such as potassium iodide and sodium azide was also reported, which suggests the involvement of hydroxyl radicals and singlet oxygen [124]. Therefore, species which are responsible for the curcumin-mediated DNA damage in the presence of copper(II) were identified to be hydrogen peroxide, hydroxyl radicals and singlet oxygen [124,125,131]. Furthermore, the inhibition of cur-

Scheme 5. Proposed mechanism for DNA damage by curcumin in the presence of copper(II). This figure is reprinted with permission from ref. [125]. Copy-

right 2002 Elsevier.

O

HO

H3C

O O

O

OH

CH3

Cu

O

HO

H3C

O O

O

OH

CH3

Cu

CH3CN NCCH3

CH3CN NCCH3

II

I


cumin-mediated DNA damage with copper(II) in the presence of bathocuproine, a Cu(I) specific quenching agent, indicates the in-volvement of copper(I), which is resulted in the reduction of cop-per(II) to copper(I) (Fig. 10) [125]. These results further support the generation of reactive oxygen species due to oxidation of curcumin in the presence of oxygen through the reduction of copper(II) to copper(I) to give rise to DNA damage.

Fig. (10). Electrophoresis gel demonstrating the effect of different types of

radical scavengers on the curcumin-induced DNA damage. This figure is

reprinted with permission from ref. [125]. Copyright 2002 Elsevier.

The generation of hydroxyl radicals was assayed with salicylate in the presence of curcumin and copper(II) [124]. In this assay, a hydroxylated product of salicylate was detected, strongly indicating the presence of hydroxyl radicals due to curcumin and copper(II) [124,126]. Moreover, DNA damage is dependent on the concentra-tion of curcumin, although this copper(II)-mediated DNA damage by curcumin was inhibited at a curcumin to copper(II) molar ratio above 2.5 [124,125]. It was suggested that a high concentration of curcumin decreases the concentration of free copper(II) as the -diketone moiety of curcumin chelates with copper(II) strongly. In short, the ability of curcumin to scavenge radicals and the redox reaction between copper(II) and curcumin catalyze the generation of reactive oxygen species, giving rise to DNA damage. Therefore, it has been proposed that curcumin has dual roles in the carcino-genesis of cells as an anti-oxidant and pro-oxidant, depending on the redox environment and concentration of free copper.

Yoshino et al. have shown that hydroxyl radicals generated from hydrogen peroxide leads to DNA cleavage and formation of 8-hydroxy-2’-deoxyguanosine DNA adduct [132]. However, there was no DNA adduct formation in other o-methoxy phenols without the -diketone moiety. Therefore, it was proposed that the -diketone moiety is essential in the pro-oxidant effect of curcumin. As discussed earlier, the formation of the -diketonate moiety is essential in the binding with copper(II) (Scheme 4) [117]. Negligi-

ble DNA damage was observed with capsaicin and gingerol, which have the phenolic hydroxyl group but not the -diketone moiety [132]. However, this observation is contradictory to the proposed mechanism in Scheme 5, which suggests that the phenolic hydroxyl functional group is essential for the generation of reactive oxygen species. Therefore, the formation of the copper(II)-curcumin com-plex and hence the reduction of copper(II) to copper(I) should be included in the mechanism of curcumin-induced DNA damage.

CURCUMIN-INDUCED APOPTOSIS IN THE PRESENCE

OF COPPER

As discussed earlier, the anti-cancer properties of curcumin may arise from the ability of curcumin to cause DNA damage in the presence of elevated copper(II) concentrations. This phenomenon suggests that curcumin induces apoptosis in cells, in particular can-cer cells [120,130,132,133]. Recently, enhancement of cell apopto-sis by curcumin was observed in the presence of copper(II) in ovar-ian and breast cancer cells [120]. However, no generation of reac-tive oxygen species in cells was observed, which is contradictory to the previous studies. This result implies the possibility of other mechanisms for curcumin-induced apoptotic cell death. Lou et al.suggested that curcumin acts as a copper(II) ionophore and it in-creases the cellular concentration of free copper(II) (Fig. 7), result-ing in accumulation of I B protein, an inhibitor of NF- B trans-ferase, which results in inhibition of the NF- B signaling pathway [120]. Additionally, they also reported inhibition of the mammalian target of rapamycin (mTOR) signaling pathway by curcumin in the presence of copper(II) due to an enhancement in the expression of both total and phosphor-p70-S6 kinase. This result suggests that the mechanism of curcumin-induced cell apoptosis in the presence of copper(II) occurs through the inhibition of both NF- B and mTOR pathways.

CONCLUSION

Curcumin, a yellow dye molecule found in the powder of Cur-cuma longa rhizome (turmeric), has shown enhancement in wound healing and multiple medicinal properties against inflammation, cystic fibrosis, Alzheimer’s disease and cancer. In order to utilize curcumin as a therapeutic agent, its limited aqueous solubility and stability are improved using a number of delivery systems including micelles, liposomes, polymer nanoparticles, casein micelles, plasma proteins and cyclodextrins. The interaction of curcumin with transi-tion metals has been shown to be important in its medicinal effects. On one hand, curcumin and its copper(II) complexes exhibit anti-oxidant effects. It has been demonstrated that they have abilities to scavenge radicals as well as maintain the levels of anti-oxidant enzymes. On the other hand, curcumin exhibits pro-oxidant effects in the presence of copper(II), causing DNA damage. Under certain disease conditions where a free copper level is elevated, the pro-oxidant effects of curcumin may dominate over its anti-oxidant effects. Curcumin forms stable complexes with copper(II) and this strong interaction between curcumin and copper(II) is essential in its DNA damage activity. Under a reducing environment, such as in the presence of ascorbic acid, the redox reaction between curcumin and copper generates reactive oxygen species, which mediate DNA damage. Additionally, curcumin can also alter the signaling path-ways which are crucial to cell survival in the presence of copper. The copper-mediated curcumin-induced apoptosis has been linked to the generation of reactive oxygen species, which leads to DNA damage as well as disturbance in the vital signaling pathway in cells. Therefore, curcumin has dual roles in the carcinogenesis with copper as an anti-oxidant and a pro-oxidant.

CONFLICT OF INTEREST

The authors confirm that this article content has no conflicts of interest.


ACKNOWLEDGEMENTS

The authors acknowledge financial support from the Australian Research Council, National Health and Medical Research Council Network “Fluorescence Applications in Biotechnology and Life Sciences” (FABLS), the Faculty of Sciences and School of Chemis-try and Physics at the University of Adelaide.

REFERENCES

[1] Sa G, Das T. Anti-cancer effects of curcumin: cycle of life and

death. Cell Division 2008; 3(1): 1-14. [2] Ammon HPT, Wahl MA. Pharmacology of curcuma-longa. Planta

Med 1991; 57(1): 1-7. [3] Mishra S, Palanivelu K. The effect of curcumin (turmeric) on

Alzheimer's disease: an overview. Ann Indian Acad Neur 2008; 11(1): 13-9.

[4] Roughley PJ, Whiting DA. Experiments in biosynthesis of curcumin. J Chem Soc Perkin Trans 1 1973(20): 2379-88.

[5] Vogel E, Pellertier S. J Pharm 1815; 2: 50-4. [6] Kiuchi F, Goto Y, Sugimoto N, Akao N, Kondo K, Tsuda Y.

Nematocidal activity of turmeric - synergistic action of curcuminoids. Chem Pharm Bull 1993; 41(9): 1640-3.

[7] Kita T, Imai S, Sawada H, Kumagai H, Seto H. The biosynthetic pathway of curcuminoid in turmeric (Curcuma longa) as revealed

by 13C-labeled precursors. Biosci Biotechnol Biochem 2008; 72(7): 1789-98.

[8] Katsuyama Y, Kita T, Horinouchi S. Identification and characterization of multiple curcumin synthases from the herb

Curcuma longa. FEBS Lett 2009; 583(17): 2799-803. [9] Payton F, Sandusky P, Alworth WL. NMR study of the solution

structure of curcumin. J Nat Prod 2007; 70(2): 143-6. [10] Chignell CF, Bilskj P, Reszka KJ, Motten AG, Sik RH, Dahl TA.

Spectral and photochemical properties of curcumin. Photochem Photobiol 1994; 59(3): 295-302.

[11] Wang ZF, Leung MHM, Kee TW, English DS. The role of charge in the surfactant-assisted stabilization of the natural product

curcumin. Langmuir 2010; 26(8): 5520-6. [12] Adhikary R, Carlson PJ, Kee TW, Petrich JW. Excited-state

intramolecular hydrogen atom transfer of curcumin in surfactant micelles. J Phys Chem B 2010; 114(8): 2997-3004.

[13] Adhikary R, Mukherjee P, Kee TW, Petrich JW. Excited-state intramolecular hydrogen atom transfer and solvation dynamics of

the medicinal pigment curcumin. J Phys Chem B 2009; 113(15): 5255-61.

[14] Nardo L, Andreoni A, Masson M, Haukvik T, Tønnesen HH. Studies on curcumin and curcuminoids. XXXIX. Photophysical

properties of bisdemethoxycurcumin. J Fluorescence 2011; 21(2): 627-35.

[15] Priyadarsini KI. Photophysics, photochemistry and photobiology of curcumin: Studies from organic solutions, bio-mimetics and living

cells. J Photochem Photobiol C 2009; 10(2): 81-95. [16] Bernabe-Pineda M, Ramirez-Silva MT, Romero-Romo M,

Gonzadlez-Vergara E, Rojas-Hernandez A. Determination of acidity constants of curcumin in aqueous solution and apparent rate

constant of its decomposition. Spectrochim Acta A 2004; 60(5): 1091-7.

[17] Shen L, Ji H-F. Theoretical study on physicochemical properties of curcumin. Spectrochim Acta A 2007; 67(3-4): 619-23.

[18] Leung MHM, Colangelo H, Kee TW. Encapsulation of curcumin in cationic micelles suppresses alkaline hydrolysis. Langmuir 2008;

24(11): 5672-5. [19] Kaminaga Y, Nagatsu A, Akiyama T, et al. Production of unnatural

glucosides of curcumin with drastically enhanced water solubility by cell suspension cultures of Catharanthus roseus. FEBS Lett

2003; 555(2): 311-6. [20] Letchford K, Liggins R, Burt H. Solubilization of hydrophobic

drugs by methoxy poly(ethylene glycol)-block-polycaprolactone diblock copolymer micelles: theoretical and experimental data and

correlations. J Pharm Sci 2008; 97(3): 1179-90. [21] Anand P, Kunnumakkara AB, Newman RA, Aggarwal BB.

Bioavailability of curcumin: problems and promises. Mol Pharm 2007; 4(6): 807-18.

[22] Tønnesen HH, Karlsen J. Studies on curcumin and curcuminoids .6. kinetics of curcumin degradation in aqueous-solution. Z

Lebensm Unters Forsch 1985; 180(5): 402-4.

[23] Wang YJ, Pan MH, Cheng AL, et al. Stability of curcumin in

buffer solutions and characterization of its degradation products. J Pharm Biomed Anal 1997; 15(12): 1867-76.

[24] Wennerstrom H, Lindman B. Micelles - physical-chemistry of surfactant association. Phys Rep 1979; 52(1): 1-86.

[25] Nagarajan R, Ruckenstein E. Theory of surfactant self-assembly - a predictive molecular thermodynamic approach. Langmuir 1991;

7(12): 2934-69. [26] Iwunze MO. Binding and distribution characteristics of curcumin

solubilized in CTAB micelle. J Mol Liq 2004; 111(1-3): 161-5. [27] Tønnesen HH. Solubility, chemical and photochemical stability of

curcumin in surfactant solutions - studies of curcumin and curcuminolds, XXVIII. Pharmazie 2002; 57(12): 820-4.

[28] Bangham AD. Physical structure and behavior of lipids and lipid enzymes. Adv Lipid Res 1963; 1: 65-104.

[29] Alving CR. Liposomes as carriers of antigens and adjuvants. J Immunol Methods 1991; 140(1): 1-13.

[30] Lian T, Ho RJY. Trends and developments in liposome drug delivery systems. J Pharm Sci 2001; 90(6): 667-80.

[31] Huwyler J, Wu DF, Pardridge WM. Brain drug delivery of small molecules using immunoliposomes. Proc Natl Acad Sci USA 1996;

93(24): 14164-9. [32] Tønnesen HH, Smistad G, Agren T, Karlsen J. Studies on curcumin

and curcuminoids .23. effects of curcumin on liposomal lipid-peroxidation. Int J Pharm 1993; 90(3): 221-8.

[33] Chen C, Johnston TD, Jeon H, et al. An in vitro study of liposomal curcumin: Stability, toxicity and biological activity in human

lymphocytes and Epstein-Barr virus-transformed human B-cells. Int J Pharm 2009; 366(1-2): 133-9.

[34] Barry J, Fritz M, Brender JR, Smith PES, Lee DK, Ramamoorthy A. Determining the effects of lipophilic drugs on membrane

structure by solid-state NMR spectroscopy: the case of the antioxidant curcumin. J Am Chem Soc 2009; 131(12): 4490-8.

[35] Bisht S, Feldmann G, Soni S, et al. Polymeric nanoparticle-encapsulated curcumin ("nanocurcumin"): a novel strategy for

human cancer therapy. J Nanobiotechnology 2007; 5(3): 18. [36] Mohanty C, Acharya S, Mohanty AK, Dilnawaz F, Sahoo SK.

Curcumin-encapsulated MePEG/PCL diblock copolymeric micelles: a novel controlled delivery vehicle for cancer therapy.

Nanomedicine 2010; 5(3): 433-49. [37] Scott MJ, Jones MN. The biodegradation of surfactants in the

environment. BBA Biomemb 2000; 1508(1–2): 235-51. [38] Yam J, Booman KA, Broddle W, et al. Surfactants: a survey of

short-term genotoxicity testing. Food Chem Toxicol 1984; 22(9): 761-9.

[39] Sahu A, Kasoju N, Bora U. Fluorescence study of the curcumin casein micelle complexation and its application as a drug

nanocarrier to cancer cells. Biomacromolecules 2008; 9(10): 2905-12.

[40] Huppertz T, Fox PF, Kelly AL. Properties of casein micelles in high pressure-treated bovine milk. Food Chem 2004; 87(1): 103-

10. [41] Esmaili M, Ghaffari SM, Moosavi-Movahedi Z, et al. Beta casein-

micelle as a nano vehicle for solubility enhancement of curcumin; food industry application. LWT-Food Sci Technol 2011; 44(10):

2166-72. [42] Yazdi SR, Corredig M. Heating of milk alters the binding of

curcumin to casein micelles, a fluorescence spectroscopy study. Food Chem 2012; 132(3): 1143-9.

[43] Kragh-Hansen U. Molecular aspects of ligand binding to serum albumin. Pharmacol Rev 1981; 33(1): 17-53.

[44] Barik A, Priyadarsini KI, Mohan H. Photophysical studies on binding of curcumin to bovine serum albumin. Photochem

Photobiol 2003; 77(6): 597-603. [45] Naik PN, Chimatadar SA, Nandibewoor ST. Study on the

interaction between antibacterial drug and bovine serum albumin: a spectroscopic approach. Spectrochim Acta A 2009; 73(5): 841-5.

[46] Leung MHM, Kee TW. Effective stabilization of curcumin by association to plasma proteins: human serum albumin and

fibrinogen. Langmuir 2009; 25(10): 5773-7. [47] FDA. alpha cyclodexrin. Food and Drug Administration; 2004

[updated 2012 Jun 30; cited 2012 Aug 3]; Available from: http: //www.accessdata.fda.gov/scripts/fcn/fcnDetailNavigation.cfm?rpt

=grasListing&id=155. [48] FDA. beta cyclodexrin. Food and Drug Administration; 2001

[updated 2012 Jun 30; cited 2012 Aug 3]; Available from: http:


//www.accessdata.fda.gov/scripts/fcn/fcnDetailNavigation.cfm?rpt

=grasListing&id=74. [49] FDA. gamma cyclodexrin. Food and Drug Administration; 2000

[updated 2012 Jun 30; cited 2012 Aug 3]; Available from: http: //www.accessdata.fda.gov/scripts/fcn/fcnDetailNavigation.cfm?rpt

=grasListing&id=46. [50] Szejtli J. Introduction and general overview of cyclodextrin

chemistry. Chem Rev 1998; 98(5): 1743-53. [51] Lincoln SF, Pham D-T. Cyclodextrins: From nature to

nanotechnology. Supramolecular Chemistry: From molecules to nanomaterials. John Wiley & Sons, Ltd; 2012.

[52] Hoshino M, Imamura M, Ikehara K, Hama Y. Fluorescence enhancement of benzene-derivatives by forming inclusion

complexes with beta-cyclodextrin in aqueous-solutions. J Phys Chem 1981; 85(13): 1820-3.

[53] Loftsson T, Duchene D. Cyclodextrins and their pharmaceutical applications. Int J Pharm 2007; 329(1-2): 1-11.

[54] Loftsson T, Konradsdottir F, Masson M. Influence of aqueous diffusion layer on passive drug diffusion from aqueous

cyclodextrin solutions through biological membranes. Pharmazie 2006; 61(2): 83-9.

[55] Rekharsky MV, Inoue Y. Complexation thermodynamics of cyclodextrins. Chem Rev 1998; 98(5): 1875-917.

[56] Harada T, Pham D-T, Leung MHM, et al. Cooperative binding and stabilization of the medicinal pigment curcumin by diamide linked

-cyclodextrin dimers: a spectroscopic characterization. J Phys Chem B 2011; 115(5): 1268-74.

[57] Tønnesen HH, Másson M, Loftsson T. Studies of curcumin and curcuminoids. XXVII. Cyclodextrin complexation: solubility,

chemical and photochemical stability. Int J Pharm 2002; 244(1–2): 127-35.

[58] Baglole KN, Boland PG, Wagner BD. Fluorescence enhancement of curcumin upon inclusion into parent and modified cyclodextrins.

J Photoch Photobio A 2005; 173(3): 230-7. [59] Singh R, Tønnesen HH, Vogensen SB, Loftsson T, Masson M.

Studies of curcumin and curcuminoids. XXXVI. The stoichiometry and complexation constants of cyclodextrin complexes as

determined by the phase-solubility method and UV-Vis titration. J Incl Phenom Macro 2010; 66(3-4): 335-48.

[60] Pham DT, Clements P, Easton CJ, Papageorgiou J, Maya BL, Lincoln SF. Complexation of 6-(4 '-(toluidinyl)naphthalene-2-

sulfonate by beta-cyclodextrin and linked beta-cyclodextrin dimers. New J Chem 2008; 32(4): 712-8.

[61] Pham DT, Ngo HT, Lincoln SF, May BL, Easton CJ. Synthesis of C6(A)-to-C6(A) and C3(A)-to-C3(A) diamide linked gamma-

cyclodextrin dimers. Tetrahedron 2010; 66(15): 2895-8. [62] Hatcher H, Planalp R, Cho J, Tortia FM, Torti SV. Curcumin: from

ancient medicine to current clinical trials. Cell Mol Life Sci 2008; 65(11): 1631-52.

[63] Lantz RC, Chen GJ, Solyom AM, Jolad SD, Timmermann BN. The effect of turmeric extracts on inflammatory mediator production.

Phytomedicine 2005; 12(6–7): 445-52. [64] Gopinath D, Ahmed MR, Gomathi K, Chitra K, Sehgal PK,

Jayakumar R. Dermal wound healing processes with curcumin incorporated collagen films. Biomaterials 2004; 25(10): 1911-7.

[65] Jagetia GC, Rajanikant GK. Curcumin treatment enhances the repair and regeneration of wounds in mice exposed to hemibody

gamma-irradiation. Plast Reconstr Surg 2005; 115(2): 515-28. [66] Jagetia GC, Rajanikant GK. Acceleration of wound repair by

curcumin in the excision wound of mice exposed to different doses of fractionated gamma radiation. Int Wound J 2012; 9(1): 76-92.

[67] Legeza V, Galenko-Yaroshevskii V, Zinov’ev E, et al. Effects of new wound dressings on healing of thermal burns of the skin in

acute radiation disease. B Exp Biol Med 2004; 138(9): 311-5. [68] Aggarwal BB, Kumar A, Bharti AC. Anticancer potential of

curcumin: preclinical and clinical studies. Anticancer Res 2003; 23(1A): 363-98.

[69] Suresh MV, Wagner MC, Rosania GR, et al. Pulmonary administration of water-soluble curcumin complex reduces ALI

severity. Am J Respir Cell Mol Biol 2012. [70] Park JH, Kim HA, Park JH, Lee M. Amphiphilic peptide carrier for

the combined delivery of curcumin and plasmid DNA into the lungs. Biomaterials 2012; 33(27): 6542-50.

[71] Cartiera MS, Ferreira EC, Caputo C, Egan ME, Caplan MJ, Saltzman WM. Partial correction of cystic fibrosis defects with

PLGA nanoparticles encapsulating curcumin. Mol Pharm 2010;

7(1): 86-93. [72] Egan ME, Pearson M, Weiner SA, et al. Curcumin, a major

constituent of turmeric, corrects cystic fibrosis defects. Science 2004; 304(5670): 600-2.

[73] Zeitlin P. Can curcumin cure cystic fibrosis? New Engl J Med 2004; 351(6): 606-8.

[74] Lim GP, Chu T, Yang FS, Beech W, Frautschy SA, Cole GM. The curry spice curcumin reduces oxidative damage and amyloid

pathology in an Alzheimer transgenic mouse. J Neurosci 2001; 21(21): 8370-7.

[75] Ono K, Hasegawa K, Naiki H, Yamada M. Curcumin has potent anti-amyloidogenic effects for Alzheimer's beta-amyloid fibrils in

vitro. J Neurosci Res 2004; 75(6): 742-50. [76] Yang FS, Lim GP, Begum AN, et al. Curcumin inhibits formation

of amyloid beta oligomers and fibrils, binds plaques, and reduces amyloid in vivo. J Biol Chem 2005; 280(7): 5892-901.

[77] Jiang T, Zhi X-L, Zhang Y-H, Pan L-F, Zhou P. Inhibitory effect of curcumin on the Al(III)-induced A 42 aggregation and

neurotoxicity in vitro. BBA-Mol Basis Dis 2012; 1822(8): 1207-15. [78] Anand P, Sundaram C, Jhurani S, Kunnumakkara AB, Aggarwal

BB. Curcumin and cancer: an "old-age" disease with an "age-old" solution. Cancer Lett 2008; 267(1): 133-64.

[79] Huang MT, Lou YR, Ma W, Newmark HL, Reuhl KR, Conney AH. Inhibitory effects of dietary curcumin on forestomach,

duodenal, and colon carcinogenesis in mice. Cancer Res 1994; 54(22): 5841-7.

[80] Rao CV, Rivenson A, Simi B, Reddy BS. Chemoprevention of colon carcinogenesis by dietary curcumin, a naturally-occurring

plant phenolic compound. Cancer Res 1995; 55(2): 259-66. [81] Sharma RA, Gescher AJ, Steward WP. Curcumin: the story so far.

Eur J Cancer 2005; 41(13): 1955-68. [82] Caruso F, Rossi M, Benson A, et al. Ruthenium–arene complexes

of curcumin: X-ray and density functional theory structure, synthesis, and spectroscopic characterization, in vitro antitumor

activity, and DNA docking studies of (p-cymene)Ru(curcuminato) chloro. J Med Chem 2011; 55(3): 1072-81.

[83] Lao CD, Ruffin MTt, Normolle D, et al. Dose escalation of a curcuminoid formulation. BMC Complem Altern M 2006; 6: 10.

[84] Martin P. Wound healing - aiming for perfect skin regeneration. Science 1997; 276(5309): 75-81.

[85] Hardy J, Selkoe DJ. Medicine - the amyloid hypothesis of Alzheimer's disease: progress and problems on the road to

therapeutics. Science 2002; 297(5580): 353-6. [86] Yanagisawa D, Shirai N, Amatsubo T, et al. Relationship between

the tautomeric structures of curcumin derivatives and their A -binding activities in the context of therapies for Alzheimer's

disease. Biomaterials 2010; 31(14): 4179-85. [87] Diaz-Cano SJ. General morphological and biological features of

neoplasms: integration of molecular findings. Histopathology 2008; 53(1): 1-19.

[88] Parkin DM, Bray F, Ferlay J, Pisani P. Global cancer statistics, 2002. CA Cancer J Clin 2005; 55(2): 74-108.

[89] Yallapu MM, Gupta BK, Jaggi M, Chauhan SC. Fabrication of curcumin encapsulated PLGA nanoparticles for improved

therapeutic effects in metastatic cancer cells. J Colloid Interf Sci 2010; 351(1): 19-29.

[90] Yallapu MM, Jaggi M, Chauhan SC. -Cyclodextrin-curcumin self-assembly enhances curcumin delivery in prostate cancer cells.

Colloid Surface B 2010; 79(1): 113-25. [91] Mohanty C, Sahoo SK. The in vitro stability and in vivo

pharmacokinetics of curcumin prepared as an aqueous nanoparticulate formulation. Biomaterials 2010; 31(25): 6597-611.

[92] Anitha A, Maya S, Deepa N, et al. Efficient water soluble O-carboxymethyl chitosan nanocarrier for the delivery of curcumin to

cancer cells. Carbohydr Polym 2011; 83(2): 452-61. [93] Bush JA, Cheung KJJ, Li G. Curcumin induces apoptosis in human

melanoma cells through a Fas receptor/caspase-8 pathway independent of p53. Exp Cell Res 2001; 271(2): 305-14.

[94] Jaiswal AS, Marlow BP, Gupta N, Narayan S. -Catenin-mediated transactivation and cell - cell adhesion pathways are important in

curcumin (diferuylmethane)-induced growth arrest and apoptosis in colon cancer cells. Oncogene 2002; 21(55): 8414-27.

[95] Nagai S, Kurimoto M, Washiyama K, Hirashima Y, Kumanishi T, Endo S. Inhibition of cellular proliferation and induction of


apoptosis by curcumin in human malignant astrocytoma cell lines.

Journal Neuro-Oncol 2005; 74(2): 105-11. [96] Jones PW, Taylor DM, Williams DR. Analysis and chemical

speciation of copper and zinc in wound fluid. J Inorg Biochem 2000; 81(1-2): 1-10.

[97] Sen CK, Khanna S, Venojarvi M, et al. Copper-induced vascular endothelial growth factor expression and wound healing. Am J

Physiol-Heart Circul Physiol 2002; 282(5): H1821-H7. [98] Valko M, Morris H, Cronin MTD. Metals, toxicity and oxidative

stress. Curr Med Chem 2005; 12(10): 1161-208. [99] Kehrer JP. Free-radicals as mediators of tissue-injury and disease.

Crit Rev Toxicol 1993; 23(1): 21-48. [100] Aizenman E, Stout AK, Harnett KA, Dineley KE, McLaughlin B,

Reynolds IJ. Induction of neuronal apoptosis by thiol oxidation: putative role of intracellular zinc release. J Neurochem 2000; 75(5):

1878-88. [101] Kehrer JP. The Haber-Weiss reaction and mechanisms of toxicity.

Toxicology 2000; 149(1): 43-50. [102] Miranda S, Opazo C, Larrondo LF, et al. The role of oxidative

stress in the toxicity induced by amyloid beta-peptide in Alzheimer's disease. Prog Neurobiol 2000; 62(6): 633-48.

[103] Yoshida D, Ikeda Y, Nakazawa S. Quantitative analysis of copper, zinc and copper/zinc ratio in selected human brain tumors. J Neuro-

Oncol 1993; 16(2): 109-15. [104] Lovell MA, Robertson JD, Teesdale WJ, Campbell JL, Markesbery

WR. Copper, iron and zinc in Alzheimer's disease senile plaques. J Neurol Sci 1998; 158(1): 47-52.

[105] Gupte A, Mumper RJ. Elevated copper and oxidative stress in cancer cells as a target for cancer treatment. Cancer Treat Rev

2009; 35(1): 32-46. [106] Atwood CS, Moir RD, Huang XD, et al. Dramatic aggregation of

Alzheimer a beta by Cu(II) is induced by conditions representing physiological acidosis. J Biol Chem 1998; 273(21): 12817-26.

[107] Brewer GJ, Yuzbasiyangurkan V. Wilson disease. Medicine 1992; 71(3): 139-64.

[108] Cherny RA, Atwood CS, Xilinas ME, et al. Treatment with a copper-zinc chelator markedly and rapidly inhibits beta-amyloid

accumulation in Alzheimer's disease transgenic mice. Neuron 2001; 30(3): 665-76.

[109] Sigurdsson EM, Brown DR, Alim MA, et al. Copper chelation delays the onset of prion disease. J Biol Chem 2003; 278(47):

46199-202. [110] Barik A, Mishra B, Kunwar A, et al. Comparative study of

copper(II)-curcumin complexes as superoxide dismutase mimics and free radical scavengers. Eur J Med Chem 2007; 42(4): 431-9.

[111] Barik A, Mishra B, Shen L, et al. Evaluation of a new copper(II)-curcumin complex as superoxide dismutase mimic and its free

radical reactions. Free Radical Bio Med 2005; 39(6): 811-22. [112] Baum L, Ng A. Curcumin interaction with copper and iron suggests

one possible mechanism of action in Alzheimer's disease animal models. J Alzheimers Dis 2004; 6(4): 367-77.

[113] Zhao X-Z, Jiang T, Wang L, Yang H, Zhang S, Zhou P. Interaction of curcumin with Zn(II) and Cu(II) ions based on experiment and

theoretical calculation. J Mol Struct 2010; 984(1-3): 316-25. [114] Krishnankutty K, Venugopalan P. Metal chelates of curcuminoids.

Syn React Inorg Met 1998; 28(8): 1313-25. [115] Dismukes JP, Jones LH, Bailar JC. Measurement of metal-ligand

bond vibrations in acetylacetonate complexes. J Phys Chem 1961; 65(5): 792-&.

[116] Addicoat MA, Metha GF, Kee TW. Density functional theory investigation of Cu(I)- and Cu(II)-curcumin complexes. J Comput

Chem 2010; 32(3): 429-38.

[117] Girolami GS, Jeffries PM, Dubois LH. Mechanistic studies of

copper thin-film growth from Cu(I) and Cu(II) beta-diketonates. J Am Chem Soc 1993; 115(3): 1015-24.

[118] Leung MHM, Lincoln SF, Kee TW. Femtosecond transient absorption spectroscopy of copper(II)-curcumin complexes. Phys

Chem Chem Phys 2012; 14(39): 13580-7. [119] Dutta S, Murugkar A, Gandhe N, Padhye S. Enhanced antioxidant

activities of metal conjugates of curcumin derivatives. Met-Based Drugs 2001; 8(4): 183-8.

[120] Lou JR, Zhang X-X, Zheng J, Ding W-Q. Transient metals enhance cytotoxicity of curcumin: potential involvement of the NF-kB and

mTOR signaling pathways. Anticancer Res 2010; 30(9): 3249-55. [121] Koiram PR, Veerapur VP, Kunwar A, et al. Effect of curcumin and

curcumin copper complex (1:1) on radiation-induced changes of anti-oxidant enzymes levels in the livers of Swiss albino mice. J

Radiat Res 2007; 48(3): 241-5. [122] Kunwar A, Narang H, Priyadarsini KI, Krishna M, Pandey R,

Sainis KB. Delayed activation of PKC delta and NF kappa B and higher radioprotection in splenic lymphocytes by copper(II)-

curcumin (1:1) complex as compared to curcumin. J Cell Biochem 2007; 102(5): 1214-24.

[123] Serpi C, Stanic Z, Girousi S. Electroanalytical study of the interaction between dsDNA and curcumin in the presence of

copper(II). Talanta 2010; 81(4-5): 1731-4. [124] Ahsan H, Hadi SM. Strand scission in DNA induced by curcumin

in the presence of Cu(II). Cancer Lett 1998; 124(1): 23-30. [125] Sakano K, Kawanishi S. Metal-mediated DNA damage induced by

curcumin in the presence of human cytochrome P450 isozymes. Arch Biochem Biophys 2002; 405(2): 223-30.

[126] Ahsan H, Parveen N, Khan NU, Hadi SM. Pro-oxidant, anti-oxidant and cleavage activities on DNA of curcumin and its

derivatives demethoxycurcumin and bisdemethoxycurcumin. Chem-Biol Interact 1999; 121(2): 161-75.

[127] Leung MHM, Mohan P, Pukala TL, Scanlon DB, Lincoln SF, Kee TW. Reduction of copper(II) to copper(I) in the copper-curcumin

complex induces decomposition of curcumin. Aust J Chem 2012; 65(5): 490-5.

[128] Kamau P, Jordan RB. Complex formation constants for the aqueous copper(I)-acetonitrile system by a simple general method.

Inorg Chem 2001; 40(16): 3879-83. [129] Kolthoff IM, Coetzee JF. Polarography in acetonitrile .2. metal ions

which have significantly different polarographic properties in acetonitrile and in water - anodic waves - voltammetry at rotated

platinum electrode. J Am Chem Soc 1957; 79(8): 1852-8. [130] Nair J, Strand S, Frank N, et al. Apoptosis and age-dependant

induction of nuclear and mitochondrial etheno-DNA adducts in Long-Evans Cinnamon (LEC) rats: enhanced DNA damage by

dietary curcumin upon copper accumulation. Carcinogenesis 2005; 26(7): 1307-15.

[131] Sahu SC, Washington MC. Effect of ascorbic-acid and curcumin on quercetin-induced nuclear-DNA damage, lipid-peroxidation and

protein-degradation. Cancer Lett 1992; 63(3): 237-41. [132] Yoshino M, Haneda M, Naruse M, et al. Prooxidant activity of

curcumin: copper-dependent formation of 8-hydroxy-2-deoxy-guanosine in DNA and induction of apoptotic cell death. Toxicol In

Vitro 2004; 18(6): 783-9. [133] Urbina-Cano P, Bobadilla-Morales L, Ramirez-Herrera MA, et al.

DNA damage in mouse lymphocytes exposed to curcumin and copper. J Appl Genet 2006; 47(4): 377-82.

Received: August 31, 2012 Accepted: October 22, 2012

delivery of curcumin and medicinal effects of the copper(ii)-curcumin complexes

Documents