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The inhibitory activity of aldose reductase in vitro by constituents of Garcinia mangostana Linn.

ABSTRACT

We investigated aldose reductase inhibition of Garcinia mangostana Linn, from Indonesia. Dichloromethane extract of the root bark of this tree was found to demonstrate an [IC.sub.50] value of 11.98 [micro]g/ml for human aldose reductase in vitro. From the dichloromethane fraction, prenylated xanthones were isolated as potent human aldose reductase inhibitors. We discovered 3-isomangostin to be most potent against aldose reductase, with an [IC.sub.50] of 3.48 [micro]M.

Keywords:

Aldose reductase

Mangostin

Garcinia mangostana

Xanthone

Diabetic complications

Introduction

The number of diabetes mellitus (DM) sufferers worldwide is increasing by about 35% annually. According to data from the IDF Diabetes Atlas, in 2013 the number of people with the disease has reached 382 million and is expected grow to 592 million people or more by 2035. The number of diabetics in Indonesia has reached 8.5 million, indicating that diabetes is now a serious problem there (IDF 2013). Diabetes mellitus is a chronic metabolic disorder characterized by excess glucose in the blood. Either the body cannot produce enough insulin (diabetes type I) or develops insensitivity in the insulin target cells, so that the body cannot use insulin effectively (diabetes type II). In DM type II indicates that it is caused by impaired insulin secretion and reduced [beta]-cell mass and appears to be a link between these two mechanisms (Talchai et al. 2009). Diabetes mellitus leads to long-term complications such as hypoglycemia, hyperglycemia, macro-vascular complications and micro-vascular complications which finally result in death.

Aldose reductase (alditol:[NAD(P).sup.+] 1-oxidoreductase, EC 1.1.1.21) is an enzyme which plays important role in the polyol pathway. This enzyme catalyzes the reduction of glucose to sorbitol with oxidation of NADPH to [NADP.sup.+]. Diabetic complications are considerately activated with increases of sorbitol (Bhatnagar and Srivastava 1992). One of common diabetic complication is cardiovascular autonomic neuropathy (CAN). Aldose reductase inhibitors can block the polyol pathway and have been used to deliberate the development of diabetic cardiovascular autonomic neuropathy (DCAN). Recently, the effectiveness and safety of ARIs in the treatment of DCAN have been reviewed. ARIs have been reported to improve cardiac automatic neuropathy especially mild or asymptomatic DCAN (Hu et al. 2014).

The use of traditional medicines has expanded internationally; they are becoming very popular as a health supplements worldwide. Recently, people have expressed interest in edible plants that can be used for diabetic treatment. In Indonesia, people use Garcinia mangostana Linn.as a health supplement for preventing everything from ulcer, diarrhea, fever, hypertension, and obesity to diabetes mellitus. G. mangostana Linn. is known for having a delicious fruit. This tree, also called "Manggis" in Indonesian, "Mangosteen" in English, "Manggoustan" in French, is known to bear the queen of tropical fruits. G. mangostana Linn, has been reported to produce many biologically active compounds especially xanthones (Syam et al. 2014; Wittenauer et al. 2012; Chae et al. 2012; Won et al. 2014, Sukma et al. 2011).

In the course of our investigation for potential aldose reductase inhibitors from natural sources, we focused on G. mangostana Linn. The aim of the current study was to evaluate the extract of this plant and its constituents for the ability to inhibit aldose reductase in vitro.

Materials and methods

Chemicals and reagents

Human recombinant aldose reductase (HRAR) and DL-glyceraldehyde were purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan), quercetin is kind of property from Laboratory of Systematic Forest and Forest Product Science, Kyushu University (Fukuoka, Japan) and [beta]-NADPH from Oriental Yeast Co., Ltd. (Osaka, Japan). Dichloromethane, n-hexane, ethyl acetate, acetone, dimethyl sulfoxide, methanol and all the other chemicals were of analytical grade.

Fractionation and isolation

Root bark of G. mangostana Linn. (Clusiaceae) was obtained from Kebun Raya Purwodadi (East Java, Indonesia). We ground 2.5 kg of root bark to powder. Then, extraction by dichloromethane (101) was performed three times at room temperature for 2 x 24 h and yielded 82 g dichloromethane extract. A further experiment partitioning with methanol and n-hexane 130 ml four times yielded a 2.5 g methanol fraction, a 47 g dichloromethane fraction and a 17 g n-hexane fractions.

A portion of the dichloromethane fractions (24 g) was fractionated by vacuum liquid chromatography employing a gradient of hexane to methanol resulting in 7 fractions: Fr A (711 mg), Fr B (2,2 g), Fr C (445 mg), Fr D (10.7 g), Fr E (659 mg), Fr F (6.4 g); Fr G (445 mg). Fraction D was subjected to vacuum liquid chromatography employing a hexane/methanol gradient to afford [alpha]-mangostin, [beta]-mangostin and 3-isomangostin. 1H, 13C, DEPT, HMQC and HMBC spectra were recorded on a JNM-AL400 FT NMR spectrometer (JEOL) (Mahabusarakam and Wiriyachitra 1987; Fukuyama et al. 1991; Ito et al. 1997).

Aldose reductase assay

Aldose reductase activity was determined spectrophotometrically on a JASCO V-530 UV- vis spectrophotometer. The activities HRAR were according to the procedure of Ueda (Ueda et al. 2004) with minor modifications. The reaction mixture contained 0.15 mM [beta]-NADPH, 10 mM DL-glyceraldehyde, 5 [micro]l of HRAR and 100 [micro]l of test sample solution or dimethyl sulfoxide (DMSO) in a total volume of 1.0 ml of 100 mM sodium phosphate buffer (pH 6.2). After the reaction mixtures were incubated at 25[degrees]C for 5 min in advance, the reaction was started by addition of the enzyme, and then the decrease of absorbance was measured at 340 nm for 10 min using a JASCO V-530 UV-vis spectrophotometer. Each plant extract was dissolved in dimethyl sulfoxide, which was found to have no effect on the enzyme activity at less than a 1% concentration. The inhibitory activity (%) was estimated as follows: [1 - ([DELTA]A sample/min - [DELTA]A control/min)] x 100. [DELTA]A sample/min showed a decrease of absorbance for 1 min with a sample and [DELTA]A control/min with DMSO instead of a sample.

Results and discussion

The present study shows the aldose reductase inhibitory activity of root bark of G. mangostana Linn., a plant that has been used for preventing diabetes mellitus in some regions of Indonesia. This is the first report on the aldose reductase inhibitory activity of C. mangostana Linn. The [IC.sub.50] value of aldose reductase from dichloromethane extract of G. mangostana Linn, was 11.98 [micro]g/ml (Fig. 1). A partition with n-hexane, dichloromethane, acetone, and methanol was carried out. The dichloromethane fraction was selected for further experimentation because it had the highest inhibition among the fractions. Thus, we continued the fractionation process from the extract to yield three known xanthones: [alpha]-mangostin, [beta]-mangostin and 3-isomangostin (Fig. 2).

In vitro assay results showed that 3-isomangostin had the highest aldose reductase inhibitory activity among isolated compounds with [IC.sub.50] value of 3.48 [micro]M, while [alpha]-mangostin and [beta]-mangostin had [IC.sub.50] values of 31.9 [micro]M and 30.6 [micro]M, respectively (Table 1). As a positive control, we used quercetin, a naturally occurring compound that has been demonstrated to be a potent aldose reductase inhibitor in vitro. The aldose reductase inhibitory values of isolated compounds from C. mangostana Linn, were compared to that of quercetin, with an [IC.sub.50] value of 2.98 [micro]M.

From Fig. 2, the only difference between the chemical structure of [alpha]-mangostin and [beta]-mangostin is the presence of methoxy instead of hydroxy at [alpha]-mangostin's position-three carbon. There was no difference in [IC.sub.50] values between the two compounds, which means that neither the methoxy nor hydroxy at this position is the source of major inhibition. The only difference between the chemical structure of 3-isomangostin and those of [alpha]-mangostin and [beta]-mangostin is the cyclization of the prenyl group at the position-two carbon. The 3-isomangostin as one xanthone derivative showed a significant value in the inhibition of aldose reductase since its [IC.sub.50] value was close to quercetin as the positive control.

Due to the limited number of currently available aldose reductase inhibitors, the result of our present study is significant for isolating a natural product that can potentially be optimized as an aldose reductase inhibitors. On the other hand, our results suggest that C. mangostana Linn, should be evaluated for controlling different diabetic complications through the pharmacologic action of inhibiting aldose reductase. The present study results open wide the possibilities for the use this genus in aldose reductase inhibition for treatment of diabetes.

In conclusion, three known xanthones, [alpha]-mangostin, [beta]-mangostin and 3-isomangostin, were isolated from dichloromethane fractions of the root bark of G. mangostana Linn. The aldose reductase inhibitory effects of the compounds were determined, and 3-isomangostin was found to be the most active against the enzyme when tested in vitro. These data validate mangosteen as an abundant natural source of potent aldose reductase inhibitors.

Conflict of interest

There is no conflict of interest.

ARTICLE INFO

Article history:

Received 26 June 2014

Revised 24 August 2014

Accepted 12 November 2014

Acknowledgments

This work was supported by a grant from research project for International Research Collaboration and Scientific Publication, Directorate General of Higher Education, Ministry of Education and Culture, Indonesia.

Supplementary materials

Supplementary material associated with this article can be found, in the online version, at doi:10.1016/j.phymed.2014.11.001.

References

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Chae, H.S., Oh, S.R., Lee, H.K., Joo, S.H., Chin, Y.W., 2012. Mangosteen xanthones, [alpha]-and [gamma]-mangostins, inhibit allergic mediators in bone marrow-derived mast cell. Food Chem. 134, 397-400.

Fukuyama, Y., Kamiyama, A., Mima, Y., 1991. Prenylated xanthones from Garcinia subelliptica. Phytochemistry 30, 3433-3436.

Hu, X., Li, S., Yang, G., Liu, H., Boden, G., Li, L, 2014. Efficacy and safety of aldose reductase inhibitor for the treatment of diabetic cardiovascular autonomic neuropathy: systematic review and meta-analysis. PLoS ONE 9, e87096. doi:10.1371/journal.pone.0087096.

IDF (International Diabetes Federation). Diabetes Atlas, 2013. http://www.idf.org/atlasmap/atlasmap. Accessed 13.05.14..

Ito, C., Miyamoto, Y., Nakayama, M., Kawai, Y., Rao Sundar, K., Furukawa, H., 1997. A novel depsidone and some new xanthones from Garcinia species. Chem. Pharm. Bull. 45, 1403-1423.

Mahabusarakam, W., Wiriyachitra, P., 1987. Chemical constituents of Garcinia mangostana. J. Nat. Prod. 50, 474-478.

Sukma, M., Tohda, M., Suksamran, S., Tantisira, B., 2011. [gamma]-Mangostin increases [serotonin.sub.2A/2C], muscarinic, histamine and bradykinin receptor mRNA expression. J. Ethnopharmacol. 135, 450-454.

Syam, S., Bustamam, A., Abdullah, R., Sukari, M.A., Hashim, N.M., Ghaderian, M., Rahmani, M., Mohan, S., Abdelwahab, S.I., Ali, H.M., 2014. [beta]-Mangostin induces p53-dependent G2/M cell cycle arrest and apoptosis through ROS mediated mitochondrial pathway and NfkB suppression in MCF-7 cells. J. Funct. Foods 6, 290-304.

Talchai, C., Lin, H.V., Kitamura, T., Accili, D., 2009. Genetic and biochemical pathways of beta-cell failure in type 2 diabetes. Diab. Obes. Metab. 11, 38-45.

Ueda, H., Kuroiwa, E., Tachibana, Y., Kawanishi, K., Ayala, F., Moriyasu, M., 2004. Aldose reductase inhibitors from the leaves of Myrda dubia (H. B. & K.) McVaugh. Phytomedicine 11, 652-656.

Wittenauer, J., Falk, S., Schweiggert-Weisz, U., Carle, R., 2012. Characterisation and quantification of xanthones from the aril and pericarp of mangosteens (Garcinia mangostana L) and a mangosteen containing functional beverage by HPLC-DAD-MSn. Food Chem. 134, 445-452.

Won, Y.S., Lee, J.H., Kwon, S.J., Kim, J.Y., Park, K.H., Lee, M.K., Seo, K.I., 2014. [alpha]-Mangostin-induced apoptosis is mediated by estrogen receptor [alpha] in human breast cancer cells. Food Chem. Toxicol. 66, 158-165.

Sri Fatmawati (a), *, Taslim Ersam (a), Kuniyoshi Shimizu (b)

(a) Department of Chemistry, Faculty of Mathematics and Natural Sciences, (nsritut Teknologi Sepuluh Nopember, Kampus ITS-Sukolilo, Surabaya 60111, Indonesia

(b) Department of Agro-environmental Sciences, Faculty of Agriculture, Kyushu University, Fukuoka 812-8581, Japan

* Corresponding author. Tel.: +62 31 594 3353; fax: +62 31 594 3353.

E-mail address: fatma@chem.its.ac.id (S. Fatmawati).

http://dx.doi.org/10.1016/j.phymed.2014.11.001

Table 1
Inhibitory effect of isolated compounds from
G. mangostana on aldose reductase activity.

Inhibitors                      [IC.sub.50] ([micro]M)

[alpha]-Mangostin               31.9
[beta]-Mangostin                30.6
3-Isomangostin                  3.48
Quercetin (positive control)    2.98
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Author:Fatmawati, Sri; Ersam, Taslim; Shimizu, Kuniyoshi
Publication:Phytomedicine: International Journal of Phytotherapy & Phytopharmacology
Article Type:Report
Date:Jan 15, 2015
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