Original Papers

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Authors

Rebamang A. Mosa 1, Javan J. Naidoo 1, Fezile S. Nkomo 1, Sithandiwe E. Mazibuko 1, 2, Christo J. F. Muller 2, Andy R. Opoku 1

Affiliations

1 2

Key words " Protorhus longifolia l " Anacardiaceae l " triterpenes l " adipogenesis l " antihyperlipidemic activity l

received revised accepted

March 22, 2014 Sept. 19, 2014 October 12, 2014

Bibliography DOI http://dx.doi.org/ 10.1055/s-0034-1383262 Published online November 11, 2014 Planta Med 2014; 80: 1685–1691 © Georg Thieme Verlag KG Stuttgart · New York · ISSN 0032‑0943 Correspondence Rebamang A. Mosa Department of Biochemistry and Microbiology University of Zululand Private Bag X1001 KwaDlangezwa 3886 Republic of South Africa Phone: + 27 3 59 02 60 99 Fax: + 27 3 59 02 65 68 [email protected]

Department of Biochemistry and Microbiology, University of Zululand, KwaDlangezwa, Republic of South Africa Diabetes Discovery Platform, Medical Research Council (MRC), Tygerberg, South Africa

Abstract !

Two lanostane triterpenes, 3β-hydroxylanosta9,24-dien-21-oic acid (1) and methyl-3β-hydroxylanosta-9,24-dien-21-oate (2), were isolated from the stem bark of Protorhus longifolia. Their structures were deduced on the basis of spectroscopic analysis (NMR, HRMS, IR). This study investigated the in vitro anti-adipogenic activity of the two triterpenes. Their inhibitory activity was evaluated on selected lipid digestive enzymes (pancreatic lipase and cholesterol esterase). The inhibitory activity of the compounds on hormone-sensitive lipase and their ability to bind bile acids were also evaluated. The effect of the compounds on glucose uptake in C2C12 muscle cells and 3T3-L1 adipocytes, and on triglyceride accumulation in 3T3-L1 adipocytes was investigated. The triterpenes effectively inhibited the activities of the enzymes with IC50 values ranging from 0.04 to 0.31 mg/mL. The compounds showed a high affinity for secondary bile acids. Both compounds stimulated glucose uptake in C2C12 muscle cells and 3T3-L1 adipocytes. Compound 1

Introduction !

Obesity, caused by the increased accumulation of lipid within the fat tissues, is becoming a major global health problem [1]. Pathophysiologically, obesity is associated with conditions such as chronic inflammation, hyperlipidemia, hypertension, diabetes, and cardiovascular disease [2]. Its prevalence is currently increasing at an alarming rate worldwide [3]. Some of the factors that cause obesity include a sedentary lifestyle and an imbalance between energy intake and expenditure. Currently antiobesity therapies include drugs such as tetrahydrolipstatin (orlistat) that inhibit intestinal lipase activity, thereby reducing fat absorption or alternatively sibutramine (Reductil)

significantly reduced triglyceride accumulation in mature differentiated 3T3-L1 adipocytes. It is apparent that these lanostane triterpenes enhance glucose uptake and suppress adipogenesis, which together with their inhibitory effects on lipid digestive enzymes suggests that they have antihyperlipidemic potential.

Abbreviations !

DCA: DMEM: FCS: GCA: HS: HSL: IBMX: KRB: LCA: TCA:

deoxycholic acid Dulbeccoʼs modified Eagleʼs medium fetal calf serum glycocholic acid horse serum hormone sensitive lipase 3-isobutyl-1-methylxanthine Krebs-Ringer bicarbonate buffer lithocholic acid taurocholic acid

Supporting information available online at http://www.thieme-connect.de/products

that suppresses food intake [4]. However, these drugs have unpleasant side effects [5, 6]. The use of natural antiobesity products is gaining popularity mainly because they are perceived to be effective with less side effects. These natural products act either by increasing cellular energy expenditure and/or by inhibiting adipogenesis and stimulating lipolysis in adipocytes [3, 7]. Plants have always been a rich reservoir of bioactive compounds with diverse therapeutic properties. Consumption of plant sterols and their esters has been reported to not only lower intestinal cholesterol absorption but to decrease blood levels of the atherogenic low-density lipoprotein cholesterol [8, 9]. Antihyperglycemic [10], antihyperlipidemic [11], and anti-inflammatory activ-

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Planta Med 2014; 80: 1685–1691

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In Vitro Antihyperlipidemic Potential of Triterpenes from Stem Bark of Protorhus longifolia

Original Papers

Fig. 1 Chemical structures of 3β-hydroxylanosta-9,24-dien-21-oic acid (1) and methyl-3βhydroxylanosta-9,24-dien-21-oate (2).

ities [12] of some triterpenes from various plants have been reported. Antiobesity properties of some other triterpenoid compounds such as ursolic acid and acetyl-keto-β-boswellic acid have also recently been documented [3, 13]. The diverse pharmacological effects possessed by plant-derived triterpenes have made these compounds new targets for drug development against an array of life-threatening diseases. The stem bark of Protorhus longifolia (Benrh.) Engl. (Anacardiaceae), a tree indigenous to South Africa, is traditionally used to cure various diseases including bleeding from the stomach, heart burn, and hemiplegic paralysis [14]. Crude extracts and lanosteryl triterpenes isolated from the stem bark of the plant have been reported to exhibit antiplatelet aggregation activity [15, 16]. In this study, the activity of the two lanosteryl triterpenes isolated from P. longifolia on lipid enzymes, cellular glucose uptake, and lipid accumulation in 3T3-L1 adipocytes was investigated.

Results and Discussion ! " Fig. 1) were estabThe structures of the isolated compounds (l lished and confirmed through 1H and 13C NMR. The physical and spectral data of 3β-hydroxylanosta-9,24-dien-21-oic acid (1) have been previously described in Mosa et al. [16]. The IR spectrum showed absorption bands for hydroxyl (3360, 2581 cm−1) and carbonyl (1702 cm−1) functional groups, which further confirmed the structure. The 1H‑NMR of compound 2 followed the same triterpenoid pattern with a large cluster of signals of CH3, CH2, and CH between δH 2.5 and 0.8 observed in 3β-hydroxylanosta-9,24-dien-21-oic acid [16]. The 13C‑NMR of this compound was also similar to that of 3β-hydroxylanosta-9,24-dien-21-oic acid, with the presence of four olefinic carbon atoms between 145–118 ppm, and five quaternary carbon atoms confirming the lanosteryl skeletal structure. The presence of an ester carbon atom at δC 177.3 instead of a carboxylic carbon at δC 181.5 suggested that this compound is the methyl ester of 3β-hydroxylanosta-9,24-dien-21-oic " Table 1 presents a detailed assignment of the 13C‑NMR acid. l and significant 1H‑NMR of the triterpene. The absorption bands for hydroxyl (3469 cm−1) and carbonyl (1683 cm−1) functional groups observed on the IR spectrum also further assisted in con-

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Table 1 13C‑NMR data and significant 1H‑NMR data of compound 2. Chemical shifts are expressed in δ (ppm). Position

δC (ppm)

Type

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 -OCH3

30.4 23.9 74.3 37.4 44.4 17.3 25.8 49.6 145.9 34.7 118.3 28.6 43.3 51.0 31.3 27.0 47.7 13.3 21.7 48.4 177.3 34.7 25.5 125.6 136.4 17.4 25.8 21.9 27.4 22.1 59.8

CH2 CH2 CH C CH CH2 CH2 CH C C CH CH2 C C CH2 CH2 CH CH3 CH3 CH C CH2 CH2 CH C CH3 CH3 CH3 CH3 CH3

δH (ppm)

4.25 (1H, s, OH)

5.11 (1H, t)

5.21 (1H, t) 1.61 (3H, s) 1.65 (3H, s) 1.21 (3H, s) 0.90 (3H, s) 1.16 (3H, s) 3.85 (3H, s)

firming the NMR structure. See Supporting Information for spectra of the compounds. Inhibition of lipid digestive enzymes and, therefore, limiting intestinal absorption of dietary lipids, as well as inhibition of adipogenesis and lipolysis could be an ideal therapeutic approach to reduce incidences of hyperlipidemia and its related diseases [17, 18]. The literature lists some plant-derived triterpenes such as ursolic acid, oleanolic acid, and acetyl-keto-β-boswellic acid as having antiobesity properties [3, 13, 19]. The two lanosteryl triterpenes isolated from the stem bark of P. longifolia effectively inhibited the activities of pancreatic lipase, cholesterol esterase, and HSL " Table 2). with IC50 values ranging from 0.04 to 0.31 mg/mL (l These IC50 values are comparable to those of the positive controls used. Inhibitors of HSL are important drug targets in the prevention of hyperlipidemia and consequent peripheral insulin resistance [20]. The inhibitory activity of the triterpenes from P. longifolia on pancreatic lipase and HSL is evidence that both compounds have the potential to regulate dyslipidemia by not only inhibiting absorption of exogenous lipids but lypolysis as well. Bile acid sequestrants are important in limiting the absorption of dietary lipids, stimulating cholesterol catabolism, and preventing diseases associated with secondary bile acids accumulation [21, 22]. The bile acid binding ability of the compounds was deter" Fig. 2. mined on some bile acids, and the results are shown in l Both triterpenes exhibited a concentration-dependent affinity for bile acids. A higher affinity for the secondary bile acids (DCA

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Sample

Pancreatic lipase

Hormone sensitive lipase

Cholesterol esterase

Compound 1 Compound 2 Orlistat Simvastatin

0.15 ± 0.02 0.17 ± 0.01 0.01 ± 0.06 –

0.04 ± 0.05a 0.24 ± 0.11 0.01 ± 0.09a –

0.31 ± 0.07 0.11 ± 0.04b – 0.16 ± 0.01

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Table 2 Inhibitory activity (IC50 mg/mL) of the triterpenes on some lipid digestive enzymes.

Fig. 2 Bile acid binding ability of the triterpenes. Bile acids (0.5–4 mM) were incubated with the test compound at 1 % w/v for 30 min at 37 °C. The concentration of the residual bile acids was determined, and the results are presented as the percentage of adsorption ability of the compounds on the various bile acids. Data are expressed as mean ± SEM. a P < 0.05 compared to 1; b p < 0.05 compared to 2.

and LCA) indicates the potential benefits of the compounds in maintaining cholesterol homeostasis and prevention of colorectal cancer and chronic diarrhea [21, 22]. The 3T3-L1 cell line is widely used to study adipogenesis, lipolysis, and lipid metabolism. These adipocytic cells have contributed to the current understanding of the biochemical mechanisms that antiobesity agents have at a cellular level. Compound 1 at 1, 10, and 25 µM concentrations significantly (p ≤ 0.05) reduced triglyceride accumulation in differentiated 3T3-L1 adipocytes fol" Fig. 3). However, there was no effect lowing a 48-h treatment (l observed at the highest concentration of 100 µM, thus indicating a concentration-dependent activity. The crystal violet stain used to normalize the cell count confirmed that the observed effect was not due to cytotoxicity. It is apparent that the anti-adipogenic activity of compound 1 could also be through the inhibition of endogenous triglyceride synthesis. The reduction of triglyceride accumulation in 3T3-L1 adipocytes as a measure of anti-adipogenic activity has been previously described for a number of plant-derived compounds [23], including lanostane triterpenes from the fruiting bodies of Ganoderma lucidum [24]. An increase in adipocytes size due to excess lipid accumulation is characteristic in obesity [25]. Therefore, the ability of the compound to reduce lipid accumulation in adipocytes suggests that this compound could have antiobesity activity and therefore potentially protect against the development of serious metabolic disease such as diabetes [25]. The effect of the triterpenes on cellular glucose uptake was evaluated in C2C12 myotubules and 3T3-L1 adipocytes. Both compounds at 50 µg/mL effectively stimulated cellular glucose uptake " Fig. 4). Lee and Phuong [26] also reported stimulation of cellu(l

lar glucose uptake in L6 myotubules by triterpenoids from Weigela subsessilis. Some other medicinal plants have shown the ability to enhance cellular glucose uptake without stimulating adipogenesis [27]. It is apparent that the triterpenes from P. longifolia also have a similar effect. The ability of hypolipidemic agents to lower blood glucose levels is important for the management of diabetes mellitus [28]. This study demonstrated the potential antiobesity activity of the two triterpenes from P. longifolia. Apparently, their antiobesity activity could be through the inhibition of exogenous lipid absorption, stimulation of lipolysis, and increased glucose uptake in muscle and fat tissue. Further inhibitory studies on the enzyme kinetics are required to elucidate the mechanism(s) whereby the compound affects lipid accumulation. Further studies such as glycerol release, a measure of lipolysis, and adipogenic studies involving gene and protein expression are recommended. An evaluation of beneficial effects of the triterpenes on metabolic disorders in vivo is necessary for further studies. The in vivo study of compound 2 on hyperlipidemia has recently been evaluated [29]. The compound effectively reduced serum cholesterol levels, the atherogenic index, and the coronary risk index.

Materials and Methods !

Cell lines, chemicals, and reagents The 3T3-L1 (CL-173) fibroblast and C2C12 myoblasts (CRL-1772) were obtained from American Type Culture Collection. The bile acid kit (Cat No: BQ 042A-EALD) was purchased from BQ Kits. HS was obtained from Highveld Biological, DMEM from Lonza,

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Data are expressed as the mean ± SEM. a P < 0.05 vs. 2; b p < 0.05 vs. 1

Original Papers

Fig. 3 Effect of compound 1 on lipid accumulation. 3T3-L1 cells were exposed to the compound at a range of concentrations (1, 10, 25, and 100 µM) for 48 h. A Undifferentiated controls showed a lack of staining (I); differentiated 3T3-L1 media control cells (II) and vehicle controls (III) showed an increased lipid accumulation, while treatment with the compound (IV) reduced lipid accumulation. B Quantification of the intracellular lipid accumulation. Data are expressed as mean of three independent experiments relative to DMSO at 100 %; * p ≤ 0.05 and ** p ≤ 0.001 vs. control (ADM). (Color figure available online only.)

and FCS from GIBCO/Invitrogen Life Technologies. Orlistat (≥ 98 % purity), cholestyramine (≥ 98% purity), insulin (≥ 95 % purity), and metformin (≥ 97% purity) were purchased from Sigma-Aldrich. All organic solvents used were purchased from Merck. Unless otherwise stated, all other chemicals and reagents used were obtained from Sigma-Aldrich Chemical Co. All the chemicals and reagents were of analytical grade.

Plant material Fresh plant material (stem barks) of P. longifolia was collected in March 2012 from Hlabisa, KwaZulu-Natal, South Africa. The plant was collected under the authority of the indigenous knowledge system (IKS), authenticated by Dr. N. R. Ntuli, Department of Botany, University of Zululand, and a voucher specimen (RA01UZ) was deposited in the University herbarium. The air-dried plant material was ground to powder and stored in a sterile brown bottle at 4 °C until required.

Extraction and isolation The method of extraction and isolation of the triterpenes from the stem bark of P. longifolia was followed as previously described by Mosa et al. [16]. The powdered plant material was first defatted with n-hexane and then extracted (1 : 5 w/v) with chloroform. The triterpenes were isolated from the chloroform extract (13 g) using silica gel column chromatography (24 × 700 mm; silica gel 60; 0.063–0.2 mm; 70–230 mesh ASTM, Merck), and

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eluted with a n-hexane-ethyl acetate solvent system (9 : 1 to 3 : 7, gradient). The active compounds 1 (0.72 g) and 2 (1.15 g) were obtained following their further purification in n-hexane and ethyl acetate, respectively. See Supporting Information for details.

Compounds Physical and spectral data of 3β-hydroxylanosta-9,24-dien-21oic acid (1) have previously been given by Mosa et al. [16]. Methyl-3β-hydroxylanosta-9,24-dien-21-oate (2) was obtained as white crystals, > 95 % pure, m. p. 204–205 °C, IR (KBr) vmax = " Table 1) data suggested 3469, 1683 cm−1. 1H and 13C NMR (see l the molecular formula C31H50O3, calcd. 470.736. The compound dissolves in DMSO and is highly soluble in methanol.

Preparation of compounds for enzyme activity assays The compounds were dissolved in methanol at a concentration of 2000 µg/mL (Stock) and stored at 4 °C. For the working solutions, the stock solution was further diluted with methanol to yield final concentrations of 25, 50, 100, 250, 500, and 1000 µg/mL.

Pancreatic lipase inhibition Inhibitory activity of the triterpenes on pancreatic lipase was evaluated following the method described by Slanc et al. [30]. The enzyme activity was determined by measuring the release of p-nitrophenol in the presence and absence of the test com-

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pound (0–500 µg/mL in methanol) at 405 nm using a BioTek® plate reader (ELx 808 UI, BioTek Instrument Supplies) equipped with Gen5 software. Three independent experiments, with each experiment replicated three times, were done and the mean ± SEM are reported. The percentage of inhibition was calculated using the formula % Inhibition = [(Ac − At)/Ac] × 100 where Ac = absorbance of control sample and At = absorbance of test sample. IC50 values were determined using GraphPad Prism v 5.03.

Hormone-sensitive lipase extraction HSL was extracted from the rat epididymal adipose tissue of Sprague-Dawley male rats as previously described [28, 31]. Briefly, fat cell suspension in Krebs-Ringer bicarbonate buffer supplemented with 4 % BSA (KRB‑BSA, pH 7.4) was diluted (1.0 : 1.125 mL) with homogenization buffer (100 mL; 50 mM Tris-HCl, pH 7.0, 250 mM sucrose, and one crushed protease inhibitor tablet). HSL was then obtained following a series of centrifugations, and diethyl ether (250 µL) was used to dissolve the fat contents. Following centrifugation of the homogenate at 1200 × g, the diethyl ether layer was aspirated and the subsequent supernatant was taken as the HSL extract. The HSL extract was stored at − 80 °C until required.

Hormone-sensitive lipase inhibition The inhibitory effect of the triterpenes on HSL activity was evaluated as described for the pancreatic lipase activity [30].

ported. The percentage of inhibition was calculated using the formula % Inhibition = [(Ac – At)/Ac] × 100

In vitro bile acid binding assay In vitro bile acid binding activity of the triterpenes was evaluated following the method described by Matsumoto et al. [33]. Different concentrations (0.5–4.0 mM) of each bile acid, GCA, TCA, DCA, and LCA, were used. Cellulose and cholestyramine were used as negative and positive controls, respectively. The concentration of the residual bile acid (water insoluble binding products) was determined using a bile acid kit following the manufacturerʼs instructions. The experiment was replicated three times. The percentage of adsorption of the compounds was then calculated using the formula %Adsorption ¼

ðAstd sample  Atest sampleÞ  100 Astd sample

Preparation of compounds for cell culture The compounds were dissolved in 100 % DMSO at a concentration of 87 mM and stored in 0.2 mL vials at − 80 °C. A stock solution was prepared by diluting the aliquoted compounds in DMSO with DMEM without phenol red to yield a concentration of 1 mM. For the working solutions, the stock solution was further diluted in DMEM without phenol red to yield final concentrations of 1, 10, 25, and 100 µM, respectively. The final concentration of DMSO present in the working solution was < 0.1 %.

Cell culture Cholesterol esterase inhibition The inhibitory activity of the triterpenes on cholesterol esterase was evaluated following the method described by Pietsch and Gütschow [32] with some modifications. The composition of the reaction mixture was 50 µL of 100 mM sodium phosphate buffered saline (pH 7.0), 50 µL of 5.16 mM taurocholic acid, 50 µL of 100 µg/mL cholesterol esterase in the buffer, 50 µL of the compound (0–1000 µg/mL in methanol), and 75 µL of 0.2 mM paranitrophenol-butyrate (p-NPB) in 6% acetonitrile. In the controls, the test compound was replaced with a mixture of methanol and water (1 : 1). The mixture was incubated at 25 °C for 10 min before the addition of the substrate. The reaction mixture was further incubated for 5 min at 25 °C. Simvastatin (≥ 97%, Sigma) was used as a standard drug. The absorbance was read at 405 nm. Three independent experiments, with each experiment replicated three times, were done and the mean ± SEM are re-

3T3-L1 fibroblasts were cultured and differentiated as previously described by Dudhia et al. [34] with slight modifications. Briefly, cells were seeded into 24-well culture plates at a density of 20 × 104 cells per well. The cells were cultured in DMEM growth medium with 10 % newborn calf serum (NCS) at 37 °C in humidified air with 5% CO2 until confluent. The confluent cells were differentiated in differentiating media containing DMEM with 10 % NCS, 1 µM dexamethasone, 1 µg/mL insulin, and 0.5 mM IBMX for three days. After differentiation, the media were substituted with adipocyte maintenance medium (DMEM, 10% FCS, 1 µg/mL insulin), and cells were cultured for a further two days (day five post-differentiation). On day five, the media was changed to insulin-free DMEM containing 10% FCS and the cells were cultured for a further two days (day seven). On day seven, the fully differentiated 3T3-L1 adipocytes were treated with different concentrations (1, 10, 25, and 100 µM) of the triterpene and the vehicle

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Fig. 4 Effect of the triterpenes on C2C12 (A) and 3T3-L1 (B) glucose uptake. The cells were exposed to 50 µg/mL of the compound in serum-free media supplemented with 8 mM glucose for 1 h at 37 °C. Data are expressed as the mean of three independent experiments relative to the glucose (8 mM) control at 100%; * p ≤ 0.05; *** p ≤ 0.0001.

Original Papers

control containing 0.1 % DMSO for 48 h. After the treatment, the intracellular lipid content was quantified. C2C12 myoblasts were cultured and differentiated as previously described by Muller et al. [35]. Briefly, C2C12 myoblasts were seeded into 24-well culture plates at a density of 25 × 104 cells per well in DMEM containing 10 % FCS until confluence. Upon confluence, myoblasts were then differentiated for two days in DMEM containing 2 % HS at 37 °C in humidified air and 5 % CO2 prior to performing the glucose uptake assay.

Intracellular lipid content determination The lipids accumulated in the adipocytes were quantified using the method described previously by Sanderson et al. [36].

Glucose uptake assay The glucose uptake assays were performed on 3T3-L1 adipocytes and C2C12 myocytes using the method previously described by Muller et al. [35] with slight modifications. Briefly, C2C12 myocytes and 3T3-L1 adipocytes were serum starved for 30 min using Krebʼs buffer containing 0.1 % BSA, 3.7 g/L NaHCO3, and no glucose. The cells were then exposed to the compounds (50 µg/mL) in serum-free media supplemented with 8 mM glucose, 3.7 g/L NaHCO3, and 0.1% BSA, and incubated for 1 h at 37 °C in humidified air with 5 % CO2. The media were collected and a glucose oxidase assay kit was used to determine the amount of glucose taken up by the cells. Glucose (8 mM) was used for the normal control, while insulin and metformin (each at a concentration of 1 µM) were included as positive controls.

Statistical analysis Data are presented as the mean ± SEM. Statistical differences were determined using one-way analysis of variance (ANOVA), followed by Dunnettʼs post hoc test if p ≤ 0.05 was deemed to be significant (GraphPad Prism version 5.03).

Supporting information Details about the plant material, extraction and isolation procedure, physical data of compound 1, as well as spectra of compounds 1 and 2 are available as Supporting Information.

Acknowledgements !

The studentship awarded to J. J. Naidoo and F. S. Nkomo by the National Research Foundation of South Africa is acknowledged. The authors are grateful to the South African Medical Research Council and University of Zululand Research Committee for funding this work.

Conflict of Interest !

Authors declare no conflict of interest

References 1 Chan RSM, Woo J. Prevention of overweight and obesity: how effective is the current public health approach. Int J Environ Res Public Health 2010; 7: 765–783 2 Greenberg AS, Obin MS. Obesity and the role of adipose tissue in inflammation and metabolism. Am J Clin Nutr 2006; 83: 461S-465S 3 He Y, Li Y, Zhao T, Wang Y, Sun C. Ursolic acid inhibits adipogenesis in 3T3-L1 adipocytes through LKB1/AMPK pathway. PLoS ONE 2013; 8: e70135

Mosa RA et al. In Vitro Antihyperlipidemic …

Planta Med 2014; 80: 1685–1691

4 Yun JW. Possible anti-obesity therapeutics from nature. Phytochemistry 2010; 71: 1625–1641 5 Chaput JP, St-Pierre S, Tremblay A. Currently available drugs for the treatment of obesity: sibutramine and orlistat. Mini Rev Med Chem 2007; 7: 3–10 6 Díaz EG, Folgueras TM. Systematic review of the clinical efficacy of sibutramine and orlistat in weigth loss, quality of life and its adverse effects in obese adolescents. Nutr Hosp 2011; 26: 451–457 7 Seo JB, Choe SS, Jeong HW, Park SW, Shin HJ, Choi SM, Park JY, Choi EW, Kim JB, Seen DS, Jeong JY, Lee TG. Anti-obesity effects of Lysimachia foenum-graecum characterized by decreased adipogenesis and regulated lipid metabolism. Exp Mol Med 2011; 43: 205–215 8 Sudhahar V, Kumar SA, Sudharsan PT, Varalakshmi P. Protective effect of lupeol and its ester on cardiac abnormalities in experimental hypercholesterolemia. Vasc Pharmacol 2007; 46: 412–418 9 Brown AW, Hang J, Dussault PH, Carr TP. Plant sterol and stanol substrate specificity of pancreatic cholesterol esterase. J Nutr Biochem 2010; 21: 736–740 10 Ghosh T, Maity TK, Singh J. Antihyperglycemic activity of bacosine, a triterpene from Bacopa monnieri, in alloxan-induced diabetic rats. Planta Med 2011; 77: 804–808 11 Liu J, Sun H, Wang X, Mu D, Liao H, Zhang L. Effects of oleanolic acid and maslinic acid on hyperlipidemia. Drug Dev Res 2007; 68: 261–266 12 Yadav VR, Prasad S, Sung B, Kannappan R, Aggarwal BB. Targeting inflammatory pathways by triterpenoids for prevention and treatment of cancer. Toxins 2010; 2: 2428–2466 13 Liu JJ, Toy WC, Liu S, Cheng A, Lim BK, Subramaniam T, Sum CF, Lim SC. Acetyl-keto-β-boswellic acid induces lipolysis in mature adipocytes. Biochem Biophys Res Commun 2013; 431: 192–196 14 Hutchings A, Scott AH, Lewis G, Cunningham A. Zulu medicinal Plants: an Inventory. Pietermaritzburg: University of Natal Press; 1996 15 Mosa RA, Lazarus GG, Gwala PE, Oyedeji AO, Opoku AR. In vitro antiplatelet aggregation, antioxidant and cytotoxic activity of extracts of some Zulu medicinal plants. J Nat Prod 2011; 4: 136–146 16 Mosa RA, Oyedeji OA, Shode FO, Singh M, Opoku AR. Triterpenes from the stem bark of Protorhus longifolia exhibit anti-platelet aggregation activity. Afr J Pharm Pharmacol 2011; 5: 2698–2714 17 Ahn JH, Liu Q, Lee C, Ahn MJ, Yoo HS, Hwang BY, Lee MK. A new pancreatic lipase inhibitor from Broussonetia kanzinoki. Bioorg Med Chem Lett 2012; 22: 2760–2763 18 Park CH, Chung BY, Lee SS, Bai HW, Cho JY, Jo C, Kim TH. Radiolytic transformation of rotenone with potential anti-adipogenic activity. Bioorg Med Chem Lett 2013; 23: 1099–1103 19 de Melo CL, Queiroz MG, Fonseca SG, Bizerra AM, Lemos TL, Melo TS, Santos FA, Rao VS. Oleanolic acid, a natural triterpenoid, improves blood glucose tolerance in normal mice and ameliorates visceral obesity in mice fed a high-fat diet. Chem Biol Interact 2010; 185: 59–65 20 Ali YB, Verger R, Carrière F, Petry S, Muller G, Abousalham A. The molecular mechanism of human hormone-sensitive lipase inhibition by substituted 3-phenyl-5-alkoxy-1,3,4-oxadiazol-2-ones. Biochimie 2012; 94: 137–145 21 Peterlik M. Role of bile acid secretion in human colorectal cancer. Wien Med Wochenschr 2008; 158: 539–541 22 Dibaise JK, Islam RS. Bile acids: an underrecognized and underappreciated cause of chronic diarrhea. In: Parrish CR, editor. Practical gastroenterology. Nutrition issues in gastroenterology, Series 110. Westhampton Beach: Practical Gastroenterology Publishing; 2012 23 Liu Q, Ahn JH, Kim SB, Hwang BY, Lee MK. New phenolic compounds with anti-adipogenic activity from the aerial parts of Pulsatilla koreana. Planta Med 2012; 78: 1783–1786 24 Lee I, Kim H, Youn U, Kim J, Min B, Jung H, Na M, Hattori M, Bae K. Effect of lanostane triterpenes from the fruiting bodies of Ganoderma lucidum on adipocyte differentiation in 3T3-L1 cells. Planta Med 2010; 76: 1558–1563 25 Zeng XY, Zhou X, Xu J, Chan SMH, Xue CL, Molero JC, Ye JM. Screening for the efficacy on lipid accumulation in 3T3-L1 cells is an effective tool for the identification of new anti-diabetic compounds. Biochem Pharmacol 2012; 84: 830–837 26 Lee MS, Phuong TT. Stimulation of glucose uptake by triterpenoids from Weigela subsessilis. Phytother Res 2010; 24: 49–53 27 Yang MH, Avula B, Smillie T, Khan I, Khan SI. Screening of medicinal plants for PPARα and PPARγ activation and evaluation of their effects on glucose uptake and 3T3-L1 adipogenesis. Planta Med 2013; 79: 1084–1095

This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited.

1690

Original Papers

33 Matsumoto K, Kadowaki A, Ozaki N, Takenaka M, Ono H, Yokoyama S, Gato N. Bile acid-binding ability of Kaki-tannin from young fruits of Persimmon (Diospyros kaki) in vitro and in vivo. Phytother Res 2011; 25: 624–628 34 Dudhia Z, Louw J, Muller C, Joubert E, de Beer D, Kinnear C, Pheiffer C. Cyclopia maculata and Cyclopia subternata (honeybush tea) inhibits adipogenesis in 3T3-L1 pre-adipocytes. Phytomedicine 2013; 20: 401– 408 35 Muller CJF, Joubert E, de Beer D, Sanderson M, Malherbe CJ, Fey SJ, Louw J. Acute assessment of an aspalathin-enriched green rooibos (Aspalathus linearis) extract with hypoglycemic potential. Phytomedicine 2012; 20: 32–39 36 Sanderson M, Mazibuko SE, Joubert E, de Beer D, Johnson R, Pheiffer C, Louw J, Muller CJF. Effects of fermented rooibos (Aspalathus linearis) on adipocyte differentiation. Phytomedicine 2014; 2: 109–117

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28 Bustanji Y, Issa A, Mohammad M, Hudaib M, Tawah K, Alkhatib H, Almasir I, Al-Khalid B. Inhibition of hormone sensitive lipase and pancreatic lipase by Rosmarinus officinalis extract and selected phenolic constituents. J Med Plants Res 2010; 4: 2235–2242 29 Machaba KE, Cobongella SZZ, Mosa RA, Lawal AO, Djarova TG, Opoku AR. In vivo anti-hyperlipidemic activity of the triterpene from the stem bark of Protorhus longifolia (Benrh) Engl. Lipids Health Dis 2014; 13: 131–137 30 Slanc P, Doljak B, Kreft S, Lunder M, Janeš D, Štrukelj B. Screening of selected food and medicinal plant extracts for pancreatic lipase inhibition. Phytother Res 2009; 23: 874–877 31 Morimoto C, Sumiyoshi M, Kameda K, Tsujita T, Okuda H. Relationship between hormone-sensitive lipolysis and lipase activity in rat fat cells. J Biochem 1999; 125: 976–981 32 Pietsch M, Gütschow M. Synthesis of tricyclic 1,3-oxazin-4-ones and kinetic analysis of cholesterol esterase and acetylcholinesterase inhibition. J Med Chem 2005; 48: 8270–8288

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Mosa RA et al. In Vitro Antihyperlipidemic …

Planta Med 2014; 80: 1685–1691

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In vitro antihyperlipidemic potential of triterpenes from stem bark of Protorhus longifolia.

Two lanostane triterpenes, 3β-hydroxylanosta-9,24-dien-21-oic acid (1) and methyl-3β-hydroxylanosta-9,24-dien-21-oate (2), were isolated from the stem...
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