Effect of Curcuma longa Rhizome Powder on Metabolic Parameters and Oxidative Stress Markers in High-Fructose and High-Fat Diet-Fed Rats

Metabolic disorders have become a major and growing global health problem, so fnding potentially novel solutions with fewer harm is favourable to solving this problem. Tus, this research aimed to determine the efect of a diet supplemented with Curcuma longa rhizome powder on markers of oxidative stress as well as biochemical and haematological parameters of rats with diet-induced metabolic disorders. Te powder was obtained from fresh Curcuma longa rhizome for preparation of extracts, which were used for in vitro analysis. After induction of metabolic disorders in male Wistar rats, they were separated into fve groups: base, control


Introduction
Environmental and behavioural changes such as the adoption of a westernised diet (high energy intake) and sedentary habits due to socioeconomic development are recognised as the main causes of metabolic disorders [1].
Tis complex disease state is defned by the occurrence of type 2 diabetes (insulin resistance), obesity, atherogenic hypertension, and dyslipidaemia that predisposes the population to an increased risk of developing cardiovascular diseases [1,2].Tese disorders are characterised by an increase in oxidative processes, promoting and aggravating other metabolic conditions, thus increasing death worldwide [3].Based on the diagnostic criteria, the world prevalence of these disorders varies from 13.6% to 46.0% [4].Improving means used to prevent risk factors include drug treatment and adjuvant therapy [5].However, the low efectiveness, high cost, and adverse efects of the latter have prompted the quest for the search of natural alternatives in the form of natural products, functional foods, or nutraceuticals [5].
Among these alternatives, spices have been recognised as high potential sources of renewable and biodegradable chemicals that are efcient in preventing and/or treating metabolic disorders.Indeed, spices are gaining interest due to anti-infammatory, carminative, antioxidant, and antiseptic properties, thanks to which their use in producing nutraceuticals or functional foods has been enhanced [6].
Curcuma longa, also referred to as turmeric, belongs to the family Zingiberaceae and has a golden colour with a slightly bitter taste.It is mostly produced in India and is also present in the Cameroonian fora.It is considered as an important spice used in most culinary preparations worldwide [5,6].Te phytochemical compounds of this plant are present in the rhizome, made up of 77% curcumin, 17% bisdemethoxycurcumin, and 3% demethoxycurcumin [7][8][9][10].Several research works had shown the benefcial efect of Curcuma longa in the treatment of various pathological states due to its antiinfammatory [11,12], antioxidant [13], antimicrobial, antibacterial, antiviral [14,15], antidiabetic [16], and antihypertension [17,18] properties.Curcumin competitively inhibits α-amylase and reduces blood glucose levels in diabetic rats [16,19,20].It also has protective properties against teratogenic, cytotoxic, and neurodegenerative diseases [21,22] and hepatoprotective [23] and nephroprotective activities against hepato-renal failure [24,25].To beneft from the health properties of this spice, it is consumed in the form of powder added to food during cooking, essential oils (either as nutraceutical or supplemented with food), and nanoparticles [6,23].Te essential oils as well as nanoparticles are not afordable for LMICs due to their high production costs.Tese populations mainly rely on the culinary powder form without information on the proportion that can efciently provide the health benefts awaited.Tis work was thus aimed to assess the efect of diet supplementation with various proportions of Curcuma longa rhizome powder on metabolic parameters and oxidative stress markers of rat-induced metabolic disorders.

Materials and Methods
2.1.Plant Material Sampling.Curcuma longa rhizomes were harvested in Santchou (Menoua Division, West Cameroon) and transported to the Research Unit of Biochemistry, Medicinal Plants, Food Sciences and Nutrition (URBP-MAN) for further studies.Tereafter, the plant identifcation was done at the National Herbarium of Cameroon and the reference number 42173/HNC was assigned.

Preparation of Powder.
Fresh rhizomes of Curcuma longa were cleaned, sliced, and oven-dried (Venticell, Turkey) at 45 °C for 48 hours.Te dried rhizomes were later ground using a blender (Singsung, model no: BL-500, Singapore) to obtain the Curcuma longa rhizome powder.

Preparation of Extract.
Te extracts were prepared following the method reported by Womeni et al. [26].25 g of powder was introduced in three diferent beakers containing, respectively, 500 ml of distilled water, 500 ml of ethanol 90%, and 500 ml of mixed ethanol and distilled water (50% V/V); the beakers were then manually shaken every 3 hours during 48 h storage at room temperature (25 °C) in order to obtain aqueous, hydroethanolic, and ethanolic extracts.Te mixtures were fltered using a Whatman paper (No. 1), and the fltrates were subsequently oven-dried at 45 °C until complete evaporation of the solvent.Te resulting extracts were stored at 4 °C, protected from light.

Total Phenolic Content (TPC)
Assessment.Te total phenolic content (TPC) was determined using the modifed method of Gao et al. [27].In brief, 20 μl of extracts (2 mg/ml) was mixed, respectively, with 2 ml of distilled water and 200 μl of Folin-Ciocalteu reagent, and the mixture was incubated for 3 min at room temperature.Tereafter, the mixture received 1 ml of sodium carbonate and was reincubated for 2 hours at room temperature, and the absorbance of the resulting blue colour was measured at 765 nm using a spectrophotometer (BIOMATE, France).For standard curve preparation, a 0.2 g/l gallic acid solution was used, and the results were expressed as mg gallic acid equivalents per gram of powder extract (GAE/g).

Total Flavonoid Assessment (TFC).
Te total favonoid content (FC) was assessed thanks to the method of Marinova et al. [28].Tus, 100 μl of Curcuma longa rhizome powder extracts (2 mg/ml) was mixed with 1.4 ml of distilled water and 30 μl of sodium nitrate (5% m/v).After 5 min, 200 μl of aluminium trichloride (10% w/V) was added, followed by the addition of 200 μl of sodium hydroxide (10% w/V) and 240 μl of distilled water after another 5 min.Te mixture was then shaken, and the absorbance was measured at 510 nm using a spectrophotometer (BIOMATE, France).For standard curve preparation, a 0.1 mg/ml solution of catechin was used, and the results were expressed as mg catechin equivalents per gram of powder extract (CAE/g).

Evaluation of DPPH Free Radical Scavenging.
Te DPPH free radical scavenging activity of the rhizomes powder extracts was evaluated using the method of Mensor et al. [29].In this experiment, 1 ml of a 0.3 mM methanolic solution of DPPH was mixed with 2.5 ml of each extract at increasing concentrations (12.5, 25, 50, 100, and 200 μg/ml).Tese solutions were held in the dark at room temperature for 30 min, after which, the optical density (OD) was measured at 517 nm.A 2 mg/ml vitamin C solution was used as a positive control.Te radical scavenging activity (RSA) was evaluated using the following formula: (1)

Ferric Reducing Antioxidant Power (FRAP) Assessment.
To determine the ferric reducing antioxidant power (FRAP) of powder extracts, their activity to reduce Fe 3+ to Fe 2+ was evaluated as described by Oyaizu [30].In glass tubes containing 0.5 ml of powder extracts at increasing concentrations (12.5, 25, 50, 100, and 200 μg/ml), were added 1 ml of 0.2 M phosphate bufered saline at pH 6.6 and 1 ml of 1% of aqueous potassium hexacyanoferrate solution.After incubating the mixture at 50 °C for 30 min, 1 ml of 10% trichloroacetic acid was introduced, followed by centrifugation for 10 min at 3000 × g.Te supernatant (1.5 ml) was removed and mixed with 1.5 ml of distilled water as well as 0.1 ml of iron trichloride aqueous solution 0.1% (w/V), and the absorbance of the mixture was measured at 700 nm.Vitamin C (2 mg/ml) prepared under the same conditions was used as control.An increase in the absorbance of the mixture attested a rise in reducing the power of the powder extracts to reduce Fe 3+ to Fe 2+ .
2.8.Alpha-Amylase Inhibition Assay.Te alpha-amylase inhibition assay was performed using the method reported by Manikandan et al. [31].Te starch solution (0.1% w/V) was obtained by adding 0.1 g of starch to 100 ml of sodium acetate bufer (16 mM).Ten, the enzyme solution was obtained by dissolving 27.5 mg of alpha-amylase in 100 ml of distilled water.To prepare the colorimetric reagent, 3,5-dinitrosalicylic acid (96 mM) and sodium potassium tartrate solutions were mixed together.Subsequently, the control and powder extracts were added to the starch solution and allowed to react with the alphaamylase solution under alkaline conditions at 37 °C and the reaction was measured over 15 min.Te reaction products released were measured using a spectrophotometer at 540 nm. ).Te report on this section adheres to the guidelines for reporting animal research [32].
Te modifed base diet proposed by Telefo et al. [33] was used in this study.Te high-fat diet was prepared in the laboratory by mixing the powdered ingredients and fat (beef suet) as shown in Table 1.

Experimental Design and Collection of Blood and Organs.
Te experimental rats were divided into fve groups of 4 rats each, and each rat was fed with 45 g of diet per day.Te induction of metabolic disorder in rats and the administration of Curcuma longa powder supplementation in order to regulate metabolic parameters lasted for 12  During the supplementation period, the weekly body weight and food intake were recorded.After a four week treatment period, the animals were allowed to fast for 12 h, followed by anaesthetisation with diazepam/ketamine (0.2 : 0.1 ml/100 g of animal), euthanisation, and blood collection by cardiac puncture in dry and EDTA tubes for haematological analyses.Te blood samples in dry tubes were centrifuged at 3000 g for 15 min at 4 °C, 3 hours following their collection.Separated serum was transferred into cryotubes and stored at −20 °C for further analysis.Liver, peritoneal, and mesenteric fats were immediately collected and weighed.One gram (1 g) of liver tissue was homogenised in 10 ml of phosphate bufer (pH 7.5), centrifuged at 3000 g, and used for in vivo antioxidant tests.

Animal's Glycaemia and Lee Index.
Te device used for the measurement of blood glucose was a commercial glucometer (VERI Q, MGD-2001, Germany).On VERI Q strips, we placed a drop of fresh blood from the tail of the rat, which was allowed for 45 seconds to read the blood glucose value from the beginning to the end of the experiment.Body weight as well as the length from the nose to the anus was measured to evaluate the Lee index, which permitted determining the state of obesity in rats.Tis index was calculated by the formula which determines the link between the weight and the length of the animal from the nose to the anus [34].Day 0 (day before high-fructose high-fat diet administration) marked the start of recording parameters, while day 1 represented the day the groups fed high-fructose high-fat diet were confrmed with glycaemia ≥180 mg/dl and Lee index ≥300 in order to verify the diagnoses of insulin resistance and obesity.
where W is the body weight (g) and L is the length from the nose to the anus (cm).

Determination of Nitric Oxide (NO) Concentration.
Te NO concentrations of samples were determined by the methods described by Tracey et al. [36].For this purpose, 340 μl of sample was introduced into the tubes, then 340 μl of sulfanilamide (1% w/V) prepared in orthophosphoric acid (1% V/ V) was added, and the mixtures were kept in the dark for 5 min at room temperature.Ten, 340 μl of naphthylenediamine (NED, 1% w/V) was added to the reaction medium, and the whole solution was once more left in the dark for 5 min at room temperature.Te optical densities of samples were measured at 520 nm.Te nitric oxide concentration (NO) was determined from a standard curve obtained from various concentrations of sodium nitrate (0.1 mM).

Determination of Malondialdehyde (MDA) Concentration.
Te MDA concentration of the samples was determined following the methods proposed by Niehaus and Samuelsson [37].In tubes containing 100 μl of sample, 500 μl of thiobarbituric acid (TBA, 1% V/V) prepared in 1% trichloroacetic acid and 500 μl of phosphoric acid (1% V/V) were added.Te mixture was incubated in a water bath at 100 °C for 15 min and then cooled.Subsequently, the tubes were centrifuged at 3000 g for 10 min at room temperature (25 °C) and the absorbance of the resulting pink colour of the supernatant was measured at 532 nm.Te blank contained 100 µl of distilled water and was treated under the same conditions.Te concentration of MDA was calculated using the following formula: where Ab is the average absorbance, Vt is the total volume of the reaction medium, α is the molecular extinction coefcient (α: 1.53 × 105 M −1 cm −1 ), and V is the sample volume.

Determination of Catalase Activity.
Te catalase activity of the samples was assessed by their ability to reduce hydrogen peroxide (H 2 O 2 ) to water (H 2 O) and oxygen (O 2 ) as described by Sinha [38].In tubes containing 50 μl of sample, 750 μl of phosphate bufer and 200 μl of 50 mM hydrogen peroxide (H 2 O 2 ) were added, followed by addition of 2 ml of 5% dichromate 1 min later.Te tubes were rigorously shaken and heated for 10 min in boiling water.After cooling the tubes, the absorbance of the green colour generated was measured at 570 nm.Te catalase activity (CAT) was determined using the following formula: where Ab is the average absorbance, Vt is the total volume of the reaction medium, α is the molar extinction coefcient (40 M −1 cm −1 ), and V is the sample volume.

Statistical Analysis.
Te results of the biochemical parameters, markers of oxidative stress, total phenolic and favonoid contents, and haematological parameters were frstly expressed as mean ± standard deviation.Ten, they were analyzed using one-way analysis of variance (ANOVA), and Waller-Duncan post hoc test was used to compare the signifcance between the mean values using the IBM SPSS Statistics 22 (Chicago, IL, USA).Signifcance was considered for a p value <0.05.In addition, Pearson's correlation between total phenolic content (TPC), favonoid content (FC), DPPH free radical scavenging (DPPH), ferric reducing antioxidant power (FRAP), and inhibition of alpha-amylase (IAA) was evaluated using SPSS version 22.

Inhibition of Alpha-Amylase.
Te inhibition of alphaamylase by the diferent extracts is grouped in Table 4. Te Curcuma longa rhizome extracts revealed an important inhibitory action on the alpha-amylase enzyme.Te inhibition signifcantly increased (p < 0.05) with the increasing concentration of Curcuma longa rhizome extracts from 250 to 1000 μg/ml.Te hydroethanolic extract showed the best inhibition activity (94.12 ± 2.84%) on the enzyme, followed by the ethanolic extract (62.52 ± 3.71%) and the aqueous extract being the lowest (42.19 ± 3.43%) at the dose 1000 μg/ml.

Discussion
Plants' natural products represent the safest, efective, and alternative source of chemical drugs.Te latter possess side efects; hence, researchers are turning to the use of plants to solve health problems.In the current study, the results of the phytochemical analysis showed that the TPC in the hydroethanolic extract was higher than that of Najah et al.
[39] (51.46 ± 0.46 mg GAE/g) on methanol/chloroform extracts of Curcuma longa rhizome powder.In contrast, the TPC obtained in this study was signifcantly less than those reported by Choi [40] in ethyl acetate and chloroform extracts of Curcuma longa rhizome from Korea with respective values of 228.7 ± 2.3 and 140.7 ± 10.6 mg GAE/g extract.According to Shan et al. [41], the diferences in the level of TPC and FC may be due to the type of solvent used for extraction, time of harvesting samples, determination methods, and the diferences in the geographical conditions from one country to another such as climate, temperature, fertility, and disease.Regarding antioxidant activity results, the higher TPC in hydroethanolic extract (about 1.30-fold that of ethanolic extract and 1.57-fold that of aqueous extract) might explain its higher DPPH radical scavenging activity.Tese results are in accordance with those of Najah et al. [39] and Sabir et al. [42] which showed that DPPH scavenging activity of methanol/chloroform and ethanolic extract of Curcuma longa rhizome increased with the TPC.Also, the highest FRAP activity observed with the hydroethanolic extract may be associated with the high amount of hydroxyl groups thanks to phenolic compounds such as favonoids that can play the role of electron donors, hereby reducing Fe 3+ to Fe 2+ [43].An increase in the quantity of Fe 2+ results in the rise of the reducing power of the extract.As such, antioxidants are considered as reducers and inactivators of oxidants [44].

Journal of Food Biochemistry
Te in vitro inhibition of alpha-amylase by the diferent extracts of Curcuma longa rhizome showed a dosedependent increase in the percentage of inhibition.Te phytochemical compounds present in the extracts could account for the inhibition of the digestive enzymes that are needed for the production of energy [45].Tese compounds have hydroxyl functions similar to the substrates of amylase, which can induce competitive inhibition of the enzyme.Aqueous extract presented the lowest activity, which could be due to the low total phenolic and favonoid contents, while hydroethanolic extract showed the best inhibitory activity of alpha-amylase because of the high TPC and FC.Tese results are consistent with those of Manikandan et al. [31] and Henda et al. [46] that demonstrated that extracts from Psidium guajava and Juniperus phoenicea leaves, respectively, rich in phytochemical compounds presented inhibitory activities of alpha-amylase.
Looking at the signifcant positive correlation recorded between the phytochemical compounds' contents and the antioxidant properties of the diferent Curcuma longa rhizome powder extracts, there is a relation between the TPC, FC, DPPH, and FRAP because the extract that shows a high antioxidant activity, taking into consideration the DPPH and FRAP, also has the highest TPC and FC.Te results showing the positive and signifcant correlation between TPC, DPPH, and FRAP are in line with those of Choi [40] who showed a signifcant positive correlation between these parameters with methanolic extract of Curcuma longa rhizome.
Te in vitro antioxidant and amylase inhibitory activities of Curcuma longa rhizome powder were used to assess its efect on metabolic parameters in rat-induced metabolic disorders.Te consumption of a high-fat and high energy diet and sedentary lifestyles are recognised as paramount causes of the development of metabolic disorders [47,48].As shown in this study, rats that ate a high-fructose high-fat diet developed obesity by increasing their Lee index.Lee index is a major parameter for assessing the efect of a high-fructose high-fat diet on the development of obesity and for monitoring its treatment.When the Lee index of an animal is greater or equal to 300, it is obese [34].Te development of obesity was due to the excess of fat contained in the hyperlipidic diet, leading to an imbalance between food energy intake and the energy expenditure necessary for daily activities.Moreover, the consumption of a high-fructose high-fat diet supplemented with Curcuma longa rhizome powder signifcantly decreased Lee index of the obese rats; this may be due to the presence of phytochemical compounds in the powder that inhibited digestive enzymes and decreased the energy intake of animals [49].Te glycaemia test performed during the study also showed an increase in blood glucose level, and this may be due to the deposition of fat at the level of pancreas tissues that block the insulin activity and lead to a higher blood glucose level in rats fed with a high-fructose high-fat diet.Tese results were similar to those of Shoumen et al. [47] and Akiyama et al. [50], who showed that rats fed on a high-fructose high-fat diet developed hyperglycemia.Moreover, Curcuma longa rhizome powder supplementation decreased the glycaemia of the high-fructose high-fat diet-fed rats.Tese results could be due to the presence of phytochemical compounds in the Curcuma longa rhizome powder which inhibited the digestion of carbohydrates and as such reduced the absorption of glucose and postprandial blood glucose level [51].
Regarding the lipid profle results, the consumption of a high-fructose high-fat diet induced metabolic disorders with an increase in serum total cholesterol, LDLc, and triglycerides levels compared to the base group, while HDLc remained low.Tis could be justifed by the fact that the fat (beef suet) used for the preparation of the high-fat diet has caused hypercholesterolemia with the deposit of cholesterol plaques in the vascular walls in rats sufering from metabolic disorders.Te more severe the metabolic disorder is, the more signifcant the HDLc decreases [52].Tis decrease in good cholesterol (HDLc) and increase in bad cholesterol (LDLc) is due to high blood glucose levels, which is a sign of  [53].Based on the current investigation, Curcuma longa rhizome powder supplementation reduced the triglycerides, total-cholesterol, and LDL-cholesterol and increased the HDL-cholesterol in high-fructose high-fat diet-fed rats.Te decrease in cholesterol rate in rats that consumed Curcuma longa rhizome powder could be due to the absorption of phytochemical compounds which inhibited the activity of hydroxymethylglutaryl-CoA reductase.Tis enzyme permits endogenous synthesis of cholesterol by transforming acetyl coenzyme-A in mevalonic acid, an intermediate in the synthesis chain of endogenous cholesterol [54].Tis result could also be due to the presence of polyunsaturated fatty acids in Curcuma longa rhizome powder since the works of Munshi et al. [55] have shown that oil extracted from Curcuma longa rhizome has important content of linoleic and linolenic acids.Adipogenesis, known as cell formation, is a diferentiation process which converts undiferentiated preadipocytes into fully diferentiated adipocytes that store energy in the form of fat and render subjects obese [56].Adipose tissue is a dynamic organ whose mass changes throughout life in response to the animal's metabolic needs and therefore plays an important role in energy balance.Obesity-associated genes such as leptin, adiponectin, and fatty acid synthase regulate adipogenesis and lipid metabolism at diferent stages of adipocyte diferentiation [57].It has been reported that excess fat is positively correlated with leptin concentration in rodents and humans [58].Leptin is a protein secreted by adipocytes that plays a crucial role in the regulation of body weight by controlling the size of adipose tissue.It regulates food intake by binding to receptors in the central nervous system and modulating the activity of neurons present in the brain's appetite control centers [59].In our study, the phytochemical compounds present in the powder could lower leptin [60] expression in animals fed with a highfructose high-fat diet supplemented with Curcuma longa rhizome powder, resulting in a drop in food intake (reduced appetite), which in turn could lead to a reduction in serum triglyceride and cholesterol levels, which have a direct impact on the amount of fat stored in adipocytes.Adiponectin, acting as an antagonist of adipogenesis, plays an efective role in regulating lipid and glucose metabolism in insulin-sensitive organs in animals and humans [61].Low circulating adiponectin levels have been demonstrated in genetic and diet-induced obesity models [62].It has been reported that upregulation of adiponectin levels is inversely correlated with fat mass and insulin resistance [63].Terefore, in our study, the decrease in blood glucose, serum lipid concentrations, and body weight in rats receiving a high-fructose high-fat diet supplemented with Curcuma longa rhizome powder may be due to the upregulation of adiponectin levels by the phytochemicals present in the powder.Te antiobesity activity observed in rats fed a high-fructose high-fat diet supplemented with Curcuma longa rhizome powder could be due to the property of phytochemical compounds that have the capacity to modify energy expenditure by increasing the expression of lipolytic proteins or the inhibition of lipogenesis by reducing the expression of fatty acid synthase.Similar reports of body weight reduction have been reported previously with extracts of some plant species [60,[64][65][66].Te consumption of high-fructose high-fat diet in most experimental models to induce metabolic disorder also caused oxidative stress [47].As shown in the present experiment, the levels of lipid peroxidation products (MDA and NO) were increased in high-fructose high-fat diet-fed rats.Tis increase in oxidative stress molecules might be associated with the decrease in CAT activity recorded in the present work.Tese results were congruous with previous studies revealing that tissue antioxidant defenses may be compromised in high-fructose high-fat diet-fed animals [67,68].Curcuma longa rhizome powder supplement prevented oxidative stress and restored the tissue antioxidants in high-fructose high-fat diet-fed rats.Tis result may be explained by the presence of phytochemical compounds that positively modulate free radical metabolism by increasing the activities of antioxidant enzymes and decreasing the concentration of lipid peroxidation products (MDA and NO) [69].
Experimentally, the use of a high-fructose high-fat diet permits us to get closer to the new nutritional habits of industrialised countries.Te results of the present work showed that the consumption of a high-fructose high-fat diet signifcantly increased the body weight gain of the animals.Tis increase in body weight gain may be due to the presence of fats in the food that is likely to be accumulated in adipose tissue, leading to the development of obesity [68,70].Curcuma longa rhizome powder supplementation signifcantly decreased the body weight gain of the high-fructose high-fat diet-fed rats.Tis result can be justifed by the presence of phytochemical compounds in the powder that inhibited digestive enzymes and reduced the absorption of nutrients [45].Te results of liver weight and fat deposition showed an increase in high-fructose high-fat diet-fed rats.However, Curcuma longa rhizome powder supplemented rats exhibited signifcantly reduced liver weight and deposition of peritoneal and mesenteric fats.Tis result may be due to the presence of phytochemical compounds in the Curcuma longa rhizome powder that inhibited the digestive enzymes in a consequence of the utilisation of fats stored in adipose tissue for the production of energy necessary for the functioning of the organism.Tis leads to the reduction of peritoneal and mesenteric fats as well as the weight of the liver.Tese results are in line with those of Manikandan et al. [31].Te hepatic damage was evaluated by measuring the activities of the liver enzyme markers.Serum ALT and AST increase when hepatic damage occurs and the activities of these enzymes were measured in all groups.Te present study showed that the serum AST and ALT levels increased in high-fructose high-fat diet-fed rats compared to the base group.Hepatocyte damage caused by the consumption of a high-fructose high-fat diet is evident in most experimental models and patients with clinical conditions [50].Tese enzymes are considered markers of hepatic dysfunction.Generally, hepatocyte damage causes these enzymes to be transported in the serum [71].Induction of metabolic disorder results in the beginning of oxidative stress in several tissues, including the liver.Tis leads to the peroxidation of membrane lipids, altering hepatic lipid profle, which Journal of Food Biochemistry consequently cause cellular damage and membrane rupture, with the release of AST and ALT into the bloodstream [72,73].
Te haematological parameters' results showed that the induction of metabolic disorder leads to a signifcant decrease in white blood cells (WBC), red blood cells (RBC), haemoglobin (HGB), haematocrit (HCT), mean corpuscular haemoglobin concentration (MCHC), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), and platelet (PLT) level.Feeding on Curcuma longa rhizome powder supplementation signifcantly increased their level.Nutrients in food play vital roles in the normal function of the body.Inclusions of active secondary metabolites in human nutrition have been demonstrated to have health benefts and reduce the risk of chronic diseases such as cancer, obesity, diabetes, and anaemia [67,70].Reduction in these parameters could be due to the coexistence of metabolic disorder and anaemia in the same animal.HCT stands for erythrocyte volume fraction of the percentage of red blood cells.RBC are involved in the transport of oxygen and absorb nutrients [74]; thus a high HCT value shows better transportation of oxygen.Elsewhere, WBC help the organism's defense through phagocytosis, transportation, and distribution of antibodies, leading to immunity.Tus, individuals with low levels of WBC are exposed to a high risk of infection [75].Te platelet count is one of the most important screen tests of platelet functions.A decrease in the circulating platelet of less than 15% of the normal value will cause bleeding [76].Te reduction observed in the platelet count in the blood of high-fructose high-fat diet-fed rats compared to the base group may suggest a possible efect on blood clotting in rats sufering from metabolic disorders.In other fndings, haemoglobin levels also decreased in rats sufering from metabolic disorders, probably due to the presence of anaemia.Erythrocyte indices do not defne the cause of anaemia but rather may be useful during the diagnostic workup.Tese results are supported by those of Monteomo et al. [77] and Ochuko et al. [78] which show that rats have low haemoglobin concentration and haematocrit after consuming a high-fat diet and have similarities with iron defciency anaemia.Changes in the haematology parameters in metabolic disorders were dominated by hypochromic anaemia corresponding to a decrease in erythrocyte, haemoglobin, haematocrit, reticulocyte contents (MCH and MCHC), and MCV values.Te increase in the haematology parameters after food supplementation with Curcuma longa rhizome powder may be due to the presence of phytochemical compounds in this rhizome, but the mechanism still remains unknown.

Conclusion
Tis study reveals that the consumption of a high-fructose high-fat diet alters the normal biochemical parameters and leads to the development of metabolic disorders (obesity, hyperglycaemia, and dyslipidaemia) and oxidative stress.Food supplementation with Curcuma longa rhizome powder, an excellent source of phytochemical compounds, improved these metabolic parameters and oxidative stress markers by a signifcant reduction of Lee index, glycaemia, triglyceride, total-cholesterol, LDLc, MDA, and NO levels.Tis supplementation also upgraded haematological parameters (WBC, RBC, HGB, HCT, MCHC, MCV, MCH, and PLT) with the best hepatoprotective efects at 5% supplementation.Further research is needed to confrm these benefcial efects on a human subject through a clinical trial.

Table 2
presents the total phenolic content (TPC) and favonoid content (FC) of Curcuma longa rhizome extracts.It appears that TPC and
3.2.DPPH Free Radical Scavenging and Ferric ReducingAntioxidant Power.Table3illustrates the DPPH free radical scavenging and ferric reducing antioxidant power (FRAP) activities of Curcuma longa rhizome powder extracts.It can be observed that the antioxidant activities are signifcantly infuenced (p < 0.05) by the concentration of the extracts.At 12.5 μg/ml, representing the lowest concentration tested, the hydroethanolic extract scavenged 34.95 ± 2.05% DPPH radical compared to 18.98 ± 2.81% for ethanolic extract and only 7.91 ± 2.67% for aqueous extract.At the maximum tested concentration (200 µg/ml), a strong DPPH scavenging activity (79.46 ± 2.12%) was evaluated in hydroethanolic extract compared to the ethanolic and aqueous extracts.
Curcuma longa Rhizome Powder Supplementation on the Lee Index of High-Fructose High-Fat Diet-Fed Rats.Te Lee index of the rats in various groups is represented in Table6.High-fructose high-fat diet-fed rats showed a signifcantly (p < 0.05) increased Lee index (326.88±3.08)as compared to the base group (287.43 ± 4.32) after induction of metabolic disorder.Te glycaemia test was performed to evaluate the efect of consumption of Curcuma longa rhizomes powder supplementation on the fasting blood glucose after 28 days.Te results of the glycaemia test are depicted in Table7.At the start of the experiment, the initial blood glucose of the fve (5)groups was approximately 127 mg/dl.Te glucose concentration of the high-fructose high-fat diet-fed rats signifcantly (p < 0.05) increased (189.25 ± 5.98 mg/dl) compared to the base group (134.5 ± 3.10 mg/dl) after induction of metabolic disorder.Curcuma longa rhizome powder supplementation at 10% in high-fructose high-fat diet-fed rats signifcantly (p < 0.05) decreased the fnal blood glucose concentration (120.5 ± 4.91 mg/dl) compared to the control group (186 ± 6.94 mg/dl) after supplementation.3.7.Efect of the Curcuma longa Rhizome Powder Supplementation on the Lipid Profles of the High-Fructose High-FatDiet-Fed Rats.Table8displays the lipid profle of rats.It can be highlighted that serum triglyceride, total cholesterol, and LDL-cholesterol levels were signifcantly (p < 0.05) increased in rats that consumed the high-fructose high-fat diet (158.35 ± 5.37 mg/dl, 208.53 ± 1.53 mg/dl, and 128.73 ± 3.64 mg/dl, respectively) compared to the base group rats (128.43 ± 1.11 mg/dl, 124.40 ± 4.63 mg/dl, and 39.26 ± 3.78 mg/dl, respectively).Curcuma longa rhizome powder high-fructose high-fat diet-fed rats (74.21 ± 1.04 mg/dl) compared to the control group rats (48.12 ± 1.82 mg/dl).3.8.Efect of Curcuma longa Rhizome Powder Supplementation on Serum and Liver Levels of Oxidative Stress Markers inHigh-Fructose High-Fat Diet-Fed Rats.Te results of the

Table 2 :
Total phenolic content (TPC) and favonoid content (FC) of Curcuma longa rhizome extracts.Table10lays out the body weight of rats during the treatment period.

Table 3 :
DPPH free radical scavenging and ferric reducing antioxidant power of Curcuma longa rhizome extracts.
iValues (n � 3) with diferent superscript letters in the same row are signifcantly diferent (p < 0.05) according to Waller-Duncan post hoc test.

Table 4 :
In vitro alpha-amylase inhibition by Curcuma longa rhizome extracts.

Table 6 :
Efect of Curcuma longa rhizome powder supplementation on the Lee index in the high-fructose high-fat diet-fed rats.

Table 7 :
Efect of the Curcuma longa rhizome powder supplementation on glycaemia (mg/dl) in the high-fructose high-fat diet-fed rats.

Table 8 :
Efect of the Curcuma longa rhizome powder supplementation on the lipid profle (mg/dl) in the high-fructose high-fat diet-fed rats.Changes in the functions of hepatic enzymes are shown in Figure1.Serum AST activity was signifcantly increased (p < 0.05) (84.21 ± 3.66 U/L) in highfructose high-fat diet-fed rats compared to the base group (53.81 ± 1.12 U/L).Food supplementation with Curcuma longa

Table 9 :
Efect of the Curcuma longa rhizome powder supplementation on serum and liver levels of oxidative stress markers in high-fructose high-fat diet-fed rats.

Table 10 :
Efect of the Curcuma longa rhizome powder supplementation on the body weight gain (g) in high-fructose high-fat diet-fed rats.

Table 11 :
Efect of Curcuma longa rhizome powder supplementation on the food intake (g/day) in high-fructose high-fat diet-fed rats.

Table 12 :
Efect of Curcuma longa rhizome powder supplementation on liver wet weight and peritoneal and mesenteric fat wet weight (g/ 100 g of body weight) of high-fructose high-fat diet-fed rats.