Original Article

Effects of Oleuropein on Epirubicin and Cyclophosphamide Combination Treatment in Rats


  • Metin Deniz KARAKOÇ
  • Selim SEKKİN

Received Date: 25.06.2020 Accepted Date: 24.09.2020 Turk J Pharm Sci 2021;18(4):420-429 PMID: 34496482


Oleuropein is the main bioactive polyphenolic compound in olive leaves, olive, and olive oil. Its anticancer, antioxidant, and antiinflammatory effects have been proven through several in vitro and in vivo studies. This study aimed to explore the effects of oleuropein on cyclophosphamideand epirubicin-induced toxicity in female rats.

Materials and Methods:

Seven groups containing eight rats in each group were formed. Four cycles of 16 mg/kg/week of cyclophosphamide and 2.5 mg/kg/week of epirubicin were administered to the rats through intraperitoneal injection. Oleuropein (150 mg/kg/week) was simultaneously applied via oral gavage. The effects of oleuropein were examined with hemogram tests in whole blood samples and biochemical analysis in serum samples. Interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in the serum samples were analyzed through enzyme-linked immunosorbent assay. Subsequently, a comet assay was performed using lymphocyte DNA. The levels of oxidant [i.e., malondialdehyde (MDA)] and antioxidants [i.e., catalase (CAT), glutathione (GSH), and superoxide dismutase (SOD)] were measured in the heart, kidney, and liver tissues.


Oleuropein could reduce DNA damage and serum TNF-α and IL-6 levels. It also ameliorated some hemogram and biochemical parameters that deteriorated due to antineoplastic drugs. It increased the amounts of antioxidants (GSH, SOD, and CAT) and reduced the level of MDA in the heart, kidney, and liver tissues.


Oleuropein might be a beneficial agent against toxicity caused by the combination treatment of cyclophosphamide and epirubicin. Further studies should be performed to demonstrate the protective effects of oleuropein against antineoplastic induced-toxicity precisely.

Keywords: Oleuropein, epirubicin, cyclophosphamide, toxicity, oxidative stress


Breast tumors are the most frequently diagnosed cancer type and the leading cause of cancer-related deaths among women worldwide.1 Oxidative stress induced by chronic inflammation is an important determinant in the progression of cellular changes, which help enhance in the production of reactive oxygen species (ROS) and the proliferation of cells. Inflammation caused by cytokines, such as interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α), has been linked to the increased production of ROS and breast tumor formation.2,3

Chemotherapy is a frequently used method to treat breast cancer. Although some antineoplastic drugs can be used as a single agent for chemotherapy, two or three antineoplastic agents are generally administered together as a combination therapy regimen to enhance their effectiveness.4 For instance, cyclophosphamide is a frequently used alkylating agent in the treatment of various types of cancers.5 Epirubicin is an anthracycline used in the treatment of various tumor types. It inhibits DNA, RNA, and protein synthesis through the intercalation of DNA, the hindering of topoisomerase II activity, and the production of reactive oxygen radicals. Epirubicin and cyclophosphamide can be used as single agents in breast cancer chemotherapy. Moreover, the combination treatment of epirubicin and cyclophosphamide is one of the frequently used chemotherapy regimens in the early and metastatic stages of breast tumor development.6,7 Nonetheless, these agents are associated with an increased risk of hepatotoxicity, nephrotoxicity, cardiotoxicity, and hematologic toxicity in patients with breast cancer.8,9,10 These toxic effects may reduce the quality of life of patients and affect the success of their treatment. As such, novel remedies should be investigated to reduce the toxic effects on patients with breast cancer under the combination chemotherapy of epirubicin and cyclophosphamide; through such treatments, morbidity can be minimized, and patients’ quality of life can be improved.

Some plant-derived polyphenols have pharmacological effects, such as antiinflammatory, antioxidant, and antitumor properties. Therefore, studies have focused on the use of natural dietary antioxidants to alleviate the toxic effects of antineoplastic drugs.11 Oleuropein (3,4-dihydroxyphenylelenolic acid) is a non-toxic secoiridoid glycoside and the major polyphenolic compound in olive tree (Olea europaea L.) and olive oil. It is responsible for the bitter taste of the leaves and fruits of olive tree. Oleuropein and its bioactive derivatives, such as hydroxytyrosol, have antioxidant, cardioprotective, anticancer, antiinflammatory, neuroprotective, and hepatoprotective effects by modulating several mechanisms.12,13 In a novel study, oleuropein does not lead to a decrease in the efficacy of anthracycline-based chemotherapy in breast tumor-induced female BALB/c mice. On the contrary, oleuropein elicits a synergistic antitumoral effect with antineoplastic agents.14 Therefore, our study aimed to explore the effects of oleuropein on epirubicin- and cyclophosphamide-induced toxicity in rats.


The effects of oleuropein on cyclophosphamide- and epirubicin-induced toxicity in rats were examined using different methods. Whole blood samples were examined through hemogram tests, and serum samples were subjected to biochemical analysis. IL-6 and TNF-α in the serum samples were analyzed through enzyme-linked immunosorbent assay (ELISA). Subsequently, a comet assay was performed by using lymphocyte DNA. The levels of oxidant [i.e., malondialdehyde (MDA)] and antioxidants [i.e., catalase (CAT), glutathione (GSH), and superoxide dismutase (SOD)] were measured in the heart, kidney, and liver tissues.


The following commercially available chemicals were used in this study: Oleuropein (high-performance liquid chromatography grade ≥98%; Santa Cruz Biotechnology®, Santa Cruz Biotechnology Inc., California, USA); epirubicin (Pirucin®, Saba Ilaç AŞ, Istanbul, Turkey); cyclophosphamide (Endoxan®, Baxter Oncology GmbH, Frankfurt, Germany); ketamine (Ketalar®, Pfizer Inc., Istanbul, Turkey); and xylazine (Rompun®, Bayer LLC, Istanbul, Turkey). All the other chemicals used in this study were obtained from Sigma-Aldrich Inc. (Missouri, USA).


Fifty-six healthy 3-month-old female Sprague-Dawley rats weighing 220 g±20 g were obtained from the Aydın Adnan Menderes University Experimental Animals Research and Application Center (Aydın, Turkey). They were kept inside polycarbonate cages in an air-conditioned room (23°C±2°C) and relative humidity of 50-55% with 12 h/12 h light/dark cycle. Standard rat food and water were provided ad libitum. The rats were held in the room and acclimatized to the laboratory environment for a week before the drug administration phase. This study was approved by the Local Ethics Committee for Experiments on Animals of Aydın Adnan Menderes University (Ethics Committee permission no: 64583101/2017/117) and performed in accordance with the ethical rules of the Helsinki Declaration.

Experimental design

The rats were separated into seven equal groups (n=8 in each group): Group I (control), 1 mL of saline administered once a week for four cycles; group II (epirubicin, “E”), 2.5 mg/kg/week epirubicin administered for four cycles; group III (cyclophosphamide “C”), 16 mg/kg/week cyclophosphamide administered for four cycles; group IV (epirubicin + cyclophosphamide, “EC”), 2.5 mg/kg/week epirubicin + 16 mg/kg/week cyclophosphamide administered for four cycles; group V (epirubicin + oleuropein, “EO”), 2.5 mg/kg/week epirubicin + 150 mg/kg/week oleuropein administered for four cycles; group VI (cyclophosphamide + oleuropein, “CO”), 16 mg/kg/week cyclophosphamide + 150 mg/kg/week oleuropein administered for four cycles; and group VII (epirubicin + cyclophosphamide + oleuropein, “ECO”), 2.5 mg/kg/week epirubicin + 16 mg/kg/week cyclophosphamide + 150 mg/kg/week oleuropein administered for four cycles.

Epirubicin and cyclophosphamide were administered via intraperitoneal (i.p.) injection, and oleuropein was administered through oral gavage (p.o.). Epirubicin, cyclophosphamide, and oleuropein were freshly prepared in saline and administered at the same time of the day in every cycle. The doses of epirubicin and cyclophosphamide administered to the rats in the study were determined by converting the human doses, which were stated in the United States National Comprehensive Cancer Network Guidelines (available at https://www.nccn.org). Dose conversions between humans and rats were calculated as described in the United States Food and Drug Administration Guidelines (available at https://www.fda.gov). The dose of oleuropein used in the study was determined from previous studies.15,16,17,18,19

The rats were treated for 4 weeks and anesthetized with 50 mg/kg ketamine (i.p) and 5 mg/kg xylazine (i.p.) 1 week after the last treatment. Blood samples were taken by cardiac puncture for comet assay, ELISA, hemogram tests, and biochemical analysis. Then, the rats were sacrificed, and the heart, liver, and kidneys were taken for the analysis of oxidant/antioxidant parameters. The organs were removed immediately and kept frozen (-80°C) until they were analyzed.

Hemogram tests

Blood samples were taken into tubes containing ethylene diamine tetra acetic acid. The samples were analyzed within the first hour after they were received from the rats. Routine hematological parameters such as leukocyte (WBC), lymphocyte (LYM), monocyte (MONO), granulocyte (GRA), lymphocyte %, monocyte % (MONO %), granulocyte % (GRA %), hemoglobin (HGB), erythrocyte (RBC), hematocrit (HCT), mean cell volume, mean cell hemoglobin concentration (MCHc), MCH, erythrocyte distribution width concentration (RDWc), platelet (PLT), platelet count/the values of other cells % ratio (PCT), platelet distribution width (PDWc), and platelet/cell number ratio (MPV) were analyzed using an automated Diatron® Abacus Junior Vet (Diatron Medical Instruments Plc, Hungary) hematology analyzer.

Biochemical analysis

Plasma was separated through centrifugation from the whole blood and used to determine biochemical parameters in rat serum: Urea, uric acid, creatinine, aspartate aminotransferase (AST), creatinine kinase (CK), alanine aminotransferase (ALT), gamma glutamyl transferase (GGT), CK-myocardial band (CK-MB) isoenzyme 3, direct biluribin, and total bilirubin. Analysis was initially performed within the first hour after the blood samples were taken from the rats by using a Roche® Cobas c501 autoanalyzer (Roche Diagnostics, Switzerland) and Roche® commercial kits.

Serum IL-6 and TNF-α levels were determined using a commercially available Thermo Fisher® (Thermo Fisher Scientific Co., USA) ELISA kits in accordance with the manufacturer’s instructions.

Comet assay

The comet assay protocol was implemented as described in Singh et al.20 In brief, lymphocytes were placed to low-melting agarose (0.7% in phosphate-buffered saline) and placed in a lysis solution (2.5 M NaCl, 100 mM Na2EDTA, 10 mM Tris-HCl, pH 10, containing freshly added before use 1% Triton X-100 and 10% dimethyl sulfoxide) for 1 h at 4°C. Then, the samples were placed on slides.

The slides were placed on a horizontal Cleaver® gel electrophoresis tank connected to a Cleaver® Scientific CS 300 power supply (Cleaver Scientific Ltd., UK) and a Julabo® FL300 recirculating cooler (Julabo GmbH, Germany). The tank was filled with alkaline solution (1 mM Na2EDTA and 300 mM NaOH, pH 13) until the liquid level covered the samples. Then, the lid of the tank was closed and waited for 30 min at 4°C. Electrophoresis was continued at 4°C in the dark for 30 min at 25 V and 300 mA, and the samples were rinsed with 400 mM Tris buffer with pH 7.5 for 7 min to neutralize surplus alkali. The neutralized slides were kept in ethanol for 5 min and then dried at room temperature. The slides were stained with 70 µL (10 µg/mL) 4ʹ,6-diamidino-2-phenylindole dihydrochloride (DAPI) before microscopic examination.

Comets were analyzed after the specimens were stained at 400x magnification by using a Leica® fluorescence microscope (Leica Microsystems GmbH, Germany) equipped with a 50 W mercury lamp. Then, 100 cells were randomly counted for each slide. The extension of each comet was examined through Comet Assay® IV (Perceptive Instruments Ltd., UK) computerized image analysis. The damage ratio of each sample was expressed as “tail moment” and “% tail intensity,” as described in Collins et al.21

Measurement of oxidant/antioxidant parameters

In this procedure, 0.5 g of tissue samples was taken from each organ (kidneys, heart, and liver) for homogenization. Tissues were homogenized at 2.000 rpm for 1 min by using a Teflon-glass stirrer (IKA Overhead Stirrer; IKA-Werke GmbH & Co., KG, Germany) in a 10-fold volume of ice-cold 10% of 150 mM phosphate buffer (pH 7.4). The homogenate was centrifuged (Hettich Zentrifugen, Mikro 200 R, Germany) at 12.000 rpm for 10 min at 4°C. The supernatants, referred to as homogenate, were stored at -80°C (Glacier Ultralow Temperature Freezer, Japan) until CAT, SOD, GSH, and MDA levels were examined. All oxidant/antioxidant levels were analyzed through spectrophotometric measurements by using a Shimadzu® ultraviolet-1601 (Shimadzu Scientific Instruments, Japan) spectrophotometer. Protein concentrations were measured in accordance with the Biuret method by using a spectrophotometer and commercially available kits (Archem Diagnostic Ind., Ltd., Turkey). The results were expressed in milligrams per milliliter of protein.

MDA levels were determined as described previously by Ohkawa et al.22 The MDA concentration (nmol/mg of tissue protein) was calculated on the basis of absorbance: Absorbance coefficient (e)=1.56×105/M/cm. CAT activity (k/mg tissue protein) was evaluated as described by Aebi.23 SOD activity (U/mg of tissue protein) was determined as described by Sun et al.24 GSH levels (mg/g tissue protein) were spectrophotometrically determined at 412 nm by using the method described by Tietze.25

Statistical analysis

Data were statistically analyzed using SPSS 22.0. All the parameters were examined for the homogeneity of variance with Levene’s test and normal distribution with the Shapiro-Wilk test. The intra-group repeated measurements of rat weights were evaluated with the Friedman test. Because of the Shapiro-Wilk test results did not match normal distribution, data were compared between groups by using Kruskal-Wallis ANOVA. Post-hoc binary comparisons were performed using the Bonferroni-corrected Mann-Whitney U test. Differences were considered statistically significant if p<0.05. All data were expressed as mean ± standard deviation.


Body weight changes

In this study, the weight of the rats in the combination therapy (EC) group significantly decreased, whereas the weight of the rats in all the other groups significantly increased (p<0.05). Although the initial body weights did not vary among the groups, the weights of the control group significantly differed from those of the E, C, EC, and CO groups at the end of the study (p<0.05). The results of the body weight measurements of the rats from the day of the determination of the groups (day 0) until the day the study was terminated (day 35) are presented in Table 1.

Hemogram tests

MONO, GRA, MONO %, GRA %, RDWc, PCT, MPV, and PDWc did not significant vary (p>0.05), but these data were not shown to simplify the results in the table. The results of the hemogram tests and their statistical differences between the experimental groups are given in Table 2.

Most of the parameters generally deteriorated in all the groups administered with the antineoplastic drugs alone in the single and combination therapies. These results significantly differed from those observed in the control group (p<0.05). WBC, LYM, RBC, HGB, HCT, and PLT, which are related to neutropenia and bone marrow suppression, significantly varied between the groups treated with antineoplastic drugs alone (E, C, and EC) and the groups treated with oleuropein + antineoplastic drug (EO, CO, and ECO; p<0.05). However, the hemogram test results of the groups treated with oleuropein + antineoplastic drug were generally closer to those of the healthy control group than to the groups treated with antineoplastic drugs alone.

Biochemical analysis

The biochemical parameters and their significant differences between the experimental groups are given in Table 3. Urea, GGT, direct bilirubin, and CK parameters did not significantly differ among the groups (p>0.05). Therefore, these data were not shown to simplify the results in the table.

AST and ALT, which are important biochemical indicators of liver damage, significantly differed (p<0.05) between the groups treated with antineoplastic drugs alone (E, C, and EC) and the groups treated with oleuropein + antineoplastic drugs (EO, CO, and ECO). Oleuropein ameliorated these parameters and decreased them to levels similar to those of the healthy control group (p>0.05). These biochemical parameters indicated that oleuropein helped alleviate liver damage. Although oleuropein was beneficial to some parameters, which are related to heart and kidney damages, no considerable effects were observed in most of the other parameters.

The administered antineoplastic agents as single or in combination significantly increased the serum IL-6 and TNF-a levels compared with those in the control group (p<0.05). These levels decreased and were similar to those of the healthy control group in all oleuropein-treated groups (p>0.05). The results of the ELISA tests are presented in Figure 1.

Comet assay

The tail moment and % tail intensity results of the comet assay are presented in Figure 2. The degree of DNA damage was considerably higher in the experimental groups than in the control group. However, the considerable DNA damage caused by EC significantly decreased in the ECO (p<0.001). The fluorescent microscope images of some DNA samples are shown in Figure 3.

Oxidant/antioxidant levels

The levels of MDA and GSH and the measurement of SOD and CAT activities in the liver, heart, and kidneys of the experimental groups are given in Table 4. The SOD activity was considerably higher in the ECO group than in the EC group (p<0.05). Moreover, the SOD activity in the heart did not significantly vary between the control and ECO groups (p>0.05).

Oleuropein showed protective effects on the heart tissues by increasing GSH levels and decreasing MDA levels. The increased MDA levels were due to the decrease in the antineoplastic drugs (p<0.05) in the oleuropein-treated groups, but these levels decreased to levels similar to those of the control group (p>0.05). Although the GSH levels in the groups treated with antineoplastic drugs alone (E, C, and EC) decreased, these levels increased remarkably (p<0.05) in the oleuropein-treated groups (EO, CO, and ECO) and reached levels similar to those of the control group (p>0.05).

The SOD and CAT activities in the kidneys of the rats treated with the antineoplastic drugs alone (E, C, and EC) significantly decreased compared with those of the rats in the control group (p<0.05). The administration of oleuropein led to an amelioration of SOD and CAT activities in the kidneys, thereby reaching levels similar to those of the healthy control group.

The MDA levels in the kidneys were lower in the ECO group than in the EC group (p<0.05). However, the differences between the healthy control group and the ECO group were not significant (p>0.05). This result indicated that oleuropein could significantly lower the elevated MDA level, which is a marker of oxidative damage. Therefore, it showed cell-protective effects on the kidneys.

The results of the GSH, SOD, and CAT analysis in the liver tissues were generally compatible with one another. In most of the oleuropein-treated groups, the antioxidant parameters were similar (p>0.05) to those of the healthy control group and significantly higher (p<0.05) than those of the groups treated with antineoplastic drugs alone. Oleuropein increased the antioxidant capacity in the liver tissues to levels similar to those of the control group. By contrast, it decreased oxidative stress and degradation products. Unlike the SOD, GSH, and CAT levels in the cardiac and renal tissues, their levels in the liver were even higher in the oleuropein-treated groups than in the healthy control group.


In this study, the efficacy of oleuropein against the toxic effects of the combination therapy of epirubicin and cyclophosphamide, which are frequently used to treat breast tumors, was investigated using healthy female rats. This study was the first to demonstrate the effects of oleuropein against toxicity induced by a combination chemotherapy based on anthracycline and alkylating agents. Many parameters were investigated and important data were obtained in this study, but this study had limitations because it did not involve a histopathological examination of the tissues and an investigation of other serum cytokine levels, such as IL-8 and IL-1β, which are involved in inflammation.

Weight loss, vomiting, appetite loss, anorexia, and neutropenia are the important side effects of the combination therapy of epirubicin and cyclophosphamide.26 In our study, the initial and final body weights of the rats in the EC group significantly differed (p<0.05). Similar weight gain results were detected in the ECO group compared with those in the healthy control group (p>0.05). These results suggested that oleuropein might be a useful agent to prevent the weight loss of the rats in this combination chemotherapy group. However, a previous study showed that rats in oleuropein-treated groups that receive a fat-rich diet gain less weight than those in other groups (p<0.05).27 Another study has demonstrated that rats administered with oleuropein + bisphenol A (BPA) have less weight gains than those treated with BPA alone (p<0.05).28 Based on the outcomes of these studies, our conclusion was that oleuropein treatment likely had a regulatory role in various mechanisms by which an ideal weight could be maintained rather than inducing weight loss or weight gain.

A limited number of studies on the effects of oleuropein or other olive products (leaves, fruits, olive oil, and olive mill waste water extracts) have examined the hemogram parameters. In one study, oleuropein administered to rats with cisplatin-induced toxicity ameliorates the hemogram parameters that reach levels similar to those of the healthy control group.29 These consistent results indicated that oleuropein might have beneficial ameliorating effects on most of the hemogram parameters, which deteriorated because of epirubicin and cyclophosphamide toxicity.

The comet assay showed that oleuropein might be a beneficial agent that could reduce the oxidative DNA damage caused by antineoplastics. In literature search, no data were found in the effects of oleuropein in terms of a comet assay for epirubicin and cyclophosphamide toxicity. As such, studies have compared oleuropein-rich olive plant (leaf, fruit, and olive mill waste water extracts) and olive oil extracts between mouse and human peripheral mononuclear blood cells. These compounds significantly reduce oxidative damage levels and show DNA protective activities.30,31,32,33,34 In this regard, our results were consistent with previous findings.

The biochemical parameter results indicated that oleuropein could reduce the elevated AST and ALT levels, which are associated with liver damage. However, no significant differences were observed in most parameters. In a previous study, oleuropein administration considerably decreased the elevated levels of AST, ALT, urea, and creatinine in rats with BPA-induced toxicity.35 In another study, oleuropein significantly decreased the increased ALT and AST levels in a hepatic fibrosis mouse model and may be a pharmacologically useful agent in hepatic fibrosis.36

The increased risk of myocardial infarction is an important side effect of anthracycline-derived drugs.26 CK-MB is an important biochemical marker in monitoring the risk of myocardial infarction.37 In our study, conflicting results were obtained on the effects of oleuropein in the CK-MB parameter. These results might be related to the dose of oleuropein administered.16 Therefore, the pharmacological effects on this parameter might be observed through the administration of higher doses.

Proinflammatory cytokines, such as TNF-α and IL-6, are involved in the formation and progression of breast tumors.2,38 Most antineoplastic agents used in the treatment of breast tumors cause an increase in tissue and serum IL-6 and TNF-α levels.39,40,41 In another study, the effects of different doses of oleuropein (5, 10, and 20 mg/kg) are investigated against cisplatin-induced toxicity in mice. Cyclooxygenase-2, nuclear factor kappa B, and TNF-α levels in the kidneys decreased compared with the administered dose of oleuropein, which may help reduce cisplatin-induced toxicity in the kidneys.40 In another study on human synovial sarcoma cells (SW982), IL-6 and TNF-α levels increase as a result of the induction of IL-1β-mediated inflammation and significantly decrease after oleuropein administration compared with those in the control group.42 Several studies have investigated the effect of oleuropein against the toxicity induced by different agents in terms of serum and tissue cytokine levels in rats. They have shown that oleuropein can significantly reduce IL-6 and TNF-α levels in serum and tissues and elicit protective effects on organs. In other studies, oleuropein can decrease the elevated IL-6 and TNF-α levels induced by various compounds in direct proportion with doses.39,43,44,45 In this regard, previous findings were consistent with our results. Animal experiments have also revealed that oleuropein can decrease IL-6 and TNF-α levels in a wide dose range (10-2000 mg/kg/day). The selected dose in our study (150 mg/kg/week) also decreased the levels of these cytokines compared with that in the healthy control group. Similar results were obtained even at different doses in these studies. This situation might be related to the purity of oleuropein used in previous studies.

Oxidative stress plays a crucial role in the pathogenesis of the hepatotoxic, nephrotoxic, and cardiotoxic effects of epirubicin and cyclophosphamide.7,10 The protective effects of oleuropein have been attributed to several mechanisms, such as the reduction of nitrosative and oxidative stress and antiinflammatory and antioxidant activities.12,13,39 In our study, the reduced GSH, CAT, SOD, and MDA levels in the kidney, hepatic, and cardiac tissues were determined as indicators of oxidative stress and organ damage. Reduced GSH plays a major role in cellular defense against toxicity and ROS scavenging. SOD converts highly reactive superoxide anions to H2O2. Subsequently, CAT is the enzyme responsible for the conversion of H2O2 formed in cellular processes into molecular oxygen and water. Moreover, MDA is a lipid peroxidation product and indicator of oxidative damage caused by ROS generation.46,47,48

Our study showed that oleuropein could decrease the MDA level and increase the GSH level and SOD and CAT activities in the liver, heart, and kidneys of the rats compared with those of the rats treated with antineoplastic drugs alone. However, studies have yet to reveal the effects of oleuropein on endogenous oxidant/antioxidant parameters in the combination chemotherapy of epirubicin and cyclophosphamide. Therefore, our findings were compared with the results of studies on the effects of oleuropein against toxicity induced by cyclophosphamide, doxorubicin, and other chemical agents in rats. In these studies, oleuropein was administered in a wide dose range of between 5 and 2000 mg/kg/day. These studies reported that oleuropein decreased the amount of MDA and increased the amount of antioxidant markers (GSH, SOD, and CAT) in many organs, including the liver, heart, and kidneys, at a dose of ≥10 mg/kg/day.15,16,17,18,19,33,35,39 Our findings were parallel to previous results. Furthermore, studies have explored different doses and demonstrated that total antioxidant capacity also increases as the administration doses of oleuropein increase.16,18,39,48


Oleuropein is a promising compound with antiinflammatory and antioxidant effects against chemical toxicity. Its anti-tumoral properties and synergistic effects with some antineoplastics increase the importance of oleuropein, especially in patients with cancer. This study showed that oleuropein could reduce the levels of DNA damage and serum proinflammatory cytokines, such as TNF-α and IL-6. It ameliorated some of the deteriorated hemogram (WBC, LYM, and HGB) and biochemical parameters (AST, ALT, and total bilirubin) because of antineoplastic drugs. Oleuropein could also increase the amounts of antioxidant parameters (SOD, CAT, and GSH) in the tissues and caused a decrease in the levels of MDA, which is a cellular degradation product. These results suggested that oleuropein might have protective effects against the toxicity induced by the combination chemotherapy of epirubicin and cyclophosphamide. Further studies should be performed to demonstrate the protective effects of oleuropein against antineoplastic induced-toxicity precisely.

Conflict of interest: No conflict of interest was declared by the authors. The authors are solely responsible for the content and writing of this paper.

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