Pecan pericarp extract protects against carbon tetrachloride-induced liver injury through oxidative mechanism in rats

Abstract

The purpose of this study was to quantify the proanthocyanidin content of pecan (Carya illinoinensis) pericarp extract (PPE) and to assess its useful impacts against carbon tetrachloride (CCl₄)-induced hepatotoxicity. Rats were randomly divided into four groups: Group 1: received intraperitoneal injection of saline solution, Group 2: was injected with PPE (25 mg/kg body weight) for 10 consecutive days, Group 3: received CCl₄ (0.5 ml/kg, subcutaneous injection), Group 4: was coadministred with PPE + CCl₄. The CCl₄ was administered every 3 days during 10 days. Results revealed the presence of a high amount of total proanthocyanidins in the PPE (81.01 ± 0.21 mg TAE.g⁻¹DW). CCl₄ injection induced significant reductions in hepatic antioxidants but increased hepatic lipid peroxidation (LPO) as well as serum injury biomarkers. However, cotreatment with PPE significantly (P < 0.05) inverted CCl₄-induced increase in plasma alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and lactate dehydrogenase activities, respectively to 74%, 77%, 60%, and 82% compared with CCl₄ group. No significant toxic effects were observed following treatment with plant extract alone. PPE cotreatment also decreased significant (P < 0.05) the hepatic malondialdehyde formation (21%) and enhanced the liver catalase activity (107%) in CCl₄-intoxicated rats. The histopathological examination showed inflammatory infiltration and degenerative changes in the hepatic tissue following CCl₄ injection. The hepatoprotective activity of PPE against CCl₄ exposure was supported by the maintenance of structural integrity of liver histopathology. In conclusion, the current study illustrated that PPE pretreatment significantly improved all examined parameters, restored the hepatic architecture and successfully alleviates oxidative damage induced by CCl₄ intoxication.

Graphical Abstract

Graphical Abstract.

Practical applications

Pecan pericarp extract (PPE) was tested to prevent CCl₄-induced liver injury in male rats in an experiment conducted for 10 days. Biochemical and histological parameters were measured to evaluate both the toxic effect of CCl₄ and the protective potential effect of PPE. The results of this study revealed that PPE has a potential application as alternative natural antioxidant product for pharmaceutical, food, and cosmetic industries and can be used as a food supplement for preventing CCl₄-induced hepatotoxicity.

Introduction

The liver is the largest internal organ in the body. It is responsible for >500 different functions. The liver is liable to toxicity caused by many factors giving rise to an alteration in metabolic functions and severe health problems. One of these functions is detoxification and neutralizing toxins. It is the most metabolically active organ in the organism [1]. Hence, liver injury induce a heterogeneous metabolic disorders characterized by altered levels of various biochemical parameters [2].

Hepatotoxicity is mainly caused by inorganic compounds, organic agents, and synthetic drugs [1]. Carbon tetrachloride (CCl₄) is the most ancient chemical product usually used for experimental induction of liver injury in lab animals as it has a hepatotoxic effect [1, 3–4]. In hepatocytes, CCl₄ is converted to trichloromethyl free radicals (CCl₃^.) that can initiate cell damage through attacking cell macromolecules [4–5] resulting in oxidative stress, inflammation, and cellular necrosis [1–6] which leads to hepatocellular damages, such as fibrosis, cirrhosis, and atrophy [5].

At the molecular level, CCl₄ activates tumor necrosis factor alpha (TNF-α), nitric oxide (NO), caspase-2, and transforming growth factors alpha and beta [7]. Many researches demonstrate that the death of hepatocytes is due to oxidative stress which is a vital component in liver sickness [8–10]. Since experimental administration of CCl₄ drastically decreased cytochrome P450 (CY2E1, CYP2B, CYP3A2, CYP2C11, and CYP1A2 mRNA) and protein expressions, CYP isoenzymes others than CYP2E1 may be involved in catalyzing CCl₄ [11]. Because most of the CCl₄ taken up by the body will be eliminated by expiration through the lungs, a therapy of forced ventilation achieved as CO₂-induced hyperventilation had been developed by Teschke [12] in 16 patients with acute intoxications by CCl₄, to accelerate toxin removal via the lungs. Additional efforts are being made to reduce the microsomal production of toxic metabolites derived from CCl₄ by seeking appropriate inhibitors of microsomal drug metabolizing enzymes in addition to the cimetidine currently used for this purpose. The liver enhancement effect is probably due to an inhibitory effect of CYP isoenzymes including CYP2E1 due to substrate competition of cimetidine versus CCl₄ [12].

Accordingly, the need for consistent hepatoprotective agents is critical to combat liver problems. Therefore, research is paying attention on medicinal plants which are used in the practices and the development of newer agents from phytoconstituents which are more potential and effective with minimal side effects than the synthetic pharmaceutical compounds. A few herbal plants have been recorded to prevent and heal hepatic diseases. Hence, these plants have high contents of antioxidant substances that protect the liver by inhibiting free radicals production [13]. According to the studies of Wu et al. [14], the review of over 100 commonly consumed foods shows that pecan almonds (Carya illinoinensis) have the highest total phenols and antioxidant capacity of the nut group. Another study done on 98 common foods indicated that pecans had the second richness in proanthocyanidins [15]. Recent studies have also reported that pecan kernels are rich sources of polyphenols the most abundant are proanthocyanidins, although they also contain high quantities of phenolic acids [16–18]. Total proanthocyanidins or condensed tannins can be quantified by several colorimetric assays. Proanthocyanidins are monomers, oligomers, and polymers of catechin and other flavan-3-ols [16]. Kernels acetone extracts, showed proanthocyanidin contents from 2030 to 3950 mg as catechin equivalent (CE) per 100 g of whole kernel [16]. In by-products, nutshells contain the highest amounts of total polyphenols, around 10 times more than kernels, with values up to 59.1 g GAE per 100 g of nutshell which is 59.1% [19]. Around five times more total flavonoids have been reported in pecan nutshells (1.6–3.6 g CE/100 g of nutshell) compared to pecan kernel (345 and 640 mg CE/100 g of whole kernel [16]. Among flavonoids, monomeric flavan-3-ols catechin, epicatechin (procyanidins), epigallocatechin (prodelphinidins), and epicatechingallate are the most abundant compounds. Procyanidins are more abundant than prodelphinidins and most are B-type procyanidins. Flavonols, mainly quercetin and its derivatives (rutin and quercitrin), are the second largest group of flavonoids found in nutshell, and small amounts of flavones (hesperetin) and flavanones (naringenin) have also been reported [20]. Vázquez-Flores et al. [20] determined the presence of three different proanthocyanidin fractions with mean degree of polymerization in nutshell, after an extraction with 80% acetone, followed by an isolation through preparative Sephadex LH-20 column.

Pecan kernel extracts have also shown various in vitro effects, including antiproliferative [21], cell-protective [16], inhibition of digestive enzymes [20], and anti-inflammatory [22] via inhibition of ROS production. Biological activities of pecan by-products have been tested in vitro and in animal studies and most works have used an aqueous nutshell extract. Hawary et al. [23] determined that the acute LD50 was higher than 5 g/kg for both nutshell and leave methanolic extracts. In subacute studies, Porto et al. [24] determined that no toxic effect was observed on Wistar rats summited to doses of 100 mg of the aqueous extract of pecan shells/kg for 28 days.

The most important bioactivities are antimicrobial [17], antidiabetic [23], hepatopoprotective action [25] and protection against different forms of oxidative stress [26]. This is in part due to the remarkable content of condensed and hydrolysable tannins as well as polymeric polyphenols. The whole pecan diet also upregulated the expression of apolipoprotein B and LDL-receptor in the liver, and increased the activity of antioxidant enzymes such as catalase, glutathione-S-transferase and glutathione peroxidase, and decreased the oxidative stress in the liver of rats fed high-fat diets [27]. It inhibits free radical formation which induces LPO in hepatic cells [25]; thus, it is utilized as a tool for assessment of new hepatoprotective therapies [25].

There are also very few researches that attempt to identify, characterize and study condensed tannins in pecan by-products. However, the information related to the potential effects of pecan pericarp extract (PPE) as hepatoprotective agents against CCl₄-induced liver damage in rats is limited and little is known on its mechanism of action. Therefore, we aimed in this work to quantify the proanthocyanidins content of PPE and further to evaluate its hepatoprotective potential against CCl₄ intoxication. The liver prevention has been estimated by the levels of various biochemical biomarkers [alanine aminotransferase (ALAT), aspartate aminotransferase (ASAT), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH)] and histological analysis, whereas the antioxidant capacity was evaluated through different parameters, including MDA level and CAT activity.

Materials and Methods

Collection of plant material

The pecan samples (Carya illinoinensis) were collected from local place in Bizerta area (Tunisia), identified and authentified by Dr Mohamed Hédi El Aouni, Botanical professor in Faculty of Sciences, Bizerta (Tunisia).

Preparation of solvent extraction

Plant material was shed dried at room temperature and transformed into powder. About 5 g of powder was extracted with a mixture of 100 ml acetone: water (9:1 ratio) and was placed in a shaker for 24 h. Acetone–water mixtures are often used to isolate midrange molecular weight proanthocyanidins [28]. The solvent was completely removed by rotary vacuum evaporator. After elimination of acetone, 100 ml of ethyl acetate were added to the concentrated solution. Only amounts of ethyl acetate-soluble materials (low molecular weight oligomers up to tetraflavonoid level) where coextracted with the crude tannin polymer [28]. The mixture was then introduced into a separating funnel. The organic phase was separated from the aqueous phase which was concentrated and evaporated on a rotary evaporator under pressure and temperature below 30°C.The sample material was stored in an airtight container at 4°C and used for further experimental analysis.

Total proanthocyanidin content of pecan

The total proanthocyanidin content of samples (pericarp, fruit, leaf, and stem) was determined according to the method of Scalbert et al. [29] with slight modification. About 2.5 ml of 1% vanillin solution in methanol (MeOH), and 2.5 ml of 3.6 N H2SO4 in MeOH were added to the MeOH-diluted sample. Then the mixture was allowed to stand for 20 min at 30°C. The absorbance was measured at 500 nm. Total proanthocyanidin content of samples was expressed as milligram tannic acid equivalent per gram of dry weight plant (mg TAE.g⁻¹DW).

Experimental animals

Wistar male rats weighing 100–120 g were used. The rats were housed under controlled conditions of temperature (22°C), with 12 h light/dark cycle. They were maintained on a standard diet and water was available ad libitum.

Animals were cared for in compliance with the Tunisian code of practice for the Care and Use of Animals for Scientific Purposes. The experimental protocols were approved by the Faculty Ethics Committee (Faculty of Sciences, Bizerte, Tunisia).

Experimental pattern

The animals were randomly divided into four groups of eight animals each. Group 1 served as control and received equal daily intraperitoneal injection (i.p.) volumes of vehicle (corn oil) during 10 days, Group 2 (E) was treated only with i.p. injection of Carya illinoinensis pericarp extract for 10 days at a concentration of 25 mg/kg body weight, Group 3 (CCl₄) received a subcutaneous injection (s.c.) of CCl₄ (0.5 ml/kg (1:1 mixed with corn oil) alone every 3 days during 10 days, and Group 4 (E + CCl₄) was preadministered with the pericarp extract (25 mg/kg, i.p.), for 10 days and injected with CCl₄ (0.5 ml/kg) every 3 days [30].

All the treatments were carried out each day in the morning under similar constant conditions and rats were fed and observed daily.

Animal sacrifice

After experimental period, rats were sacrificed by decapitation. Blood was collected and the plasma was separated by centrifugation at 3000 × g for 15 min at 4°C, aliquoted and frozen at −80°C until analysis. Liver was immediately dissected out, washed, weighed, and frozen at −80°C for various biochemical evaluations and tissue samples were fixed in 4% paraformaldehyde for histopathological studies.

Evaluation of biochemical liver profiles

The plasma was used for the analysis of hepatic biochemical profile; ASAT, ALAT, LDH, and ALP were estimated by method using diagnostic reagent kit manufactured by Biomaghreb, Tunisia. Tissue homogenates were used to analyze the biochemical parameters. Proteins were estimated by the method of Lowry et al. [31] using bovine serum albumin (Sigma, St. Louis, MO) as a standard. LPO was expected according to the method given by Buege and Aust [32]. About 400 mg of liver are homogenized in TBS-buffer and then centrifuged at 10,000 rpm for 15 min. Almost 500 μl of the supernatant are recovered and resuspended in TBS and TCA-BHT then recentrifuged at 1000 rpm for 10 min. The supernatant obtained is taken up in HCl and tris-TBA. The addition of tris-TBA makes it possible to evaluate the formation of the MDA-TBA complex. Optical density is determined at 530 nm after 10 min of incubation at 80°C. Catalase (CAT) activity was determined according to the method of Claiborne [33]. The liver is excised and homogenized in TBS-buffer. Samples are then diluted in phosphate buffer. The reaction is triggered by the addition of H₂O₂ and the CAT activity was measured by measuring the initial level of H₂O₂ disappear at 240 nm.

Liver histology

Serial sections of liver were prepared according to the method previously described [34]. Small pieces of liver were fixed overnight at room temperature by direct immersion in 4% paraformaldehyde in 0.1 M phosphate buffer, pH 7.4. The samples were dehydrated with ethanol and toluene and embedded in paraffin wax. Serial sections (5 mm thick) were mounted on gelatin-coated glass slides and stained with hematoxylin and eosin.

Statistical analysis

Statistical analysis was performed using one-way ANOVA followed by Tukey’s multiple comparison as the post hoc test. Data are expressed as mean ± standard errors of the mean and significance of difference between groups was accepted at P value < 0.05.

Results

Quantification of total proanthocyanidin content

According to Fig. 1, the pericarp of pecan is the most rich in total proanthocyanidins (81.01 ± 0.21 mg TAE.g⁻¹DW) compared with fruit, leaf, or stem. Thus, in the present study we used the PPE.

Figure 1.

Proanthocyanidins concentration in various organs of Carya illinoensis

Assessment of liver injury

Subcutaneous injection of CCl₄ alone in rats caused a significant (P < 0.05) elevation in serum levels of ASAT, ALAT, ALP, and LDH compared with control rats (Table 1). In contrast, pecan cotreatment greatly (P < 0.05) restituted to near control value the activity of these enzymes respectively to 74%, 77%, 60%, and 82% compared with CCl₄ group.

Table 1.

Effect of pecan pericarp extract on biochemical liver profile

Treatments	ASAT (U/L)	ALAT (U/L)	ALP (U/L)	LDH (U/L)
Control	101.60 ± 16.15§	75.68 ± 14.78§	62.95 ± 14.24§	147.34 ± 16.99§
E (25 mg/kg)	85.09 ± 13.46§	65.82 ± 14.87§	62.56 ± 18,63§	113.70 ± 14.99§
CCl4 (0.5 ml/kg)	384.01 ± 18.56*	275.97 ± 10.88*	152.28 ± 18.51*	730.259 ± 33.66*
E + CCl4	88.37 ± 14.23§	70.78 ± 16.88§	61.18 ± 20.97§	130.28 ± 12.29§

Values [means ± standard errors of the mean (SEM), n = 8] are significantly different (P < 0.05) in Tukey’s multiple comparison post hoc test. Assays were carried out in triplicate. ASAT, aspartate aminotransferase; ALAT, alanine aminotransferase; ALP, alkaline phosphatase; LDH, lactate dehydrogenase.

^* P < 0.05 compared to control group.

^§ P < 0.05 compared to CCl₄ group.

Liver lipoperoxidation

Data from Fig. 2 showed that hepatic MDA level was significantly (P < 0.05) increased in CCl₄-intoxicated rats to 84% compared with controls and this effect was completely reversed by pecan extract coadministration. PPE cotreatment also decreased significantly (P < 0.05) the hepatic MDA formation to 21% compared with CCl₄ group.

Figure 2.

Effect of pecan pericarp extract on liver MDA level; animals were pretreated during 10 days with pecan pericarp extract [(E)25 mg/kg b.w., i.p.] or vehicle (control), challenged with a s.c. injection of CCl₄ (0.5 ml/kg b.w.) or vehicle (control) every third day for 10 days. Values (means ± standard errors of the mean (SEM), n = 8) are significantly different (P < 0.05) in Tukey’s multiple comparison post hoc test; assays were carried out in triplicate; ^*: P < 0.05 compared to control group and §: P < 0.05 compared to CCl₄ group

Liver CAT activity

Fig. 3 illustrated that CCl₄ treatment significantly (P < 0.05) decreased hepatic CAT activity. Importantly, cotreatment with PPE extract significantly (P < 0.05) ameliorated the abnormal levels of this antioxidant enzyme toward control level. PPE enhanced the liver CAT activity to 107% compared with CCl₄-intoxicated animals. Pecan extract per se had no significant effect on the enzyme activity compared with controls animals.

Figure 3.

Effect of pecan pericarp extract on liver CAT activity; values [means ± standard errors of the mean (SEM), n = 8] are significantly different (P < 0.05) in Tukey’s multiple comparison post hoc test; assays were carried out in triplicate; ^*: P < 0.05 compared to control group and §: P < 0.05 compared to CCl₄ group

Liver histology analysis

Control rats showed normal liver architecture with a well-preserved cytoplasm, protruding nucleus, nucleolus, and manifested central veins (Fig. 4a). No histopathological changes are observed in liver of rats treated with pecan extract alone (Fig. 4b). In contrast, the liver of rats exposed to CCl₄ showed various histological changes including a mixture of macrovesicular and microvesicular steatosis (Fig. 4c), lymphomononuclear inflammatory infiltration (Fig. 4d), binucleated, ballooning degeneration and mitosis (Fig. 4e). However, coadministration of PPE alleviated notably the histopathological hepatic lesions induced by the toxic. The histological structure of liver sections showed more or less normal lobular structure with a low degree of steatosis and lymphocyte infiltration quite similar to the control (Fig. 4f).

Figure 4.

Effect of pecan pericarp extract on CCl₄-induced liver damage; photomicrography of hematoxylin–eosin-stained sections of normal rat liver (a: ×25); liver from rats treated with pecan extract (25 mg/kg b. w.) (b: ×25); liver from rats treated with CCl₄ (0.5 ml/kg b. w.) (c: ×100, d: ×400, e: ×400); liver of a rat pretreated with pecan extract (f: ×100); H, hepatocytes; N, nucleus; CLV, central vein; st, steatosis; In, inflammatory infiltration; binucleated (^*), ballooning degeneration () and mitosis ()

Discussion

The present investigation revealed that Carya illinoinensis possesses an important amount of total proanthocyanidin compounds. High contents were obtained from PPE (81.01 ± 0.31 mg TAE.g⁻¹DW). The quantity extracted are superior that those previously reported from fresh apple fruits (37.99 ± 0.44 mg TAE·g⁻¹FW) [35]. Using an ultraperformance liquid chromatography coupled with quadruple time-of-flight mass Jia et al. [36] showed that (+)-Catechinis the most abundant phenolic in immature pecan kernels and it may be converted to proanthocyanidins and oligomeric proanthocyanidins, which becomes the major phenolic compound as the maturation progresses. The phenolic constituents of pecans possess bioactive properties, which might be useful in ameliorating the status of certain chronic disease states [36]. The efficacy of any hepatoprotective agent is essentially dependent on its ability to reduce the harmful effects or to maintain the normal hepatic physiology.

CCl₄ may induce liver damages at a variable extent as evidenced by increased serum activities of ASAT, ALAT, and LDH [12]. The research presented in this paper demonstrates that the toxic effects of CCl₄ led to the significantly increased activity of the ASAT, ALAT, ALP, and LDH enzymes in comparison to the results of the control-untreated group, which is in accordance with the results obtained in other studies [25, 37–38], since these enzymes represent a specific indicators of the hepatocellular damage and necrosis [39, 40]. Molecular CCl₄ is not toxic, but it is accumulated in the liver and further biotransformed in microsomes via the cytochrome P450 (CYP), especially by its isoenzyme CYP2E1, to the reactive^. CCl₃ radical. The CCl₃ radical reacts with oxygen to create a more reactive CCl₃O₂ [12, 41, 42]. The toxic metabolites of CCl₄ led to an increase in the activity of biochemical markers of damage in the serum by inducing the process of LPO and by destroying polyunsaturated fatty acids and phospholipids. These processes increased the porosity of the membrane of the hepatocyte, mitochondria, and endoplasmic reticulum, and released the enzymes from the intracellular space into the extracellular space and systemic circulation [43, 44]. Therefore, in our study, the cotreatment with PPE counteracted almost all CCl₄ deleterious plasma enzyme biomarkers, which indicates that the proanthocyanins from the PPE are noticeable in protecting the hepatocytes’ membranes and organelles from the toxic effects of CCl₄. Our results are in accordance with previous studies that make obvious the liver protective effects of proanthocyanidins compounds [45, 46].

CCl₄ toxic metabolites are tied to the lipids, and they remove the hydrogen atom from the unsaturated fat acids of the membrane, which induces the process of chain LPO that damages the liver cells [47], resulting in MDA increase as a final product in the LPO process [43]. The current study demonstrated a striking increase in MDA levels in CCl₄-treated rats compared with the control animals. Importantly, PPE cotreatment abolished CCl₄-induced MDA elevation.

According to the study accomplished by Popović et al. [17], the phenolic-proanthocyanins from the PPE led to a significant decrease of the LPO level in comparison to the CCl₄-treated animals, which was accomplished by the termination of the chain process of the LPO, by adding the hydrogen atom from the hydroxyl (OH) groups of proanthocyanin compounds to the unstable and reactive lipid peroxyl radical, which passed into an unreactive and significantly more stable state by forming the lipid hydroperoxides-LOOH. Hawary et al. [23] reported in diabetic rats treated with 125 mg of nutshell methanolic extract/kg for 4 weeks, a decrease in serum glucose, glycated hemoglobin, oxidative stress markers, and lipid peroxidation (MDA), serum insulin remained unchanged, whereas glutathione and total antioxidant capacity increased.

During the phase of oxidative stress, the toxic CCl₄ metabolites lead to the depletion and dysfunction of the antioxidative defense capacity whereas simultaneously increasing the pro-oxidative markers (Xanthine-oxidase, NADPH oxidase, GSSG, and H₂O₂), which results in oxidative stress and liver cell damage [44]. Phytochemical analyze achieved by Jia et al. [36] proved that proanthocyanidins may be the main antioxidant constituents at the kernel early stages.

Our results confirmed that CCl₄ caused a significantly decreased level of CAT activity in comparison to the untreated group. Pretreatment with PPE significantly inverted the level of this antioxidant enzyme. Similar findings are repoted for the water nutshell extract which is able to revert ethanol-induced liver damage by inducing a reduction on hepatic tissue oxidative damage and an increase of antioxidant enzymes activity and plasma antioxidant capacity according. This effect was attributed to the high content of phenolic compounds in the extract [25–48].

In addition to its significant contribution to the process of LPO, the CCl₃ radical has a crucial role in the induction of the inflammation by activating the Kupffer cells in the liver. The activated Kupffer cells then produce and release a variety of inflammatory mediators (TNF-α, IL-6, IL-1β, PGE2, ROS, and eicosanoids), thereby reinforcing the damage of the parenchymatous liver cells [49]. TNF-α represents a proinflammatory cytokine, which has a major role in the pathogenesis of various acute and chronic liver disorders. During the inflammatory phase of the liver damage caused by the CCl₄, TNF-α contributed to the worsening of the damages provoked by the oxidative stress and inflammation [50]. Furthermore, the biotransformation of CCl₄ produces numerous active metabolites, which bind to DNA and RNA, resulting in a reduction in the protein synthesis and damaging the liver structure [39]. In our study, histological examination of liver sections of rats treated with CCl₄ showed lymphomononuclear inflammatory infiltration, steatosis, binucleated, and ballooning degeneration of the liver tissue in comparison to the results obtained from the control group.

ROS, released in the CCl₄ metabolism, can induce the activation of NF-ĸB pathway and cause a significant production of proinflammatory mediators (IL-1β, IL-6, TNF-α, iNOS, COX-2, CXCL1, and NGAL), remarkably contributing to the pathogenesis of the liver injury [51].

According to Cheng et al. [43], the accumulation of MDA in excessive amounts resulted of the incapacity of the endogenous antioxidant systems to conteract the production of toxic radicals, leading to progressive peroxidation which subsequently generates severe disturbances of calcium homeostasis and necrotic cell death [52, 53] and subsequent hepatic tissue damage [1]. In contrast, cotreatment with PPE protected against CCl₄ damage to a considerable extent as evident from the preservation of normal hepatic architecture and absence of steatosis and inflammatory signs. This action may be related to the ability of PPE to maintain the structural integrity of hepatocytic cell membrane and/or regeneration of damaged liver cells. Zhong et al. [54], to explain hepatoprotective effect of polyphenols from Camellia sinenesis against cholestasis-induced liver fibrosis in rats, proposed that proanthocyanidin compounds prevented the activation of Kupffer cells by decreasing formation of inflammatory and fibrogenic mediators. The same mechanism may be proposed to explain the hepatoprotective action of proanthocyanidin compounds from Carya illinoinensis L. Accordingly, the ability of several phenolic compounds structures in pecan, particularly flavan-3-ols, anthocyanidins, proanthocyanidins (mono, dim, oligo, and polymeric), phenolic acid, and their sugar containing glycosides, to interact with other molecules in living organisms confers their beneficial properties following the synthesis carried out by Parrilla et al. [48].

Conclusion

The results obtained in this research confirm the fact that the Carya illinoinensis pericarp extract evidently decreased the subacute hepatotoxic effects caused by CCl₄. The protective properties are based on the decrease in the biochemical biomarkers of liver damage, the reduction of LPO level, the restriction of liver architecture alteration and the induction of hepatic antioxidant enzyme. This study can be considered as a significant contribution to the possible preventive use of proanthocyanins from the extract of pecan pericarp as nutraceuticals in reducing the hepatotoxic effects of drugs.

Authors’ contributions

H.D. contributed substantially to the completion of acquisition of analysis of the data; M.S. has been involved in the conception of the research and in drafting the manuscript; B.L.H. made and interpreted the histological sections; B.R.K. contributed in the phytochemical and oxidative stress analyses; S.M. contributed to the general supervision of the research; T.O. contributed to the interpretation of the data and the statistical analysis. All authors have read and approved the final manuscript.