Insights into the phytochemical composition and antioxidant potential of the Tunisian Ceratonia siliqua L.

Carob (Ceratonia siliqua L.) corresponds to an evergreen leguminous tree (Fabaceae family). The high phenolic content of numerous parts of carob has been deeply associated with several nutritional and functional benefits. The aim of this study was to investigate the physicochemical properties of ground carob pods and seeds, the effect of different extraction procedures as well a comprehensive phytochemical characterization of hydro-methanolic extracts (80/20 v/v) of pods and seeds by HPLC-DAD ESI-Q-TOF-MS/MS. Additionally, their antioxidant activity was evaluated using in vitro assays. The results showed thatthe dry matter (DM) values were 88.09% for pods and 89.10% for seeds, protein contents were 0.41 g/100 g DM for pods and 0.88 g/100 g DM for seedsand total sugars contents were 0.35 g/100 DM for pods and 26.70 g/100 g DM for seeds. Furthermore, the oil holding capacities (OHC) were 10.43 g/g for pods and 7.53 g/g for seeds, while the water holding capacities were 8.46 g/g for pods and 2.59 g/g for seeds.The hydro-methanolic extracts of both pods and seeds showed the presence of 53 secondary bioactive metabolites belonging to various classes(flavonoids, phenolic acids, tannins and non-phenolic compounds). The antioxidant activities were evidenced in DPPH (22.24 mg/ml for pods and 26.37 mg/ml for seeds), ABTS (198.50 mmol Eq Trolox/100 g for pods and 201.04 mmol Eq Trolox/100 g for seeds) and FRAP (0.39 mmol Eq Trolox/100 g for pods and 0.53 mmol Eq Trolox/100 g for seeds).Moreover,high significant (p ≤ 0.01) correlation coefficients were found between the antioxidant activity estimated by the DPPH method and total phenols (r = 0.943), orthodiphenols (r = 0.996), flavonoids (r = 0.880) and flavonols (r = 0.982). Nevertheless, lower correlations were detected with ABTS and FRAP methods.These results demonstrated that carob parts displayed an interesting potential that can be of interest for further valorizations as a natural antioxidant with multiple applications, namely functional food ingredients or prevention of many health problems.

Carob (Ceratonia siliqua L.), Pharmacological and Phytochemical Activities of Neglected Legume of the Mediterranean Basin, as Functional Food.

Carob (Ceratonia siliqua L.) has been widely cultivated in different parts of the world, particularly in the Mediterranean region, and the tree belongs to the family Leguminosae. Several studies have indicated that carobs and their products can improve human health and help prevent different specific chronic diseases. Carob can considered as functional food due to its high content in dietary fibers, low-fat content, and high content of minerals. Its fruit is a pod containing 10%-20% seeds, and the pods consist of sugars, proteins, crude fibers, minerals, vitamins, polyphenols, vitamins, and lipids. In many countries in the Middle east, carob is mainly used to prepare as a traditional drink and some kinds of confectioneries. The powders can be utilized to prepare carob juice concentrate. The systematic review of documents from clinical trials and scientific societies dedicated to traditional medicine in China has been carried out. The goal of this review article is a survey of chemical compounds, and pharmaceutical benefits of carob, especially by considering traditional medicinal sciences. Moreover, clinical trials research promotes studies to highlight and focus on the scope of application of traditional medicinal science in the growing system of medicine.

Phenolic compounds profile of macerates of different edible parts of carob tree (Ceratonia siliqua L.) using UPLC-ESI-Q-TOF-MSE: Phytochemical screening and biological activities.

Locust bean pulp and gum extracts were prepared, and phytochemical tests based on color reactions and chromatographic analyzes were performed. A profile of seventy-six phenolic compounds was obtained by the ultra-high performance liquid chromatography with electrospray ionization and quadrupole time-of-flight mass spectrometry. The main groups of phenolic compounds identified in the both extracts of Ceratonia siliqua L., were flavonoids, tannins and phenolic acids. Moreover, carob pulp and gum extracts were tested for their antimicrobial activity using disk diffusion tests which showed sensitivity of the different strains to the analyzed extracts at a concentration of 100 mg/mL. Additionally, the antioxidant activity of Ceratonia siliqua L. extracts was assessed by the 2,2-diphenyl-1-picrylhydrazyl acid test, which confirmed stronger antioxidant properties in the case of the pulp extract. To sum up, carob pulp and gum extracts present promising alternatives to synthetic additives within the medicinal industry, serving as potential antioxidant agents and preservatives that combat bacterial contamination, thereby offering a more natural approach to enhancing product safety and efficacy.

Immature carob pods extract and its fractions prevent lipid metabolism disorders and lipoprotein-rich plasma oxidation in mice: A phytochemical and pharmacological study.

ETHNOPHARMACOLOGICAL RELEVANCE: In Morocco carob fruits are used traditionally to treat hypercholesterolemia, diabetes and related diseases. AIMS: This study was designed to evaluate the hypolipidemic activity of Ceratonia siliqua green pods extract and its fractions in Triton WR-1339 and high fat/cholesterol diet (HFCD) induced hyperlipidemia mice, as well as their ability to prevent lipoproteins oxidation in vitro. MATERIALS AND METHODS: High performance liquid chromatography (HPLC) analysis was used to determine the phenolic composition of the immature carob pods extract (HWCE). Antioxidant activities were evaluated using the DPPH radical scavenging test as well as MDA measurement in oxidized lipoprotein rich plasma. Plasma lipids, glucose and biliary total cholesterol, as well as lipids level in liver and feces, were analyzed. The acute oral toxicity was performed in mice single dosed with the HWCE at 2000 and 5000 mg/kg body weight. RESULTS: HPLC analysis shows that gallic acid is the main phenolic compound in the HWCE. The acute oral toxicity assessment revealed that the HWCE is not toxic (LD50 is greater than 5000 mg/kg body weight). In the acute hypolipidemic study, mice treated with the HWCE and its fractions exhibited a significant (P < 0.001) reduction in plasma total cholesterol (TC), triglycerides (TG) and low density lipoprotein-cholesterol (LDL-C) levels. Importantly, immature carob aqueous extract was more effective in lowering mice hypercholesterolemia than its fractions. Indeed, mice fed the HFCD for 12 weeks showed a significant raise in plasma TC, TG and LDL-C, as well as in hepatic and fecal TC and TG levels. The HWCE at 100 and 200 mg/kg body weight significantly (P < 0.001) reversed the plasmatic levels of these lipid parameters, increased plasma HDL-C level, reduced hepatic lipids accumulation, but increased cholesterol level in the bile and fecal lipids excretion. The HWCE decreased also the atherogenic index, the LDL-C/HDL-C ratio and plasma glucose level after 12 weeks' experiment. On the other hand, the HWCE was more effective in preventing mice lipoprotein-rich plasma oxidation than its fractions, with a concentration-dependent manner. CONCLUSION: C. siliqua green fruits extract could be effective in preventing atherosclerosis and related cardiovascular complications through the inhibition of lipoprotein oxidation and cholesterol clearance.

Bioprotection of Olive Trees Against Verticillium Wilt by Pomegranate and Carob Extracts.

Bioprotection using plant extracts is an environmentally friendly strategy in crop protection. Effective control of Verticillium wilt of olive (Olea europaea; VWO), caused by Verticillium dahliae, has proven challenging due to the ineffectiveness of chemicals, which makes it necessary to search for new control tools. Thus, the aim of this study was to evaluate the effect of pomegranate (Punica granatum) and carob (Ceratonia siliqua) extracts against VWO. Extracts derived from pomegranate peels and carob pods and leaves were obtained using ethanol, methanol, or ethyl acetate as solvents. A targeted analysis of their metabolite composition was performed using QTRAP Ultra High-Performance Liquid Chromatography with Mass Spectrometry (QTRAP UHPLC‒MS). Remarkably, gallic acid was detected in all extracts at a high concentration. The effect of the extracts on the mycelial growth and on the germination of conidia and microsclerotia of V. dahliae was evaluated by in vitro sensitivity tests at various doses: 0 (control), 3, 30, 300 and 3,000 mg of extract/liter. Extracts obtained with ethanol or methanol significantly reduced the viability of V. dahliae structures when applied at the highest dose, while those obtained with ethyl acetate were ineffective across all doses. The most effective extracts, as determined in vitro, were then evaluated against the disease in olive plants. Potted plants of cv. Picual were treated by spraying (foliar application) or irrigation (root application) of extracts at 3,000 mg of extract/liter, followed by inoculation with V. dahliae. The results indicated that foliar applications were ineffective, while root treatments with pomegranate peel or carob leaf extracts were more effective in reducing disease severity, regardless of solvent, compared to that of the untreated control.

Exploring Carob (Ceratonia siliqua L.): A Comprehensive Assessment of Its Characteristics, Ethnomedicinal Uses, Phytochemical Aspects, and Pharmacological Activities.

The carob tree (Ceratonia siliqua L.) is currently considered one of the most valuable fruit and forest trees in various fields and sectors of activity. It is a versatile plant, belonging to the Fabaceae family. It is widely used in traditional medicine to treat many diseases such as diabetes, hypertension, and gastrointestinal disorders, given that all its parts (leaves, flowers, pods, seeds, wood, bark, and roots) are useful and hold value in many areas. Its importance has increased significantly in recent years. Originating from the Middle East, it is recognized for its ecological and industrial significance. Previous studies conducted on Ceratonia siliqua L. have revealed the presence of several compounds, including polyphenols, flavonoids, carbohydrates, minerals, and proteins. The carob tree demonstrates antihypertensive, antidepressant, anti-obesity, and antihyperglycemic activities. This plant is known for its medicinal and therapeutic virtues. Moreover, it is particularly interesting to consider the pharmacological activities of the major phytochemical compounds present in the different extracts of this plant, such as phenolic acids, for example, coumaric and gallic acids, as well as flavonoids such as kaempferol and quercetin. Therefore, this review aims to analyze some aspects of this plant, especially the taxonomy, cytogeography, traditional uses, phytochemical constituents, and pharmacological activities of Ceratonia siliqua L., in addition to its biological properties.

Biological Activities of Ceratonia siliqua Pod and Seed Extracts: A Comparative Analysis of Two Cretan Cultivars.

Ceratonia siliqua L., commonly known as the carob tree, appears in most Mediterranean countries, often cultivated for the collection of its fruits to be used as food for humans and animals. This study was aimed at the phytochemical characterization of two common Cretan C. siliqua cultivars and the biological evaluation of deseeded pod and seed extracts regarding their putative use in cosmetics. Gas and liquid chromatographic techniques were used to assess their essential oil, fatty acid, and carbohydrate profiles. Cell-free assays, including free-radical scavenging; the inhibition of tyrosinase and collagenase; the blocking of advanced glycation end product (AGE) formation; along with assays in human skin fibroblast cultures, i.e., reactive oxygen species suppression, glutathione stimulation, and protection from oxidative stress and from ultraviolet (UVB) radiation, were also used. Extracts from both cultivars were found to possess antioxidant capacity, tyrosinase- and collagenase-inhibitory activities, an ability to block glucose-induced AGEs, and in certain cases, UVB absorbance and photoprotective activities. Seed extracts were in general more active, while the use of 30% aqueous methanol seemed to be more efficient than n-hexane for extraction. Serial partition of the most active extracts resulted in fractions with enriched biological activities. These properties make Cretan carob extracts and their fractions suitable candidates for use in cosmetics.

Ceratonia siliqua L. Pod Extract: From Phytochemical Characterization to Liposomal Formulation and Evaluation of Behaviour in Cells.

The formulation of plant extracts in phospholipid vesicles is a promising strategy to exploit their biological properties while solving problems related to poor solubility in water, high instability, and low skin permeation and retention time. In this study, Ceratonia siliqua ripe pods were used for the preparation of a hydro-ethanolic extract, which showed antioxidant properties owing to the presence of biologically active compounds identified by liquid chromatography-mass spectrometry (e.g., hydroxybenzoic acid and flavonoid derivatives). To improve the applicability of the extract in therapy, a topical formulation based on liposomes was explored. The vesicles were characterized by small size (around 100 nm), negative charge (-13 mV), and high entrapment efficiency (>90%). Furthermore, they displayed both spherical and elongated shapes, with oligolamellar structure. Their biocompatibility was demonstrated in cells, including erythrocytes and representative skin cell lines. The antioxidant activity of the extract was proved by the scavenging of free radicals, the reduction of ferric ions, and the protection of skin cells from oxidative damage.

Aqueous extract of Ceratonia siliqua L. leaves elicits antioxidant, anti-inflammatory, and AChE inhibiting effects in amyloid-ß42-induced cognitive deficit mice: Role of α7-nAChR in modulating Jak2/PI3K/Akt/GSK-3ß/ß-catenin cascade.

Alzheimer's disease (AD) is a multifactorial neurodegenerative disorder attributed to several etiological factors including cholinergic dysregulation, neuroinflammation, oxidative stress, ß-amyloidogenesis, and tauopathy. This demands the search for multitarget drugs, especially of natural sources owing to their pleiotropic activities and low adverse effects. The present study was conducted to investigate the cognitive-improving potential of Ceratonia siliqua L. (Cs) extract compared with donepezil, an acetylcholinesterase inhibitor, on AD-like pathological alterations induced by single intracerebroventricular amyloid-ß42 (Aß42) injection in mice. Aß42-injected mice were treated with Cs (100 mg/kg/day, po) with or without methyllycaconitine (MLA; 1 mg/kg/day, ip), an α7-nAChR antagonist. Aß42-injected animals demonstrated an elevation of hippocampal Aß42, p-Tau, and acetylcholinesterase. They also showed a decline in phosphorylated levels of Jak2, PI3K, Akt, and GSK-3ß, leading to induction of neuroinflammation and oxidative stress. Noteworthy, Cs improved the histopathological and behavioral variables in addition to mitigating AD hallmarks. It also exerted neuroprotection by reducing NF-κBp65 and TNF-α, while elevating Nrf2 and HO-1, along with stabilizing ß-catenin under the impact of Jak2/PI3K/Akt/GSK-3ß signaling. These beneficial effects of Cs were abrogated by MLA co-administration signifying the α7-nAChR involvement in Cs-mediated effects. Therefore, Cs can ameliorate Aß42-induced neurodegeneration by modulating Jak2/PI3K/Akt/GSK-3ß/ß-catenin axis in an α7-nAChR-dependent manner.

Evaluation of The Effects of Carob (Ceratonia siliqua L.) Fruits on the Puberty of Rats

Objective: This study was planned to determine the effects of carob use on puberty because of the observation of early puberty or pubertal variants due to the long-term use of carob in our clinic. Methods: Forty-eight Wistar albino rats, on postnatal day 21, were assigned into two groups female (n=24) and male (n=24). Groups were divided into four groups Control, and Carob-150, Carob-300, and Carob-600. Ceratonia siliqua L. extract was given to rats in a 0.5% carboxymethylcellulose (CMC) solution. CMC (0.5%) was given to the control, Ceratonia siliqua L. extract was given 150 mg/kg/day to the Carob-150, 300 mg/kg/day to the Carob-300, 600 mg/kg/day to the Carob-600 by oral gavage. The treatments were performed once daily until the first sign of puberty. Serum follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol, total testosterone, leptin, glutathione, glutathione peroxidase (GPx), and malondialdehyde were measured by commercial rat-specific ELISA kits. Testis, uterus and ovarian tissue were examined histologically. Results: The median time of preputial separation in male rats was 38th, 31st, 31st, and 31st days in the Control, Carob-150, Carob-300, and Carob-600 groups, respectively (p=0.004). The median day of vaginal opening day was the 39th, 31st, 34th, and 31st days in the Control, Carob-150, Carob-300, and Carob-600 groups, respectively (p=0.059). FSH, LH, testosterone (male), estradiol (female) and leptin levels of the groups were similar. However, GPx levels were higher in male and female animals given C. siliqua extract compared to the Control (male p=0.001 and female p=0.008). Testicular and ovarian tissues were concordant with the pubertal period in all groups. As the dose of Ceratonia siliqua extract increased, it induced spermatogenesis and spermiogenesis, causing abnormal changes, such as ondulation in the basement membrane, capillary dilatation, and increased congestion in males. In females, edema in the medulla gradually increased with increased dosage, and granulosa cell connections were separated in Carob-300 and Carob-600 groups. Conclusion: This study demonstrated that C. siliqua caused early puberty and increased spermiogenesis and folliculogenesis. Antioxidant mechanisms were impaired with increasing dose, possibly leading to tissue damage at high doses.

The Effect of Ceratonia siliqua Supplement on Bone Mineral Density in Ovariectomy-induced Osteoporosis in Rats.

AIM: This study aimed to investigate the effect of Ceratonia siliqua on bone mineral density (BMD) as a non-pharmaceutical alternative treatment for postmenopausal osteoporosis. MATERIALS AND METHODS: Thirty mature female Wistar rats were randomly separated into three groups of 10: Control, ovariectomized (OVX), and ovariectomized-plus-C. siliqua (OVX+CS). Total and proximal BMD were measured by dual-energy X-ray absorptiometry (DEXA) in all groups before ovariectomy, and at 3 and 6 months postoperatively. At the end of the study, the femurs were subjected to a three-point bending test. RESULTS: DEXA revealed no statistically significant difference in absolute values or percentage changes for total tibial BMD between OVX+CS and OVX groups throughout the study. In the proximal tibia, both absolute values and BMD percentage changes from baseline were higher in the OVX+CS group compared to the OVX group after 3 and 6 months of C. siliqua administration. Three-point bending test revealed a significantly higher thickness index in the OVX+CS group compared to the OVX group and a higher cross-sectional area index compared to the control group. CONCLUSION: Long-term administration of C. siliqua may be considered a non-pharmaceutical alternative treatment for postmenopausal osteoporosis. Further research is required to properly investigate the effects, and suitable treatment dose and schedule.

Evaluation of Different Extraction Methods on the Phenolic Profile and the Antioxidant Potential of Ceratonia siliqua L. Pods Extracts.

The present work was designed to investigate the effects of different extraction processes, namely ultrasonic-assisted, supercritical fluid, microwave-assisted and Soxhlet applied to carob pods. The total phenolic quantification and the antioxidant activity were assessed by the means of rapid in vitro spectrophotometric assays; the phenolic profile was identified using ultra-high performance liquid chromatography coupled to mass spectrometry. The results revealed that the phenolic compounds and the antioxidant capacity varied significantly with the nature of the extraction process. The content of total phenolic compounds ranged from 11.55 to 34.38 mg GAE/g DW; the content of total flavonoids varied from 3.50 to 10.53 mg QE/g DW, and the content of condensed tannins fluctuated from 3.30 to 6.55 mg CE/ g DW. All extracts performed differently on antioxidant activity when determined by the DPPH assay producing a dose-dependent response, with IC50 extended from 11.33 to 6.07 µg/mL. HPLC analysis enabled the identification of nine compounds. As a function of the studied extraction methods, the phenolic compound contents were positively correlated with antioxidant activity.

Comparison of the effect of Ceratonia siliqua L. fruit oral capsule and vitamin E on semen parameters in men with idiopathic infertility: a triple-blind randomized controlled clinical trial.

OBJECTIVES: To compare the effect of oral Ceratonia siliqua (Carob) and vitamin E on sperm parameters in idiopathic infertile men. METHODS: In this triple-blind randomized controlled trial study, fifty 20-45-year-old infertile men with idiopathic abnormal sperm parameters were selected among men referring to the infertility clinic of Al Zahra-Iran Hospital, and randomly allocated into two groups. The first group received Carob capsules, 500 mg, and the second group received vitamin E capsules three times a day for 90 days. The primary outcomes of this study were the sperm parameters (count, motility, and morphology). RESULTS: After the intervention, the mean of sperm count increased from 49.08 to 60.22 million/mL in the carob vs. it increased from 47.64 to 58.88 million/mL in the vitamin E group (adjusted mean difference (aMD)=1.124 million/ml, 95% CI: -12.892 to 15.141). The mean sperm motility increased from 45.90 to 52.90% in the carob group but decreased from 55.23 to 38.10% in the vitamin E group (aMD=17.22%, 95% CI: 8.53 to 25.92). The mean sperms with normal morphology increased from 11.52 to 67.05% in the carob, and from 10.20 to 77.47% in the vitamin E group (aMD=-10.88%, 95% CI: -22.30 to 0.536). There was no serious side effect during the treatment course. CONCLUSIONS: Taking 1,500 mg oral dose of Carob capsule per day for 90 days improved sperm motility but had no significant effect for improving morphology and count comparing to vitamin E capsule.

Effect of Ceratonia siliqua L. extract on DNA Fragmentation of Sperm in Adult Male Mice Treated with Cyclophosphamide.

This study was conducted to determine the effects of Ceratonia siliqua L. (CS) extract on sperm parameters and DNA damage in adult male mice treated with cyclophosphamide (CP). Based on an initial dose response experiment on Ceratonia siliqua L. extract, five treatment groups were set up: control, sham (normal saline: 0.2 ml per day, IP), CP (15 mg kg-1 per week; IP), Ceratonia siliqua L. (100mg l-1 per day; IP), and group of Ceratonia siliqua L. along with CP for 35 days. After euthanizing the animals, sperms from caudal part of epididymis were collected, and their parameters, Malone Di-Aldehyde (MDA) level, and DNA fragmentation were analyzed. In the mice exposed to cyclophosphamide, reduction in the sperm count and viability and increase in the abnormal sperm and MDA levels were detected (p < .05). In addition, an increase in sperms with damaged DNA was detected in CP group, while the use of Ceratonia siliqua L. Extract significantly recovered these disturbances in the treatment group (p < .05). This study suggested the competence of Ceratonia siliqua L. extract in the improvement of sperm parameters and DNA fragmentation in animals treated with CP.

Ceratonia siliqua (Carob) extract improved in vitro development of vitrified-warmed mouse germinal vesicle oocytes: assessment of possible mechanism.

Oocyte banking is a vital step for safekeeping and spreading genetic resources of animals. It is also used for fertility preservation of human. Oocyte vitrification is closely related to the lower developmental competence which includes the cryo-injury arisen during vitrification. The aim of the present study was to evaluate the maturation, embryonic development and production of reactive oxygen species (ROS) of mice oocytes following the supplementation vitrification media with different concentrations of Ceratonia siliqua (carob) extracts. In this experimental study, germinal vesicle oocytes collected from 8 to 10 week-old female NMRI mice (30-40 gr) were randomly divided into six groups of vitrification media supplemented with 0 (control), 5, 10, 20, 30 and 50 µg/ml C. siliqua. After thawing, oocytes were put in an in vitro maturation medium (IVM) (α-MEM: Alpha Minimum Essential Medium). 3-4 and 24 h (hr) later, the oocyte nuclear maturity was checked. Standard in vitro fertilization was performed on the matured oocytes (MII), and embryonic development was followed. Extra- and intra-cellular ROS was measured in IVM medium after 24 h of oocyte incubation. The addition of 20 and 30 µg/ml C. siliqua extract to vitrification media improved normal morphology of warmed germinal vesicle (GV) oocytes, rate of germinal vesicle break down (GVBD), and metaphase 2 (MII) oocyte formation significantly (p < 0.05). Fertilization rate, (embryonic development to 2 cells stage, 4-8 cells stage, and > 8 cells stage increased in the 30 µg/ml C. siliqua group significantly (p < 0.05). Furthermore, supplementation of 30 µg/ml C. siliqua in vitrification media significantly decreased extra- and intra-cellular of ROS as well as embryonic fragmentation (p < 0.05). In conclusion, supplementation of GV oocyte vitrification media with carob extract improved maturation, fertilization, and embryonic development rate and decreased extra- and intra-cellular ROS levels.

Enhanced retinoid response by a combination of the vitamin A ester retinyl propionate with niacinamide and a flavonoid containing Ceratonia siliqua extract in retinoid responsive in vitro models.

OBJECTIVES: Retinoids have been used for decades as efficacious topical agents to treat photoaged skin. The purpose of our present research is to evaluate whether the activity of the vitamin A ester retinyl propionate (RP) can be enhanced by niacinamide (Nam) and a flavonoid containing Ceratonia siliqua (CS) fruit extract in retinoid responsive in vitro models. METHODS: Retinyl propionate was tested alone and in combination with Nam and CS in an RARα reporter cell line for promoter activation and compared to trans-retinoic acid (tRA) activation. These treatments were also tested in keratinocytes for gene expression profiling by qPCR using a panel of 40 retinoid responsive genes. RESULTS: tRA or RP elicited RARα reporter activation in a dose-dependent manner. The combination of 0.5 µM or 2 µM RP with 10 mM Nam had a 56% and 95% signal increase compared to RP, respectively. The addition of 1% CS to 0.5 µM or 2 µM RP with 10 mM Nam elicited a further increase of 114% and 156%, respectively, over RP and Nam combinations. All retinoids elicited an increase in expression of 40 retinoid sensitive genes over control levels. Of the 40 genes, 27 were enhanced by either 0.1 µM RP or 0.5 µM RP with 10 mM Nam and 1% CS. Nam or CS had very modest activity in both models. CONCLUSION: The combination of RP with Nam and CS showed a higher retinoid response than RP in two separate retinoid responsive in vitro models. We hypothesize Nam and CS enhances RP activity by modulating metabolism to tRA via increasing NAD+ pools and inhibiting reduction of retinal (RAL) back to retinol, respectively. The findings provide evidence that this combination may have enhanced efficacy for treating the appearance of photoaged skin.

OBJECTIFS: Les rétinoïdes sont utilisés depuis des décennies comme agents topiques efficaces pour traiter la peau photo-âgée. Le but de notre recherche actuelle est d'évaluer si l'activité du propionate rétinyl ester de vitamine A (RP) peut être augmentée par le niacinamide (Nam) et un flavonoïde contenant un extrait de fruit de Ceratonia Siliqua (CS) dans les modèles in vitro sensibles aux rétinoïdes. MÉTHODES: RP a été testé seul et en combinaison avec Nam et CS dans une ligne de cellule rapporteur de RARα pour l'activation du promoteur et par rapport à l'activation de l'acide transrétinoïque(tRA). Ces traitements ont également été testés dans les kératinocytes pour le profilage d'expression génique par qPCR à l'aide d'un panel de 40 gènes rétinoïdes sensibles. RÉSULTATS: tRA ou RP ont provoqué l'activation du promoteur RARα d'une manière dépendante de la dose. La combinaison de 0,5 µM ou 2 µM de RP avec 10 mM de Nam a permis une augmentation respectivement de 56% et 95% du signal par rapport à RP. L'ajout de 1 % de CS à 0,5 µM ou 2 µM de RP avec 10 mM de Nam a permis une nouvelle augmentation de 114 % et 156 %, respectivement, qu'avec la combinaison RP et Nam. Tous les rétinoïdes ont provoqué une augmentation de l'expression de 40 gènes sensibles aux rétinoïdes sur les niveaux de contrôle. Sur les 40 gènes, 27 ont été améliorés soit par 0,1 µM de RP ou 0.5 µM de RP avec 10 mM de Nam et 1% de CS. Nam ou CS avaient une activité très modeste dans les deux modèles. CONCLUSION: La combinaison de RP avec Nam et CS a montré une réponse rétinoïde plus élevée que RP dans deux modèles in vitro séparés sensibles aux rétinoïdes. Nous émettons l'hypothèse que Nam et CS améliorent l'activité RP en modulant le métabolisme de tRA par l'augmentation des groupement NAD+ et en inhibant la réduction du rétinal (RAL) en rétinol, respectivement. Les résultats fournissent la preuve que cette combinaison peut améliorer l'efficacité du traitement de l'aspect de la peau photo-âgée.

Biological Activities of Aqueous Extracts from Carob Plant (Ceratonia siliqua L.) by Antioxidant, Analgesic and Proapoptotic Properties Evaluation.

The present work describes the volatile compounds profile and phytochemical content of Ceratonia siliqua L. Fifty different components have been identified. Among them, three constituents are shared i.e., 2-methlybutanoic acid, methyl hexanoate and limonene by different common carob preparations: pulp decoction (PD), seeds decoction (SD) and Rob, a sweet syrup extracted from the pulp of the carob pod. Each extract exhibits different volatile aromatic emission profiles. The antioxidant activity of the extracts was evaluated using three methods, DPPH, ABTS and FRAP, producing a dose-dependent response. The IC50, when determined by FRAP, gave the lowest values (0.66 ± 0.01, 0.73 ± 0.05 and 0.55 ± 0.00 mg/mL PD, SD and Rob, respectively). The nociception essay, after intraperitoneal injection of acetic acid in mice, demonstrated that Rob, pulp and seeds decoction extracts showed an efficient inhibition of writhes over time, with persistence over 30 min. The SD decoction revealed the highest efficacy in decreasing the writhing reflex (90.3 ± 1.2%; p < 0.001). Furthermore, the proapoptotic activity of SD against three human cell line, THP-1, MCF-7 and LOVO, evaluated by flow cytometry, showed a significantly stronger proapoptotic activity on colon cancer (LOVO) than on the other cell lines, a phenomenon known as phenotypic selectivity.

A versatile model for investigating the protective effects of Ceratonia siliqua pod extract against 1,4-dioxane toxicity.

In this study, the toxic effects of 1,4-dioxane, a common contaminant, and the protective property of Ceratonia siliqua L. pod extract (Cspe) against this toxicity are aimed to be demonstrated with a versatile model. For this purpose, Allium toxicity test was used and six different experimental groups were formed. While the control group was germinated in tap water, the application groups were germinated in mediums containing 750 mg/L Cspe, 1500 mg/L Cspe, 100 mg/L 1,4-dioxane, 750 mg/L Cspe+100 mg/L 1,4-dioxane, and 1500 mg/L Cspe+100 mg/L 1,4-dioxane. Each group was germinated in related solution for 72 h and alterations in physiological, biochemical, genetic, and anatomical parameters were investigated. Germination percentage, relative injury rate, root length, and weight gain parameters were examined as physiological parameters, and no significant difference was observed in the control group and only-Cspe-treated groups. In groups treated with 100 mg/L 1,4-dioxane, germination percentage, root length, and weight gain were significantly decreased, and the relative injury rate reached the highest value as 0.48. It was determined that all physiological parameters improved in the groups where Cspe and 1,4-dioxane treated together, and the relative injury rate decreased to 0.22 in the group treated with 1500 mg/L Cspe+1,4-dioxane. Genotoxic effects were tested by the micronucleus and chromosomal abnormality frequency, and statistically insignificant micronucleus formation was found in control group and Cspe-treated groups. Micronucleus frequency were found to be 58.00 ± 12.12 and 31.00 ± 07.38 in 1,4-dioxane and 1500 mg/L Cspe+1,4-dioxane-treated groups, respectively. This result showed that the application of 1500 mg/L Cspe had a 46.5% reduction in the frequency of 1,4-dioxane-induced micronucleus and had a protective effect on genomic integrity. It has been found that 1,4-dioxane application induces lipid peroxidation and increases malondialdehyde level 4.5 times compared with control group. Oxidative stress, which was proved by increased malondialdehyde levels in 1,4-dioxane-treated group caused induction of superoxide dismutase and catalase enzymes, and it was determined that enzyme activities increased by 1.99 and 4.9 times, respectively, compared with the control group. Cspe treatment with 1,4-dioxane caused a significant decrease in malondialdehyde level, superoxide dismutase, and catalase enzyme activities, indicating that oxidative stress formation in the cells was repressed. Abnormalities such as cell deformation, cell wall thickening, and flattened cell nuclei were seen in 1,4-dioxane-treated group in the cross sections of root tips, and the frequency of these abnormalities decreased with Cspe application. As a result, it was determined that 1,4-dioxane caused a versatile toxicity in the test material Allium cepa, whereas Cspe application had a dose-dependent protective feature against toxicity in all tested parameters.

The Aqueous Extract from Ceratonia siliqua Leaves Protects Against 6-hydroxydopamine in Zebrafish: Understanding the Underlying Mechanism.

Ceratonia siliqua L. is a Mediterranean medicinal plant traditionally cultivated for its ethnopharmacological benefits, such as antidiarrheal, antidiabetic, enhance acetylcholine, antioxidant, antiatherosclerotic, and for its possible anti-neurodegenerative potential. The aim of the present study was to evaluate the chemical composition, as well as the cognitive-enhancing, anxiolytic, and antioxidant activities of the aqueous extract from C. siliqua (CsAE) leaves against 6-hydroxydopamine (6-OHDA) zebrafish Parkinson's disease (PD) model. CsAE (0.1, 0.3, and 1 mg/L) was administered by immersion to zebrafish (Danio rerio) for eight consecutive days and one hour before each behavioral test of each day, while 6-OHDA (250 µM) treatment was supplied one day before the novel tank diving test (NTT). Qualitative and quantitative analyses were performed by the ultra-high-performance liquid chromatography (UHPLC) analysis. The memory performance was evaluated through the NTT and Y-maze tests. Additionally, the in vitro and in vivo antioxidant status and acetylcholinesterase (AChE) activity was also assessed. Our finds demonstrated that CsAE presented positive antioxidant and anti-AChE activities, which contributed to the improvement of cognitive function in the 6-OHDA zebrafish PD model.

An in vitro study of the effect of carob (Ceratonia siliqua L.) leaf extracts on gilthead seabream (Sparus aurata L.) leucocyte activities. Antioxidant, cytotoxic and bactericidal properties.

Carob leaves, the main residues of the carob tree, were investigated as a renewable and abundant source of bioactive compounds for fish aquaculture. Aqueous and ethanolic extracts obtained from carob leaves were characterized in terms of biochemical composition, antiradical and cytotoxic effects and immunostimulant and antibacterial activities. The ethanolic extract showed higher levels of total phenolics, flavonoids and condensed tannins and higher antioxidant activity than the aqueous extract. No significant immunostimulant effects were observed on gilthead seabream (Sparus aurata) head kidney leucocytes (viability, phagocytosis and respiratory burst activities and peroxidase content) after incubation for 24 h with different extracts. Furthermore, the ethanolic extracts used at 0.5, 0.75 and 1 mg mL-1 and aqueous extracts at 1 g mL-1 had a cytotoxic effect on PLHC-1 cells. When the bactericidal activity was tested against three fish pathogenic bacteria (Vibrio harveyi, Vibrio anguillarum and Photobacterium damselae) notable activity of the different extracts was detected against P. damselae at all three concentrations. A similar effect was demonstrated against V. haryeri when ethanolic extracts were used in the same range of concentrations. This work demonstrates interesting in vitro effects of carob leaf extracts and suggests it could be used as an alternative to chemical compounds with farmed fish. The concentration and nature of the extracts were very important in terms of any positive results.