Activités amylase et lichenase d'une nouvelle souche de Bacillus. Production sur milieu solide et caractérisation.

L objectif de cette thèse était d isoler de nouvelles glycoside-hydrolases à partir d une souche de Bacillus issue d un Biotope sud-tunisien. Cette souche a montré des potentialités à produire une amylase et une lichenase à 45C et à pH 9. La production de ces deux hydrolases a été optimisée en fermentation solide sur millet, une agro-ressource de faible coût. Cette optimisation a été conduite en adoptant la méthodologie des plans d expériences. Nous avons ainsi obtenu des niveaux de production de l ordre de 540 Unités d activités amylase par gramme de substrat solide et 503 U/g d activité lichenase. Ces deux protéines ont été par la suite purifiées et caractérisées biochimiquement. L amylase présente un pH et une température d activité optimaux de 5 et 70C, respectivement. La lichenase a montré une thermoactivité et une thermostabilité remarquables qui la distinguent des lichenases précédemment décrites. En effet, l enzyme conserve plus de 20% de son activité à 100C, et plus de 60% de son activité après une incubation de 30 min à 90C. Le gène codant pour cette protéine a été isolé par la construction d une banque fosmidique dans E. coli. La comparaison de sa séquence avec la banque de données NCBI a montré que le gène de la lichenase UEB-S possède une très forte homologie avec celle de Bacillus subtilis 168, avec les positions de deux acides aminés seulement qui divergent. Un modèle de la lichenase construit au cours de cette étude laisse supposer que l un de ces deux acides aminés (Val 69) pourrait être impliqué dans sa thermostabilité, et ce en modifiant la géométrie du site de fixation au calciumThe aim of this thesis was to isolate new glycoside hydrolases from a Bacillus strain isolated from a Biotope in the south of Tunisia. This strain was able to produce a lichenase and an amylase at 45 C and pH 9. The production of these two hydrolases was optimized in solid state fermentaion using millet, a low cost. agro-resource as solid substrate. This optimization was carried out using response surface methodology (RSM) based on Doehlert design. We obtained production levels of around 540 units of amylase activity per gram of solid substrate and 503 U / g of lichenase activity.Both proteins were subsequently purified and characterized biochemically. The amylase has a pH and a temperature optimum of activity of 5 and 70 C, respectively. The lichenase showed a remarkable thermostability which distinguish it from described lichenases. Indeed, the enzyme retained more than 20% of its activity at 100 C, and more than 60% of its activity after incubation for 30 min at 90 C. The gene encoding this protein was isolated by the construction of genomic a library in E. coli. Comparison of its sequence with the NCBI database showed that the gene coding for UEB-S lichenase has a very high homology with that of Bacillus subtilis 168, with a difference in the position of only two amino acids A model for UEB-S lichenase built during this study suggests that one of these two amino acids (Val 69) could be involved in its thermostability probabely by changing the geometry of the calcium binding siteTOULOUSE-INSA-Bib. electronique (315559905) / SudocSudocFranceF

Water-soluble polysaccharides from Opuntia stricta Haw. fruit peels: Recovery, identification and evaluation of their antioxidant activities

Opuntia stricta Haw. is considered as one of the most common cactus plant growing in Tunisia. Extracting valuable compounds from its fruit peel, considered as by-product, is drawing more and more attention, making it on the verge of commercialization. Water-soluble polysaccharides were extracted from Opuntia stricta Haw. peels, and their chemical composition assessed using thin layer chromatography. The antioxidant activities of the extracted polysaccharides were assessed using 2,2-diphenyl-1-picrylhydrazyl free radical scavenging activity, total antioxidant activity and reducing power capacity. The extraction yield of water-soluble polysaccharides was 7.53±0.86%. The chemical composition revealed the presence of rhamnose, arabinose, glucose, mannose, galactose and galacturonic acid. The infra-red spectroscopic analysis showed a similar structure to that of Opuntia ficus-indica polysaccharide peels. Additionally, the extracted polysaccharides exhibited high antioxidant activities. In fact, the free radical scavenging activity (half inhibition concentration = 6.5 mg ml-1 with 94.9% inhibition at 50 mg ml-1), the total antioxidant activity (100 μg ascorbic acid equivalent at 50 mg polysaccharides) and the reducing power capacity (absorbance 700 nm = 0.7 at 50 mg ml-1), appeared to be interesting compared to natural and synthetic antioxidants. Therefore, water-soluble polysaccharides from Opuntia stricta Haw. fruit peels could be a natural alternative to replace synthetic antioxidants

Profiling beneficial phytochemicals in a potato somatic hybrid for tuber peels processing: phenolic acids and anthocyanins composition

The purpose of this study was to characterize the peels of a CN1 somatic hybrid obtained from two dihaploid potato lines (Cardinal H14 and Nicola H1) in terms of the health‐promoting phenolic compounds (phenolic acids and anthocyanins). The CN1 hybrid is defined by a pink tuber skin color making it different from the light‐yellow‐skinned “Spunta,” which is the most commonly grown potato cultivar in Tunisia. Oven‐dried peel samples derived from CN1 hybrid and cv. Spunta were ground, and phenolic compounds were extracted with water or methanol for quantification. Lyophilized peels were used for the phenolic acid and anthocyanin analyses. Higher total quantities of phenolic compounds were recovered in methanol extracts compared with water extracts. A slightly higher concentration of phenolic acids (100 mg/100 g DW) was obtained in the lyophilized peels extract of CN1 hybrid than in the cv. Spunta corresponding sample (83 mg/100 g DW). The profiles of the chlorogenic acid isomers were almost identical in both of CN1 hybrid and cv. Spunta. Caffeic acid (CA) and three caffeoylquinic acids (CQAs): 3‐CQA, 4‐CQA, and 5‐CQA, were identified from both genotypes, 5‐CQA being the dominant form in both potatoes. Since the CN1 hybrid has a pink skin color, its anthocyanin profile was also determined. The anthocyanin quantity in the CN1 peels was 5.07 mg/100 g DW, involving six different anthocyanins that were identified within the extract, namely, Pelargonidin‐3‐rutinoside‐5‐glucoside, peonidin‐3‐rutinoside‐5‐glucoside, coumaroyl ester of pelargonidin‐3‐rutinoside‐5‐glucoside, coumaroyl ester of peonidin‐3‐rutinoside‐5‐glucoside, feruloyl ester of pelargonidin‐3‐rutinoside‐5‐glucoside, and feruloyl ester of peonidin‐3‐rutinoside‐5‐glucoside. These results suggest that the peel waste of CN1 somatic hybrid can be considered as a promising source of high‐value compounds for food industry

Profiling beneficial phytochemicals in a potato somatic hybrid for tuber peels processing: phenolic acids and anthocyanins composition

The purpose of this study was to characterize the peels of a CN1 somatic hybrid obtained from two dihaploid potato lines (Cardinal H14 and Nicola H1) in terms of the health-promoting phenolic compounds (phenolic acids and anthocyanins). The CN1 hybrid is defined by a pink tuber skin color making it different from the light-yellow-skinned "Spunta," which is the most commonly grown potato cultivar in Tunisia. Oven-dried peel samples derived from CN1 hybrid and cv. Spunta were ground, and phenolic compounds were extracted with water or methanol for quantification. Lyophilized peels were used for the phenolic acid and anthocyanin analyses. Higher total quantities of phenolic compounds were recovered in methanol extracts compared with water extracts. A slightly higher concentration of phenolic acids (100 mg/100 g DW) was obtained in the lyophilized peels extract of CN1 hybrid than in the cv. Spunta corresponding sample (83 mg/100 g DW). The profiles of the chlorogenic acid isomers were almost identical in both of CN1 hybrid and cv. Spunta. Caffeic acid (CA) and three caffeoylquinic acids (CQAs): 3-CQA, 4-CQA, and 5-CQA, were identified from both genotypes, 5-CQA being the dominant form in both potatoes. Since the CN1 hybrid has a pink skin color, its anthocyanin profile was also determined. The anthocyanin quantity in the CN1 peels was 5.07 mg/100 g DW, involving six different anthocyanins that were identified within the extract, namely, Pelargonidin-3-rutinoside-5-glucoside, peonidin-3-rutinoside-5-glucoside, coumaroyl ester of pelargonidin-3-rutinoside-5-glucoside, coumaroyl ester of peonidin-3-rutinoside-5-glucoside, feruloyl ester of pelargonidin-3-rutinoside-5-glucoside, and feruloyl ester of peonidin-3-rutinoside-5-glucoside. These results suggest that the peel waste of CN1 somatic hybrid can be considered as a promising source of high-value compounds for food industry

Date Seeds as a Natural Source of Dietary Fibers to Improve Texture and Sensory Properties of Wheat Bread

The aim of this work was to investigate the effect of date seed water-soluble polysaccharides (DSP) and hemicellulose (DSH) as dietary fiber sources in enhancing the wheat bread’s quality. DSP and DSH were extracted from the three date seed varieties Deglet Nour, Ghars Souf, and Allig. The extraction yields ranged from 3.8% to 6.14% and from 13.29% to 18.8%, for DSP and DSH, respectively. DSP and DSH showed interesting functional properties and were incorporated at 0.5% and 0.75% (w/w) in wheat flour with low bread-making quality (FLBM). The results showed that the addition of 0.75% DSH significantly improved the alveograph profile of the dough, and in a more efficient way than that of DSP. Furthermore, bread evaluation revealed that the addition of DSH considerably improved the volume (by 24.22%) and the texture profile of bread (decrease of the hardness and chewiness by 41.54% and 33.81%, respectively), compared to control bread (prepared with FLBM). A sensory analysis showed that the better overall acceptability was found for bread supplemented with DSH. Results in this work demonstrate that hemicellulose fraction extracted from date seeds (DSH) and added with a level of 0.75% to FLBM represents the component that improved bread quality the best

Antioxidant and antimicrobial activities of solvent extract obtained from rocket (Eruca sativa L.) flowers

International audienceBackground and aim: Eruca sativa or most commonly known as rocket is a worldwide herbaceous plant usually used for salad preparations due to its astringent properties. The health benefits of its leaves and seeds were widely investigated, however no study has been conducted on its flowers demonstrating these benefits. The aim of this work is to investigate the phytochemical properties of solvent extract from rocket flowers. Method: Rocket flowers were collected at their blooming day and solvent extracts were prepared meaning hydrodistillation. Non-polar compounds were extracted using hexane and the efficiency of the obtained extract was evaluated using antibacterial and antioxidant activities. The antibacterial activities were tested against 11 pathogenic strains, whereas the antioxidant activities were tested through DPPH free radical scavenging activity, total antioxidant capacity and β-carotene bleaching test. Results: The antibacterial activities showed good growth inhibition compared to positive controls. The diameter of the inhibition zones reached a maximum of 16.7 ± 0.1 mm when tested against Salmonella typhimurium using 14 mg extract. The results obtained for the antioxidant activities showed more than 90% DPPH free radical inhibition, 315 μg AAE/ml for 71 mg/ml extract and more than 70% inhibition using β-carotene bleaching assay. Conclusion: The obtained phytochemical properties demonstrated the health benefit features of rocket flowers and their potential uses as feedstock of bioactive molecules

Date Seeds as a Natural Source of Dietary Fibers to Improve Texture and Sensory Properties of Wheat Bread

International audienceThe aim of this work was to investigate the effect of date seed water-soluble polysaccharides (DSP) and hemicellulose (DSH) as dietary fiber sources in enhancing the wheat bread’s quality. DSP and DSH were extracted from the three date seed varieties Deglet Nour, Ghars Souf, and Allig. The extraction yields ranged from 3.8% to 6.14% and from 13.29% to 18.8%, for DSP and DSH, respectively. DSP and DSH showed interesting functional properties and were incorporated at 0.5% and 0.75% (w/w) in wheat flour with low bread-making quality (FLBM). The results showed that the addition of 0.75% DSH significantly improved the alveograph profile of the dough, and in a more efficient way than that of DSP. Furthermore, bread evaluation revealed that the addition of DSH considerably improved the volume (by 24.22%) and the texture profile of bread (decrease of the hardness and chewiness by 41.54% and 33.81%, respectively), compared to control bread (prepared with FLBM). A sensory analysis showed that the better overall acceptability was found for bread supplemented with DSH. Results in this work demonstrate that hemicellulose fraction extracted from date seeds (DSH) and added with a level of 0.75% to FLBM represents the component that improved bread quality the best

High Throughput Screening for Bioactive Volatile Compounds and Polyphenols from Almond ( Prunus amygdalus ) Gum: Assessment of Their Antioxidant and Antibacterial Activities

International audienceThis work was devoted to recover and identify bioactive polyphenols and volatile compounds from almond gum, which can be used as food additives and/or nutraceuticals. For this purpose, solvent extract was obtained after hydrodistillation of almond gum and the extracted volatiles were identified using gas chromatography‐mass spectrometry (GC‐MS). The major compounds were 9‐octadecenoic acid (18.47%), 3‐eicosene (17.04%), hexadecanoic acid (13.82%), benzyl salycilate (9.54%), 1‐octadecene (7.28%), cetene (4.38%) and turmerone (3.2%). Subsequently, the antioxidant and antibacterial activities of the obtained extracts rich in polyphenols and volatile compounds were evaluated. Antioxidant activities showed 90.2% DPPH free radical inhibition and 206.8 µg ascorbic acid equivalents/mL using 50 mg/mL solvent extract, whereas the reducing capacity at 700 nm and 30 mg/mL was 0.8 ± 0.09. In addition, the antibacterial activities against 10 pathogenic strains were studied, showing the maximum growth inhibition, when using 50 mg solvent extract, against Enterobacter spp., Salmonella typhimurium, Listeria monocytogenes, Micrococcus luteus and Bacillus subtilis with diameter of inhibition zones of 30.4 ± 0.5, 19 ± 1.3, 27.5 ± 1.2, 28.2 ± 1.0 and 25.5 ± 0.3 mm, respectively

Solvent extract from Opuntia stricta fruit peels: Chemical composition and Biological activities

International audienceBackground and aim: Opuntia stricta is belonging to the Cactaceae family. Its fruit is composed of around 69% peels, 21% pulp and 10% seeds. Extracting bioactive compounds from the peels; the major part usually discarded, helps to reduce the cost and environmental concerns associated with their disposal. Many studies have been interested in extracting dyes and phenolics from O. stricta peels, showing their antioxidant properties, but no work was devoted to extract and characterize the non-polar compounds meaning hydrodistillation. Method: O. stricta fruits were collected at ripening, the peels, pulps and seeds were manually separated. Peels were then blended, hydrodistilled, and the non-polar compounds were extracted and identified using gas chromatography–mass spectrometry. Afterwards, the antioxidant and antibacterial activities of the extracted molecules were investigated. Results: The extracted non-polar compounds from O. stricta fruit peels were mainly terpene alcohols. The major components were trans-linalool oxide, cis-linalyl oxide and linalool with 38.3%, 29.6% and 23.4%, respectively. The antioxidant activities showed high inhibition of the DPPH free radicals with 84% at 50 mg/ml, higher reducing power than that of ascorbic acid, and high total antioxidant activity with 309 ± 37 μg ascorbic acid equivalent at 25 mg/ml. The antibacterial activities showed high growth inhibition against Staphylococcus aureus and partial inhibition against Enterococcus faecalis. Conclusion: Non-polar compounds extracted meaning hydrodistillation from O. stricta fruit peels exhibit high antioxidant activities and inhibit the growth of S. aureus. They represent thus a promising way for the valorization of this by-product

Efficiency of almond gum as a low-cost adsorbent for methylene blue dye removal from aqueous solutions

International audienceThis study was aimed to use almond gum as a potential adsorbent to remove methylene blue dye from aqueous solution. The adsorbent was characterized by scanning electron microscopy and Fourier transform infrared spectrometer. The adsorption of methylene blue on almond gum material was studied as a function of almond gum dose (0.05–2 g), pH solution (3–10), contact time (up to 240 min) and initial concentration (20–200 mg L−1), temperature (303.16 K, 313.16 K, and 323.16 K), and agitation(up to 250 rpm). The influence of these parameters on the adsorption capacity was studied using the batch process. The experimental data were analyzed by the Langmuir, Freundlich, and Tempkin isotherms. Results show that the data fitted well with the Freundlich isotherm (R2 = 0.99). The maximum adsorption capacities of methylene blue onto almond gum were found to be 250 mg g−1, 333.33 mg g−1, and 500 mg g−1 at 303.16 K, 313.16 K, and 323.16 K, respectively. The kinetic data were fitted to the pseudo-first-order, pseudo-second-order and intraparticle diffusion models, and it is revealed that adsorption of methylene blue onto almond gum follows closely the pseudo-second-order kinetic model. Thermodynamic parameters such as enthalpy change (ΔH), entropy change (ΔS), and free energy change (ΔG) were evaluated to predict the nature of adsorption. The calculated values of ΔH, ΔS, and ΔG for uptake of methylene blue were 52.078 kJ mol−1, 177.91 J mol−1 K−1, and −18.56 kJ mol−1, respectively. These results indicate the endothermic and spontaneous nature of the adsorption process. The results revealed that almond gum adsorbent is potentially an efficient and low-cost adsorbent for adsorption of methylene blue