Magister Scientiae - MSc / This study aimed to evaluate the antimicrobial efficacy of medicinal hemp plant extracts to determine the antibacterial effects of indigenous Sansevieria species and exotic Cannabis sativa phytotherapy varieties. This study also assessed whether aqueous o / South Africa

It is evident that herbal medicines continue to be the mainstay of healthcare systems and source of livelihoods of many local communities in South Africa and other developing countries. As a result, there is an overwhelming dependence on medicinal products harvested from natural populations. This dependence has led to local extinction of some important medicinal plants that include Warburgia salutaris and Cassine transvaalensis in South Africa. Cultivation has great potential to relieve the pressure on natural populations. However, some traditional practitioners and scientists believe that cultivation may weaken medicinal properties and that increased secondary metabolites may form only under stress conditions, respectively. This is certainly true in some cases especially where infections with pathogens, browsing by herbivores or competition takes place in nature. It is however not clear how true this is with environmental stresses. The overall aim of this study was to evaluate to what degree different environmental conditions influenced antimicrobial and antioxidant activities of plants cultivated outside their natural environment. In order to address the aim of the study, exploratory and in-depth studies were undertaken. The exploratory study comprised long-lived Combretum collinum Fresen. (Combretacea), Terminalia sericea Burch. ex DC. (Combretaceae) and Sclerocarya birrea (A. Rich.) Hochst. (Anacardiaceae). Short-lived herbaceous Tulbaghia violacea Harv. (Alliaceae) and Hypoxis hemerocallidea Fish, C.A.Mey,&Avé- Lall, (Hypoxidaceae), were included as part of the exploratory study. The in depth studies were further undertaken, also with short-lived herbaceous Leonotis dysophylla Benth. (Lamiaceae), Bulbine frutescens (L.) Willd. (Asphodelaceae) and T. violacea. Acetone leaf extracts of all plants were studied for antimicrobial activity against bacteria (Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and Enterococcus faecalis) and fungi (Candida albicans, Cryptococcus neoformans and Aspergillus fumigatus). Extracts were also studied for antioxidant activity against Trolox and L-ascorbic acid standard oxidants using 2,2’-azinobis-(3-ethyl-benzothiazoline-6-sulfonic acid) (ABTS) and 2,2- diphenyl-2-picryl-hydrazyl (DPPH) free radicals, respectively. The exploratory study tested the effect of different rates of annual rainfall (≥870 mm/year, 651 mm/year and 484 mm/year) on the antibacterial activity of C. collinum, T. sericea and S. birrea growing in nature. The minimum inhibitory concentration (MIC) of acetone extracts of air-dried leaves was determined by using microplate serial dilution technique. Thin layer chromatography (TLC) and bioautography determined chemical constituents and antibacterial activity of extracts, respectively. The majority of extracts had low MIC values, which indicated good antibacterial activity against test bacteria (MIC of 240 μg/ml - 60 μg/ml). Leaf extracts of C. collinum and S. birrea against S. aureus (range of 390 – 100 μg/ml), E. coli (310 -70 μg/ml) and P. aeruginosa (520 - 70 μg/ml) had antibacterial activity increased significantly with low rate of annual rainfall. However, extracts of T. sericea against P. aeruginosa (240 - 100 μg/ml) and E. faecalis (150 - 820 μg/ml) had antibacterial activity significantly increased and decreased, respectively. Extracts of C. collinum and S. birrea against E. faecalis as well as T. sericea against S. aureus and E. coli did not show any clear correlation between activity and different rates of annual rainfall. Inconsistent results suggest that other factors in nature such as genetic variability, age difference, pathogens, herbivores or allelopathy (competition) might have influenced the antibacterial activity of extracts. The results indicate that the antimicrobial activity of plants growing in nature may be highly variable. In order to eliminate possible effect of those factors common in nature, another exploratory study was undertaken using clone T. violacea and H. hemerocallidea of similar age (Chapter 3). Plants were grown under controlled conditions that included irrigation with 1000 ml of distilled water in intervals of 3, 14 and 21 days outside natural environment. Dry mass of all plants was reduced significantly (P≤0.05) with watering interval of 21 days, which indicated the effect of water stress. Air-dried leaves of all plants were finely ground and extracted with acetone. Extracts had good antibacterial activity as attested by low MIC values (< 1 mg/ml) across watering intervals. Differences in the antibacterial activity of the extracts against test bacterial between water treatments were not statistically significant (P≤0.05). Furthermore, there was no clear correlation between the activity of extracts and water treatments in terms of the MIC and total activity values or chemical constituents. The results in general suggest that cultivation under optimal watering intervals may not necessarily weaken the biological activity of extracts. To complement the above findings, in depth studies were also undertaken with clone L. dysophylla, T. violaceae and B. frutescens of similar age growing under controlled conditions outside natural environment. The studies determined the influence of a wide range of water (50 ml – 500 ml) and temperature (15°C and 30°C) treatments on antibacterial, antifungal and antioxidant of extracts. With the exception of a crassulacean acid metabolism (CAM) plant, B. frutescens, transpiration, dry mass and leaf areas of the other two plants were reduced significantly (P 0.05) under high temperature of 30°C and lowest water supply of 50 ml. Acetone leaf extracts had some biological activity. Differences in the majority of antibacterial and antifungal activities of extracts between water and temperature treatments were not statistically significant. With the exception of the influence of temperature, the majority of the antioxidant activity of extracts was almost similar between water treatments. However, the significant reduction of the antioxidant activity of all extracts under high temperature of 30°C was indicative of great sensitivity to high temperatures. The overall findings suggest that the biological activity of plants is more likely to vary widely in nature than under controlled conditions outside the natural environment. This is an indication that natural environment cannot always guarantee high and stable biological activity. As a result, beliefs by some traditional practitioners and scientists that cultivation weakens medicinal properties and good secondary metabolites form only under stress, respectively, cannot be widely substantiated. Therefore, the study encourages cultivation of medicinal plants. It has potential to optimise yield of biomass production, and ensure uniform and quality biological activity as well as reduce misidentification. / Thesis (PhD)--University of Pretoria, 2012. / Paraclinical Sciences / unrestricted

Helicobacter pylori classified as a class 1 carcinogen is a common human pathogen implicated in certain gastrointestinal diseases. Helicobacter pylori infection is acquired mainly in childhood, especially in developing countries. H. pylori infection causes peptic ulcer, duodenitis, gastritis and cancer. The growing resistance of H. pylori to antibiotics used in its treatment as well as other innate limitations of the triple therapy has necessitated a search for alternative treatment from natural sources which could be readily available, less cost effective. The antimicrobial activity of solvents (acetone, ethanol, methanol, chloroform and water) crude extracts of Lippia javanica were investigated against 31 H. pylori strains by the agar well diffusion technique. The minimum inhibitory concentration (MIC) was determined by spectrophotometric analysis at 620 nm using the broth micro dilution method and the rate of kill by broth dilution method. Phytochemical analysis was also performed. H. pylori standard strain NCTC 11638 was included as a positive control. Metronidazole and amoxicillin were used as positive control antibiotics. The ANOVA test was used to analyze the results using SPSS version 17.0. The strains were inhibited by all the extracts with inhibition zones of diameter ranging from 0-36 mm and 0-35 mm for the control antibiotic, clarithromycin. The MIC90 ranged from 0.039- 0.625 mg/mL for acetone; 0.039-1.25mg/mL for methanol, 0.00195-0.313 mg/mL for ethanol; 0.01975-2.5 mg/mL for metronidazole and 0.0048-2.5 mg/mL for amoxicillin. Acetone extract completely inhibited strain PE369C at MIC (0.1 mg/mL) and 2× MIC (0.2 mg/mL) in 18h and at ½× MIC (0.05 mg/mL) in 36h. Strain PE466C was completely inhibited at 4× MIC in 72h. Phytochemical analysis revealed the presence of flavonoids, saponins, tannins, steroids and alkaloids. The results indicate that the extracts of the leaves of L. javanica may contain compounds with anti-H. pylori activity and merits further study to identify the compounds.

The purpose of this investigation is to determine the possible presence and the extent of nutritional material in the extracts of the green leaves of a selected group of common trees in an effort to devise simpler and more economical, yet useful and satisfactory, culture media for the use in bacteriological laboratories, particularly those on the secondary level.

Medicinal plants are the focus of intense study, in particular whether their traditional uses are supported by real pharmacological effects, or merely based on folklore. Piper capense L.f. (Piperaceae) is used traditionally for the treatment of infectious diseases, and has the potential to be a source of novel antimicrobial compound(s). Crude solvent extracts (water, methanol, hexane and acetone) and sequentially extracted subfractions of the root-bark of P. capense were prepared, of which the hexane-soluble subfraction MsAsHs was identified as the most promising antimicrobial subfraction. Phytochemical analyses of the various extracts and subfractions using TLC with numerous mobile phases and compound selective visualising reagents revealed the presence of quinones in all of the crude solvent extracts. Alkaloids, lipids/sterols/steroids, phenolic compounds and amino acids/peptides were detected in select subfractions. Gradient reverse phase HPLC analyses using 0.1% formic acid and methanol indicated three major peaks in MsAsHs. IR spectroscopy indicated that carbonyl and hydroxyl functional groups, and aromatic characteristics were present in the major compound present in MsAsHs. Further analysis using targeted LC-MS Q-TOF and quadrupole LC-MS/MS analyses indicated an empirical formula of C11H8O3. This formula was confirmed for the isolated compound by GC-MS (HP5-MS column) that identified the compound as 5-hydroxy-2-methyl-1,4-naphthoquinone (C11H8O3 MW: 188.18) with 98% certainty using the database. Although 5-hydroxy-2-methyl-1,4-naphthoquinone (also known as plumbagin) is well-known, this is the first time that the presence of this compound is reported in the Piper genus. Antimicrobial activities of P. capense root-bark extracts and the subfractions were determined against Gram-negative and Gram-positive bacteria and a yeast strain using the disk diffusion and broth micro-dilution assays. Antimicrobial activity was observed against Gram-positive bacteria, Gramnegative bacteria as well as a yeast strain, indicating broad spectrum activity. The antimicrobial activities of the crude solvent extracts decreased in the order: acetone > methanol > hexane > water. The MsAsHs subfraction demonstrated the highest antimicrobial activity with an MIC of 29 μg/ml against both Staphylococcus aureus (ATCC 12600) and Candida albicans (ATCC 10231). HPLC eluents of this subfraction that were collected in a drop-wise fashion onto silica TLC plates and assayed by bioautography, indicated that the major compound eluting at 13.6 minutes accounted for most of the antimicrobial activity. Antioxidant activity was observed for the crude water extract, crude methanol extract, crude acetone extract, MsAsAs subfraction as well as the MsAsHs subfraction. Cytotoxicity against mammalian cells in culture was observed for the crude methanol extract, crude acetone extract, crude hexane extract and the MsAsHs subfraction when determined using C2C12 cells as well as resting and PHA stimulated lymphocytes. Stability testing of the MsAsHs subfraction revealed that the antimicrobial compounds found in this subfraction appear to be stable up to 30 days at both 25°C and 40°C when assayed against S. aureus. However, when assayed against C. albicans, there was an increase in antifungal activity from 29 μg/ml to < 7 μg/ml after 30 days at both temperatures tested. This study provides scientific support for the ethnomedical use of the rootbark of P. capense as an antimicrobial. To date, the presence of plumbagin has not been reported in any other plant in the Piper genus. Due to the significant cytotoxic activity against mammalian cells reported in the current study and the mechanism of action of plumbagin, the therapeutic potential of P. capense extracts is very limited due to non-selective cytotoxicity, despite its marked antimicrobial activity. / Dissertation (MSc)--University of Pretoria, 2011. / Pharmacology / unrestricted

Magister Scientiae - MSc / Early man dabbled with the use of plant extracts to cure ailments. This practice has been passed down from generation to generation and today more than 50% of the world'sdrugs are natural products or derivatives thereof. Scientists have thus established a branch of research called natural product research. This branch of research involves the identification and purification of secondary metabolites with a specific biological activity. The methodology involves the screening of plant products for a specific biological activity, purification of the biologically active natural product by separation technology and structure determination. The biologically active natural products is then further scrutinized to serve as a novel drug or lead compound for the development of a novel drug. This research exploited this research methodology. / South Africa

In preliminary work done in a tree leaf screening project in the Phytomedicine Programme (www.up.ac.za/phyto) Ochna pretoriensis acetone leaf extracts had good antibacterial activity against several important bacterial pathogens. The main aim of this study was to isolate and characterize antibacterial compounds present in the acetone leaf extract of Ochna species growing in South Africa. In a preliminary screening, the minimum inhibitory concentration of acetone leaf extracts of Ochna natalitia, Ochna pretoriensis, Ochna pulchra, Ochna gamostigmata, and Ochna. Serullata, against Staphylococcus aureus, Escherichia coli, Enterococcus faecalisand Pseudomonas aeruginosa were determined by using a serial microplate dilution assay. The number of antibacterial compounds in the extracts was also determined by bioautography against the same bacteria. The MIC values of the five species ranged from 0.039 mg/ml to 1.25 mg/ml. The lowest average MIC values observed were for O. Pretoriensis especially against E. Faecalis and E. Coli. The most sensitive organism to all the plants was E. coli. O. Pretoriensis had the lowest average MIC value and the highest total activity value of 1538 ml/g. Based on bioautography some of the Ochna species had antibacterial compounds with similar Rf values. The thin layer chromatography chemical profiles of the five plant extracts may be useful in the taxonomy of the genus. O. Pretoriensis was chosen for fractionation and isolation of antibacterial compound because it has the lowest average MIC values and highest total activity especially against E. Faecalis and E. coli. The acetone extract of O. Pretoriensis was fractionated into seven fractions (hexane, carbon tetrachloride, chloroform, ethyl acetate, 35% water in methanol, 70% water in methanol and butanol) by solvent-solvent fractionation. Only three of the seven fractions (carbon tetrachloride, chloroform, ethyl acetate fractions) had clearly defined antibacterial spots/lines on bioautograms. The three fractions were further fractionated using column chromatography from which three compounds were successfully isolated. The chemical structures of the isolated compounds were determined using NMR spectroscopy as β-Sitosterol (SS), ochnaflavone (OF) and ochnaflavone 7-O- methyl ether (OFME). Compounds that are related to sitosterol have activity against neurodegenerative disorders as well as estrogenic, analgesic, anti-inflammatory, anthelminthic and antimutagenic activity. OF and OFME are biflavonoids which belong to the group ochnaflavones previously characterized from Ochna obtusata. These compounds have anti-atherosclerotic, anti-inflammatory, and anti-tumor activity. They also inhibit lymphocytes proliferation, archidonic acid release and phospholipase activity. Moreover, OFME was reported to inhibit HIV-1 activity as well as HIV-1 reverse transcriptase activity. The antibacterial activity, and potential cytotoxic, genotoxic and antigenotoxic effects of the isolated compounds were determined. The MIC values ranged from 31.3 to 250 μg/ml. SS was more active against P. Aeruginosa with an MIC of 62.5 μg/ml, OF against P. Aeruginosa and E. Faecalis with MICs of 0.03 mg/ml and OFME against P. Aeruginosa with an MIC of 31.3 μg/ml. The isolated compounds were much less active than the positive control gentamycin. The compounds had low cytotoxic activity, with LC50 values of 193.8 μg/ml for β-Sitosterol, 125.9 μg/ml for OF and 125.9 μg/ml for OFME against Vero cells. The therapeutic indexes of the crude extract and the isolated compounds varied between 0.77 and 3.27, which is an indication of non-specific antibacterial activity i.e. general toxicity, thus the crude plant extract and compounds isolated from O. Pretoriensis can only be recommended for external applications. e.g. topical treatments. None of the compounds tested had potential genotoxic and/or antigenotoxic effects. The number of revertants in the mutagenicity experiments was less than twice the number of revertants in the negative control. The percentage inhibition of 4NQO in the antimutagenicity experiments were less than 45%. The results obtained in this case may be principally associated with the general toxicity of the test samples to the bacteria used in this study. Comparison of the total activity of the crude extract and the fractions gave a clear indication of synergic interaction of compounds in the crude extract to successfully inhibit the growth of the test pathogens. Approximately 76% of activity was lost in the 34% of dry mass lost during fractionation. Twelve percent of activity was present in the chloroform fraction and 6% in the carbon tetrachloride fraction. Despite the evidence for synergistic activity, the crude extract was also relatively toxic to the Vero cells with a therapeutic index of 0.8. As far as could be established, the antibacterial activity of members of the Ochna genus and the cytotoxicity of ochnaflavones were determined for the first time in the current study. The two most active antibacterial compounds (ochnaflavone and ochnaflavone 7-O- methyl ether) are being reported from this species for the first time. The relative safety of the crude extract and the compounds isolated from this plant was relatively low. Preparations of O. Pretoriensis may be safe in a topical application but internal use cannot be recommended for treating antibacterial infections before animal toxicity studies have been carried out. Caution is also required in using the isolated compounds or crude extracts for other applications. Copyright / Dissertation (MSc)--University of Pretoria, 2009. / Paraclinical Sciences / unrestricted

Medicinal plants have played an important role in drug discovery, with many pharmaceutical products originating from plants. Isolation and characterization of antibacterial compounds is still relevant today because of continuing development of resistance of bacteria to antibiotics. The aim of the study was to evaluate the antibacterial activity of leaf extracts of nine tree species (Acalypha sonderiana, Androstachys johnsonii, Dracaena mannii, Drypetes natalensis, Funtumia africana, Necepsia casteneifolia, Oncinotus tenuiloba, Turraea floribunda, and Xylia torreana) selected from the Phytomedicine Programme Database based on good antimicrobial activities. The next step was to select the most active plant species and to isolate and characterize the antibacterial compounds. A serial microplate dilution method was used to determine the minimal inhibitory concentration and bioautography was used to determine the number of antibacterial compounds in the extract and their Rf values. Four nosocomial infection pathogens (Escherichia coli, Enterococcus faecalis, Pseudomonas aeruginosa, and Staphylococcus aureus) were used as test organisms. Extracts of all the plant species were active with average MIC values ranging from 0.13 to 2.0 mg/ml against the four bacteria. MIC values as low as 0.08 mg/ml was obtained with F. africana and O. tenuiloba extracts against S. aureus. In bioautography seven of the nine leaf extracts had activity with clear zones of inhibition on bioautograms against the red background. F. africana was active against all four bacteria while O. tenuiloba had selective activity against P. aeruginosa with clear bands on the bioautogram. F. africana was chosen for further investigation because (a) it had good antibacterial activity against the four tested bacteria with MIC value as low as 0.08 mg/ml, (b) there were several active compounds against all the tested bacteria based on bioautography, (c) it is common in nature, and (d) as far as our literature survey could ascertain there was no published information on the antimicrobial activity of this plant species. The bulk powdered leaves of F. africana were extracted with acetone. The acetone extract was fractionated into five fractions (hexane, chloroform, butanol, H2O and 30% H2O in methanol) using solvent-solvent fractionation, to group the phytochemicals based on their polarity. Hexane and chloroform fractions were the most active with MIC values as low as 0.02 mg/ml for the chloroform fraction. One of the traditional uses of F. africana is to treat burns. As a result, the crude extract and its five fractions were also tested for anti-inflammatory activity using both the COX-1 and COX-2 assays. The crude extract and the hexane and chloroform fraction had moderate activity against both cyclooxygenase 1 and 2. The chloroform fraction was more active than the crude extract (59.7 ± 1.4%)with an inhibition of 68.2 ± 6.6%. Because there was no activity in the aqueous extracts and traditional healers usually use water as extractant, the pain relief experiences traditionally must be due to another anti-inflammatory mechanism. One antibacterial compound was isolated from the hexane fraction using column chromatography with silica gel as the stationary phase and a hexane ethyl acetate gradient as the mobile phase from low to high polarity. The isolated compound was identified as methyl ursolate using nuclear magnetic resonance (NMR) and mass spectrometry. Methyl ursolate has been isolated from a number of plant species. However, this is the first report on the isolation from Funtumia genus and the first report of its antimicrobial activity. Previous phytochemical investigation from the stem bark of F. africana led to the isolation of steroidal alkaloids of the conanine group. Methyl ursolate had a low activity with MIC values of >250 μg/ml against the four tested bacteria, but had better activity against five fungal (Candida albicans, Cryptococcus neomeforms, Fusarium oxysporum, Penicillium janthinellium, and Rhizoctonia solani) species with an MIC value of 63 μg/ml against F. oxysporum. The chloroform fraction had excellent activity with an MIC of 20 μg/ml and may be developed to become a useful complex drug. The more than one hundred fold lower activity of the isolated methyl ursolate compared to the activity of the chloroform fraction from which it was isolated, provides strong evidence of synergism. This may be good model system for studying synergism in antimicrobial preparations. / Dissertation (MSc)--University of Pretoria, 2010. / Paraclinical Sciences / unrestricted

Tesis por compendio / [EN] The main goal of this Thesis was to determine the influence of the main processing stages involved in obtaining natural extracts with high antioxidant potential from byproducts originating in the olive oil industry. Firstly, the effect of freezing and/or the drying methods applied to olive oil byproducts on the polyphenol content and antioxidant capacity of the extracts subsequently obtained was addressed. For this purpose, two byproducts were considered: olive leaves and olive pomace. Secondly, the feasibility of intensifying the extraction of olive leaf polyphenols by means of a new technology, such as power ultrasound, was approached taking both compositional and kinetic issues into account. Thirdly, how the processing conditions (drying and extraction) influence the extract's stability was evaluated. Thus, on the one hand, extracts obtained from olive leaves were subjected to in vitro digestion or dehydrated and stored at different conditions. Finally, the possibility of obtaining a dried vegetable matrix (apple) rich in olive leaf phenolic compounds was explored by addressing the influence of apple pretreatments (blanching and freezing) and drying on the final retention of infused phenolics. The antioxidant potential of extracts and the retention of infused polyphenols in apple were evaluated by means of the total phenolic content and antioxidant capacity analysis, as well as the identification and quantification of the main olive leaf polyphenols by HPLC-DAD/MS-MS. Moreover, in apple samples, the polyphenol oxidase and peroxidase activity and microstructure were also analyzed. The experimental results highlighted that both drying and freezing methods significantly (p<0.05) influenced the concentration of the main polyphenols identified in the olive leaf extracts. Thus, drying at the highest temperature tested was the best processing condition in which to obtain extracts with high antioxidant capacity and phenolic content. Ultrasound application was found to be a relevant, non-thermal way of speeding-up the antioxidant extraction from olive leaves. Thus, by appropriately tuning-up the process variables, the ultrasonic assisted extraction shortened the extraction time from the 24 h needed in conventional extraction to 15 min, without modifying either the extract composition or the antioxidant potential. As far as extract stability is concerned, the processing conditions used for obtaining the olive leaf extracts did not have a meaningful influence on bioaccessibility. Regardless of the method used, stabilizing the extracts by means of dehydration only reduced both the antioxidant capacity and the total phenolic content by around 10 %. Moreover, storage conditions did not show a significant (p<0.05) effect on the antioxidant potential of the extracts for 28 days of storage. A stable dried product (apple), rich in natural phenolic compounds (from olive leaves or tea extracts), was obtained by combining drying-impregnation-drying steps. However, it should be considered that the role of fresh apple drying on the retention of infused olive leaf polyphenols was more important than the further drying of the impregnated apple. In overall terms, olive leaves can be considered a potential source of natural phenolic compounds. Notwithstanding this, the previous drying and freezing steps applied in the raw material processing are decisive factors in the obtaining of natural extracts with high antioxidant potential. Moreover, enhancing the extraction by applying power ultrasound was stated as a non-thermal way of shortening processing times. The stability of olive polyphenols during storage and in vitro digestion was closely related to the individual component considered. Finally, the exploitation of olive leaf extracts as a means of enriching solid foodstuffs requires the use of porous solid matrices free of oxidative enzymes. / [ES] El objetivo principal de esta Tesis fue determinar la influencia de las principales etapas de procesado implicadas en la obtención de extractos naturales con alto potencial antioxidante a partir de los subproductos originados en la industria del aceite de oliva. En primer lugar, se evaluó el efecto de los métodos de congelación y/o secado de la materia prima (hojas y orujo), sobre el contenido polifénolico y la capacidad antioxidante de los extractos. En segundo lugar, se abordó la intensificación de la extracción de polifenoles de hoja de olivo con ultrasonidos de potencia, teniendo en cuenta: composición y la cinética del proceso. A continuación, se estudió cómo las condiciones de procesado (secado y extracción) podían influir en la estabilidad de los extractos. Así, extractos de hojas de olivo fueron sometidos a digestión in vitro o deshidratados y almacenados a distintas condiciones. Por último, se exploró la posibilidad de obtener una matriz vegetal deshidratada (manzana) y rica en compuestos fenólicos de hoja de olivo. Para ello, se evaluó la influencia de los pretratamientos de la manzana (escaldado y congelación) y del secado en la retención final de los polifenoles impregnados. El potencial antioxidante se determinó a través del contenido total en compuestos fenólicos y la capacidad antioxidante y la identificación y cuantificación (HPLC-DAD/MS-MS) de los principales polifenoles. Además, en manzana, se midió la actividad enzimática de la polifenol oxidasa y peroxidasa y se analizó la microestructura. Los resultados manifestaron que el método de secado y el de congelación influyeron significativamente (p<0.05) en la concentración de los principales polifenoles en los extractos. Así, el secado a mayor temperatura resultó ser el mejor tratamiento para obtener extractos con alta capacidad antioxidante y alto contenido fenólico. La aplicación de ultrasonidos resultó ser una alternativa no térmica muy interesante para acelerar la extracción de antioxidantes de hojas de olivo. Con la combinación adecuada de las variables del proceso, la aplicación de ultrasonidos redujo el tiempo de extracción de 24 h necesarias en extracción convencional a 15 min, sin modificar la composición de los extractos y su potencial antioxidante. En cuanto a la estabilidad del extracto, las condiciones de procesado no tuvieron una influencia significativa en la bioaccesibilidad de los extractos. Independientemente del método utilizado, la estabilización de extractos por deshidratación sólo redujo la capacidad antioxidante y el contenido total en compuestos fenólicos en torno a un 10 %. Además, las condiciones de almacenamiento no mostraron ningún efecto significativo (p<0.05) sobre el potencial antioxidante durante los 28 días de almacenamiento. Combinando secado-impregnación-secado, fue posible desarrollar un producto deshidratado (manzana), estable y rico en compuestos fenólicos naturales (de hojas de olivo o extractos de té). No obstante, cabe destacar que el secado de la manzana fresca jugó un papel más importante en la retención de los polifenoles de hoja de olivo infundidos que el secado final de la manzana impregnada. En términos generales, las hojas de olivo pueden considerarse como una fuente potencial de compuestos fenólicos naturales. No obstante, el secado y la congelación durante el procesado de la materia prima son factores decisivos para la obtención de extractos naturales con alto potencial antioxidante. Además, la aplicación de ultrasonidos de potencia durante la extracción puede resultar una alternativa no térmica muy interesante de cara a acortar el tiempo de procesado. La estabilidad de los polifenoles de la hoja de olivo, durante el almacenamiento y la digestión in vitro, dependió claramente del compuesto individual considerado. Finalmente, el empleo del extracto de hoja de olivo como medio para enriquecer alimentos sólidos requiere del uso de matrices s / [CA] L'objectiu principal d'aquesta tesi va ser determinar la influència de les principals etapes de processament implicades en l'obtenció d'extractes naturals amb alt potencial antioxidant procedents de subproductes de la indústria de l'oli d'oliva. En primer lloc, es va estudiar l'efecte de la congelació i/o els mètodes d'assecatge aplicats a fulles d'olivera i pinyolada sobre el contingut fenòlic i la capacitat antioxidant dels extractes. En segon lloc, es va avaluar, tenint en compte la composició i la cinètica del procés, la intensificació de l'extracció de polifenols de fulla d'olivera amb ultrasons de potència. En tercer lloc, es va avaluar com les condicions de processament (assecatge i extracció) poden influir en l'estabilitat dels extractes. Així, extractes de fulles d'olivera van ser sotmesos a una digestió in vitro o deshidratats i emmagatzemats a distintes condicions. Finalment, es va explorar la obtenció d'una matriu vegetal deshidratada (poma) i rica en compostos fenòlics de fulla d'olivera considerant la influència del pretractament de la poma (escaldament i congelació) i de l'assecatge sobre la retenció final dels fenòlics introduïts en la poma. El potencial antioxidant es va avaluar determinant el contingut fenòlic total i la capacitat antioxidant, així com identificant i quantificant els principals polifenols (HPLC-DAD/MS-MS). A més, en poma l'activitat enzimàtica de la polifenoloxidasa i la peroxidasa i la microestructura. Els resultats experimentals van destacar que el mètode d'assecatge i el de congelació van influir significativament (p<0,05) en la concentració dels principals polifenols identificats en els extractes. L'assecatge a la temperatura més alta que es va provar va resultar la millor condició de processament per a obtenir extractes amb una alta capacitat antioxidant i un alt contingut fenòlic. L'aplicació d'ultrasons va ser una manera rellevant i no tèrmica d'accelerar l'extracció d'antioxidants de les fulles d'olivera. Així, amb la combinació adequada de les variables del procés, l'extracció assistida per ultrasons va escurçar el temps d'extracció, de les 24 h requerides en l'extracció convencional a 15 min, sense modificar la composició de l'extracte ni el potencial antioxidant. Quant a l'estabilitat de l'extracte, les condicions de processament utilitzades per a l'obtenció dels extractes de fulla d'olivera no van tenir una influència significativa en la bioaccessibilitat. Independentment del mètode utilitzat, l'estabilització dels extractes per mitjà de la deshidratació només va reduir la capacitat antioxidant i el contingut fenòlic total al voltant d'un 10 %. A més, les condicions d'emmagatzematge (temperatura i forma de l'extracte: líquid o pols) no van mostrar cap efecte significatiu (p<0,05) en el potencial antioxidant dels extractes durant els 28 dies d'emmagatzematge. Combinant etapes d'assecatge-impregnació-assecatge fou possible obtenir un producte assecat estable (poma) i ric en compostos fenòlics naturals (de fulles d'olivera o te). No obstant això, cal destacar que l'assecatge de la poma fresca va ser més important i determinant en la retenció dels polifenols de fulla d'olivera que no l'assecatge de la poma impregnada. En termes generals, les fulles d'olivera es poden considerar com una font potencial de compostos fenòlics naturals. No obstant això, l'aplicació d'assecatge i congelació durant el processament de la matèria primera són factors decisius per a l'obtenció d'extractes naturals amb un alt potencial antioxidant. A més, l'aplicació d'ultrasons de potència durant l'extracció resultà ser una forma no tèrmica de millorar el procés, tot reduint-ne el temps d'extracció. L'estabilitat dels polifenols d'olivera durant l'emmagatzematge i la digestió in vitro va dependre del compost individual considerat. Finalment, la utilització d'extractes de fulla d'olivera per a desenvolupar aliments sòlids enriquits requ / Ahmad-Qasem Mateo, MH. (2015). ASSESSMENT OF THE INFLUENCE OF PROCESSING CONDITIONS ON THE ANTIOXIDANT POTENTIAL OF EXTRACTS OBTAINED FROM OLIVE OIL INDUSTRY BYPRODUCTS [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/53452 / TESIS / Premios Extraordinarios de tesis doctorales / Compendio

Yes / This study used fluorescence excitation-emission matrices (EEMs) analysis to investigate the characteristics of natural organic matter (NOM) in treated water using okra crude extract (OCE), sabdariffa crude extract (SCE) and kenaf crude extract (KCE) as coagulants. In addition, an assessment of the impact of purified okra protein (POP), purified sabdariffa protein (PSP) and purified kenaf protein (PKP) was undertaken. The performance evaluation of these coagulants in terms of increase or decrease in dissolved organic carbon (DOC) was compared with Peak T fluorescence intensity observed at excitation wavelength 220–230 nm, and emission wavelength 340–360 nm. Fluorescence analysis of water treated with the crude extracts identified the removal of DOC in peaks A and C region whereas the increase in DOC from the protein was predominantly found in peaks T and B region. Furthermore, it was observed that the purified proteins were noted to be capable of reducing the DOC concentration in raw water where all fluorophores were not detected. The application of OCE, SCE and KCE yielded an increase in DOC of 65, 61 and 55% respectively, corresponding to increases of 65, 29 and 54% in peak T fluorescence intensities, at 100 mg/l dose. Furthermore, DOC concentration was reduced by 25, 24 and 18% using POP, PSP and PKP respectively as coagulants with corresponding decreases in fluorescence intensity of 46%, 44 and 36% in POP, PSP and PKP, at a lower dose of 0.1 mg/l. Therefore, it is clear that Peak T fluorescence intensity could be used to characterise organic matter in treated water using natural extracts to assess final water quality. / Financial support given to this research work by the Nigerian Government through the Tertiary Education Trust Fund (TETfund/AST &D/2013/2014/CE/02)