<p>U okviru ovog rada izvr&scaron;eno je ispitivanje različitih procesa ekstrakcije herbe industrijske konoplje. Za ekstrakciju određenih bioaktivnih jedinjenja primenjene su klasične tehnike ekstrakcije, maceracija i Soxhlet ekstrakcija, dok su od savremenih metoda primenjene ultrazvučna ekstrakcija, mikrotalasna ekstrakcija, ekstrakcija vodom u subkritičnom stanju i ekstrakcija ugljendioksidom u superkritičnom stanju. Praćenjem uticaja ulaznih promenjivih (procesnih parametara) na odabrane izlazne promenjive (sadržaj bioaktivnih jedinjenja ili antioksidativnu aktivnost) za svaku tehniku ekstrakcije pojedinačno utvrđeni su optimalni uslovi ekstrakcije. Nakon izvr&scaron;enog poređenja odabrana je najadekvatnija tehnika ekstrakcije za pripremu tečnog ekstrakta koji će biti transformisan u suvi ekstrakt primenom metode su&scaron;enja raspr&scaron;ivanjem. Prilikom dobijanja suvog ekstrakta ispitan je uticaj različite koncentracije nosača na kvalitet ekstrakata i efikasnost su&scaron;enja. Dobijeni ekstrakti su okarakterisani u pogledu fizičkih i hemijskih osobina. Sadržaj kanabinoida u tečnim ekstraktima suve herbe industrijske konoplje ispitanih u ovoj doktorskoj disertaciji je bio u opsegu od 0,4314 mg/mL CBD-a i 0,0364 mg/mL THC-a (u optimalnom ekstraktu dobijenom maceracijom) do 1,0420 mg/mL CBD-a i 0,0430 mg/mL THC-a (u optimalnom ekstraktu dobijenom mikrotalasnom ekstrakcijom). U ekstraktima koji su nepolarnog karaktera i koji su dobijeni primenom Soxhlet ekstrakcije sadržaj CBD-a i THC-a je bio 64,40 i 2,90 mg/g, dok je primenom superkritične ekstrakcije dobijen sadržaj CBD-a i THC-a u opsegu od 76,1193 do 163,1111 mg/g i od 4,1746 do 6,5803 mg/g. Tečni ekstrakti dobijeni primenom vode i etanola kao ekstragensa bili su bogati flavonoidima i fenolima, a utvrđeno je i da imaju značajnu antioksidativnu aktivnost, dok su ekstrakti dobijeni Soxhlet ekstrakcijom i ekstrakcijom ugljendioksidom u superkritičnom stanju bili bogati lipofilnim jedinjenjima i terpenima. Ovako dobijeni ekstrakti predstavljaju visokovredne proizvode koji imaju veliki potencijal za upotrebu u farmaceutskoj industriji.</p> / <p>Various extraction processes of the aerial parts of industrial hemp were examined. From classic extractions, maceration and Soxhlet extraction were applied, while from novel methods, ultrasound-assisted extraction, microwave-assisted extraction, subcritical water extraction and supercritical carbon dioxide extraction were applied. By monitoring the influence of the input variables on the selected output variables for each extraction, the optimal extraction conditions were determined separately. After a comparison of all the extraction processes tested, microwave-assisted extraction was selected to prepare the liquid extract for spray drying. The effect of different carrier concentrations was examined during the preparation of the dry extract. The extracts obtained were characterized in terms of physical and chemical properties. The content of cannabinoids in liquid extracts of dry Cannabis sativa herba examined ranged from 0.4314 mg/mL for CBD and 0.0364 mg/mL for THC in the optimum extract obtained by maceration to 1.0420 mg/mL for CBD and 0.0430 mg/mL for THC in the optimum extract obtained by microwave-assisted extraction. The CBD and THC content in extract obtained by Soxhlet extraction was 64.4000 mg/mL and 2.9000 mg/mL, respectively, while the highest content of CBD and THC in the supercritical carbon dioxide extract was 163.11 and 6.5803 mg/g. In addition to cannabinoids, the liquid extracts obtained were rich in phenols and flavonoids, and have considerable antioxidant activity, while the extracts obtained by Soxhlet extraction and supercritical carbon dioxide were rich in lipophilic compounds and terpenes. The extracts mentioned above are high value products which have great potential for implementation in the pharmaceutical industry.</p>

The diploma thesis deals with Cannabis sativa. The aim is to focus on possible utilization of Cannabis sativa in teaching chemistry. The theoretical part describes the botany of cannabis, overviews phytocannabinoids, substances contained in cannabis, and it also includes wide use of cannabis in agriculture and industry. The practical part deals with the selection and arrangement of a possible curriculum about chromatography as the most affordable technique for analysis of cannabis at school, including the formulation of school experiment methodology using the thin-layer chromatography of Cannabis sativa extract to demonstrate the presence of phytocannabinoids. The practical part also includes the scheme of an educational project on cannabis and describes the latest techniques, as well as the methodology of work with them, which are suitable for analysis of phytocannabinoids.

Submitted in fulfillment of the requirement for the degree of Doctor of Philosophy in the Faculty of Health Sciences at the University of the Witwatersrand, Johannesburg / Renewed interest in cannabinoid compounds arose since the discovery of the endocannabinoid system in the early 1990’s and its role in mediating the body’s energy balance. The aim of this study was to investigate the effect of an organic Cannabis sativa (hereafter referred to as C. sativa) extract on β-cell secretory function using an in vivo diet-induced obese rat model and an in vitro isolated rat pancreatic islet model and to determine the associated molecular changes within the pancreatic tissue. Materials and methods: Diet-induced obese Wistar rats and rats fed on standard pellets were subcutaneously injected, over a 28 day period, with an organic C. sativa extract or the vehicle (1% Tween 80® in saline). The effect of diet and treatment was evaluated using the intraperitoneal glucose tolerance tests (IPGTTs) and quantitative polymerase chain reaction (qPCR) analysis on rat pancreata. In vitro studies were conducted using isolated rat islets exposed to 11.1 (representative of normoglycemic conditions) and 33.3 mM glucose levels (representative of hyperglycemic conditions) over a 24-(D1; acute) and 96-hour (D4; chronic) period, and treated with C. sativa extract containing an equivalent of 2.5 (T1) and 5 ng/mL (T2) tetrahydrocannabinol (THC). Glucose-stimulated insulin secretion (GSIS), immunohistocytochemistry for apoptosis and proliferation detection and western blotting for detection of cannabinoid receptor type 1 (CB1), CB2 receptors and specific transduction factors were undertaken. Antagonist studies were conducted using AM251 (A1) and AM630 (A2) to block CB1 and CB2, respectively, to determine the role of cannabinoid receptors in insulin secretion. Results: The overall increase in body weight in the experimental groups occurred at a significantly slower rate than the control groups (P < 0.01), irrespective of diet. In the lean group, the area under the curve for glucose (AUCg) was significantly higher compared to the diet-induced obese group (P < 0.001), while C. sativa treatment significantly improved the AUCg in the lean rats (P < 0.05). The cafeteria diet did not induce hyperglycemia and insulin resistance in the obese rats and C. sativa treatment maintained a plasma glycemic profile similar to the obese control rats. The lower AUCg values in the obese group may, in part, be due to the inclusion of milk products (shown to be beneficial in reducing diabetes) in the cafeteria diet. qPCR analysis showed that the cafeteria diet induced down-regulation of the following genes in the obese control group, relative to lean controls: UCP2 (P < 0.01), c-MYC (P < 0.05) and FLIP (P < 0.05), and upregulation of CB1 (P < 0.01), GLUT2 (P < 0.001), UCP2 (P < 0.001) and PKB (P < 0.05), relative to the obese control group, while c-MYC levels were down-regulated (P < 0.05), relative to the lean control group. In the in vitro study, results showed C. sativa treatment decreased chronic insulin secretion in islets cultured under normoglycemic condition for D1 (P < 0.05), but not for D4. In islets cultured under hyperglycemic conditions, C. sativa treatment for the D4 period showed a significant increase in their chronic insulin secretion (HD4T1, P = 0.07; HD4T2, P < 0.001), increase in basal insulin secretion (HD4T1, P < 0.001; HD4T2, P < 0.001), increase in GSIS (HD4T1, P < 0.05; HD4T2, P < 0.001), reduction in glucose-stimulated:basal insulin production (HD4T1, P < 0.05; HD4T2, P < 0.05), reduction in insulin content (HD4T1, P < 0.001), increase in the percentage basal : content ratio (HD4T1, P < 0.001; HD4T2, P < 0.01) and increase in the percentage GSIS : content ratio (HD4T1, P < 0.001; HD4T2, P < 0.05), relative to ND4C islets. In antagonist studies, A2 preconditioning did not affect suppress the stimulatory effect of C. sativa treatment on chronic insulin secretion under normo- and hyperglycemic conditions, relative to the NC and HC islets, respectively. qPCR studies showed that C. sativa exposure induced a 2.2-fold increase in CB1 gene expression, relative to normoglycemic control islets (P < 0.05), while c-MYC and FLIP expression was significantly reduced by 12% (ND4T1, P < 0.05) and 37% (HD4T1, P < 0.05), respectively. C. sativa treatment also induced increased secretion of anti-inflammatory cytokines/chemokines under hyperglycemic conditions. Conclusion: These results suggest that C. sativa protects pancreatic islets against the negative effects of obesity (in vivo studies) and hyperglycemia (in vitro studies). In light of these findings, further investigation into the potential of C. sativa as a complementary therapeutic agent in the treatment of the deleterious effects of hyperglycemia in diabetic patients is warranted. In addition, the significant effect of C. sativa treatment on adipose tissue in experimental rats needs further investigation to determine how the cannabinoids affect the mechanisms of adipogenesis and lipolysis in diet-induced obesity. Keywords: Diet-Induced Obesity, Cannabinoids, C. sativa, THC, β-cell, AM251, AM630.

Submitted by PPG Odontologia (odontologia-pg@pucrs.br) on 2018-03-15T18:13:31Z No. of bitstreams: 1 MARIANA_KLEIN_DIS.pdf: 2818382 bytes, checksum: 8bd8dc12dea147fdea294b9c63661941 (MD5) / Approved for entry into archive by Tatiana Lopes (tatiana.lopes@pucrs.br) on 2018-03-28T11:55:01Z (GMT) No. of bitstreams: 1 MARIANA_KLEIN_DIS.pdf: 2818382 bytes, checksum: 8bd8dc12dea147fdea294b9c63661941 (MD5) / Made available in DSpace on 2018-03-28T11:59:45Z (GMT). No. of bitstreams: 1 MARIANA_KLEIN_DIS.pdf: 2818382 bytes, checksum: 8bd8dc12dea147fdea294b9c63661941 (MD5) Previous issue date: 2018-01-22 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / Ulcerative lesions are some of the most common injuries of the oral mucosa. Damage to the epithelial barrier results in discomfort, which may interfere with oral hygiene, chewing, swallowing and speech. The management of these lesions basically aims to optimize and accelerate tissue repair, in addition to reducing its symptomatology. However, there is no option considered gold standard for the treatment of traumatic mouth ulcers. Cannabidiol (CBD) is the main non-psychomimetic component of Cannabis sativa. It exerts potent antiinflammatory, antioxidant and analgesic effects when tested in various pathological conditions. The present dissertation is structured in the form of 2 scientific papers. The first article consists of a literature review to evaluate the different mechanisms of action of CBD that may be involved in the repair of ulcerative lesions, suggesting that this drug is promising for the treatment of oral inflammatory disorders. The second article is a report of an experimental study conducted in an animal model, to assess the effect of 5 and 10 mg/kg CBD on the repair of an ulcerative lesion induced on the ventral tongue of 60 Wistar rats after 3 and 7 days. Treatment with CBD decreased inflammatory scores after 3 days (p < 0.05), but was unable to clinically influence the size of ulcerative lesions. Thus, it is concluded that CBD has an inflammation modulating effect and may represent a promising alternative in the management of oral wounds. / As les?es ulceradas correspondem ?s enfermidades mais frequentes da mucosa bucal. O dano ? barreira epitelial resulta em desconforto, podendo interferir na higiene oral, mastiga??o, degluti??o e fala. O manejo dessas les?es objetiva basicamente otimizar e acelerar o reparo tecidual, al?m de diminuir a sintomatologia dolorosa. No entanto, n?o est? dispon?vel atualmente uma alternativa considerada o padr?o-ouro para o tratamento das ?lceras traum?ticas bucais. O canabidiol (CBD) ? o principal componente n?o-psicomim?tico da Cannabis sativa e desempenha potentes efeitos anti-inflamat?rios, antioxidantes e analg?sicos em diversas condi??es patol?gicas. A presente disserta??o est? estruturada na forma de 2 artigos cient?ficos. O primeiro consiste em uma revis?o de literatura, cujo objetivo foi avaliar os diferentes mecanismos de a??o do CBD que possam estar envolvidos no reparo de les?es ulceradas traum?ticas, sugerindo o car?ter promissor desta droga na terapia de dist?rbios inflamat?rios orais. O segundo trata de um estudo experimental desenvolvido em modelo animal, com objetivo de avaliar cl?nica e histologicamente o efeito da administra??o intraperitoneal do CBD, nas doses de 5 e 10 mg/kg, por 3 e 7 dias, no reparo de les?o ulcerada induzida no ventre lingual de 60 ratos Wistar. O tratamento com CBD foi capaz de diminuir os escores inflamat?rios das les?es ap?s 3 dias (p < 0,05), contudo n?o foi suficiente para interferir no tempo de cicatriza??o das ?lceras. Dessa forma, concluiu-se que o CBD ? capaz de modular o processo inflamat?rio em les?es ulceradas orais, podendo representar uma alternativa promissora no manejo dessa condi??o.

Submitted by William Justo Figueiro (williamjf) on 2015-06-15T22:51:58Z No. of bitstreams: 1 17.pdf: 23802324 bytes, checksum: 8005f41c1cffc71f7faca250bc920a4d (MD5) / Made available in DSpace on 2015-06-15T22:51:58Z (GMT). No. of bitstreams: 1 17.pdf: 23802324 bytes, checksum: 8005f41c1cffc71f7faca250bc920a4d (MD5) Previous issue date: 2013-11-28 / CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / UFPI - Universidade Federal do PIAUI / Este estudo, realizado a partir da percepção da existência de um discurso favorável à legalização do uso medicinal e recreativo da Cannabis sativa, nas atividades jornalísticas e culturais, produzido em uma nova ambiência semiótica que se articula com diversos campos do saber, como os da medicina, da antropologia, do direito, da indústria, da moda e das artes; procura compreender como os discursos antiproibicionistas foram historicamente restritos às bordas dos sistemas de comunicação, e de que modo as mobilizações discursivas que se opõem à proibição ou criminalização da Cannabis sativa, materializadas em diferentes plataformas midiáticas, constituem acontecimentos jornalísticos, que semioses acionam e que tensões de sentidos desencadeiam. Foram estudados textos jornalísticos, letras de músicas, vídeos e fotos contendo posicionamentos antiproibicionistas, e postagens de usuários no fórum de debates Growroom, sobre a organização das “marchas da maconha” de São Paulo, em 2011. / This study, developed from the perception of the existence of a speech favorable at the legalization of recreational and medicinal use of Cannabis sativa in journalistic and cultural activities, produced in a new ambience semiotics that articulates with several disciplines, such as medicine, anthropology, law, industry, fashion and arts, seeks to understand how discourses prohibitionist historically are restricted to the edges of the communication systems, and how the discursive mobilizations who oppose to prohibition or criminalization of Cannabis sativa, materialized in different media platforms constitute journalistic events, what semiosis and tension of senses are triggered. We studied newspaper articles, lyrics, videos and photos with anti-prohibitionist placements and postings of users on the Growroom discussion forum about the organization of the "marijuana marches" of São Paulo, in 2011.

Les procédés industriels de transformation des grandes productions végétales génèrent des quantités importantes de coproduits qui peuvent très souvent trouver une valorisation en tant que sources de molécules à valeur ajoutée pour l'agrochimie. Les travaux de thèse se polarisent sur une matière première originale : le chanvre (Cannabis sativa L.), plante riche en une lignine particulière, de caractéristiques différentes de celle du bois et beaucoup plus accessible. Le procédé de défibrage du chanvre (Cannabis sativa L.) génère 30% de fibres pour 70% de co-produits lignocellulosiques : chènevotte (50%) et poudre organique (20%) lesquels ont été étudiés ici comme sources potentielles d'acides hydroxycinnamiques (AHC) tels que les acides férulique (AF) et p-coumarique (ApC). Leur répartition structurale dans la matrice lignocellulosique a été évaluée analytiquement par hydrolyses séquencées. L'AF est majoritairement éthérifié à la structure lignocellulosique, et ce pour les deux matières, tandis que l'ApC est principalement sous forme estérifiée dans la poudre organique et lie en proportions équivalentes par des liaisons ester et éther dans la chènevotte. Le fractionnement des coproduits du chanvre pour l'obtention d'extraits et raffinats performants en acides phénoliques a été étudié par extraction assistée par micro-ondes et extraction thermo-mécano-chimique en extrudeur bi-vis. Ces méthodes ont toutes deux permis d'intensifier l'extraction des AHC. Pour la chènevotte, un solvant hydro-alcoolique alcalin conduit aux rendements optimaux en ApC en réacteur micro-ondes et en AF par extrusion bi-vis tandis que, pour la poudre organique, ce même solvant est le plus efficace pour extraire les deux acides phénoliques par extrusion bi-vis. L'enrichissement en AHC des extraits par adsorption sur différents solides microporeux a été étudié. La zéolithe *BEA (beta) a démontré un fort potentiel tant pour l'adsorption d'AHC de solutions modèles que pour ceux contenus dans les extraits issus des schémas de fractionnement. / The industrial transformation of common cultivated crops generates significant amounts of by-products that can often be valorized as a source of value- added molecules for biochemistry. The present work focuses on an original raw material: hemp (Cannabis sativa L.), rich in a particularly, more accessible, lignin having different characteristics than that of wood. Hemp defibering process (Cannabis sativa L.) generates 30% fibers and 70% lignocellulosic by-products: hemp shives (50%) and hemp dust (20%) which were studied in this work as a potential source of hydroxycinnamic acids (HCA) such as ferulic (FA) and p-coumaric (pCA) acids. Their structural distribution in the lignocellulosic matrix was analytically evaluated by multistage hydrolysis. FA is mostly etherified to the lignocellulosic structure, while pCA is mainly esterified in hemp dust and equally bound through ester and ether linkages in hemp shives. Biorefinery of hemp by-products, to obtain extracts and raffinates which are rich in phenolic acids, was studied by using microwave-assisted extraction and thermo-mechano-chemical extraction using twin-screw extruder. Both methods have helped to intensify the extraction of HCA. For hemp shives, alkaline-hydro alcoholic solvent lead to the optimum yields of pCA by microwave extraction and of FA by twin-screw extrusion, while for hemp dust, the same solvent is the most effective for the extraction of both phenolic acids using twin-screw extraction. The enrichment of HCA extracts by adsorption on different microporous solids has been investigated. The zeolite *BEA (beta) showed a high potential of HCA adsorption from both model solutions and extracts obtained from fractionation.

This diploma thesis deals with fractionation of protein from hemp (Cannabis sativa L.) meal according to their solubility of the hemp varieties - Fedora 17 and Uso 31. The content of nitrogenous substances and protein content in the hemp meal of these studied varieties ranged from 23,6 % (Uso 31) to 31,5 % (Fedora 17) and from 100,1 mg.g-1 (Uso 31) to 130,7 mg.g-1 respectively. The fractionation of hemp protein (Fedora 17, Uso 31) confirmed that the main hemp protein fraction is an albumin fraction, with its relative abundance - 20,5 % of protein. As a next step, the spectra of hemp proteins were determined. The use SDS-PAGE analysis revealed albumin proteins in area of 7,4 53,6 kDa. In the globulin fraction was revealed by SDS-PAGE basic subunits of the major protein edestin in the area 19-21 kDa and acidic subunits in the area 34 kDa.

Cannabinoids and cannabinoid receptors have been shown to play important roles in immune regulation particularly as modulators of anti-inflammatory cytokines and antibody production. The predominant cannabinoid receptor involved in this immune regulation is cannabinoid receptor 2 (CB2), which is robustly expressed in B cells. Utilizing a combination of bioinformatics, 5' RACE, real time RT-qPCR, and reporter assays, we showed that human B cells from peripheral blood mononuclear cells (PBMC) expressed one CB2 transcript while mouse B cells from spleen express three CB2 transcripts. Alignment of the sequenced B cell RACE products to either the mouse or human genome, along with the GenBank mRNA sequences, revealed that the transcripts isolated in this study contained previously unidentified transcriptional start sites (TSSs). In addition, expression construct testing of the genomic region containing the TSSs of the mouse CB2 exon 1 and 2 transcripts showed a significant increase of promoter activity. Bioinformatics analysis for cis-sequences in the promoter regions identified DNA binding sites for NF-kB, STAT6, and Elk1 transcription factors activated by LPS, IL-4 and anti-CD40. Regarding variations in CB2 transcript expression among the immune cell subtypes, RACE analysis showed that the exon 1b transcript is seen in B cells but not in T cells, dendritic cells or macrophages. Furthermore, RT-qPCR showed variations in transcript expression during B cell development as well as in resting versus LPS or IL-4/anti-CD40 stimulated B cells. The exon 1a transcript was predominant in pre-, immature and resting B cells whereas the exon 1b and 2 transcripts were enhanced in mature and activated B cells. These data showed for the first time that human B cells use one TSS for CB2 expression while mouse B cells use multiple TSSs for the expression of three CB2 transcripts, in which the expression of the individual transcript is related to immune cell type and/or cell activation state. Additionally, this is the first report in mouse B cells defining TSSs that are in genomic areas with promoter activity thus suggesting the location of two promoter regions. Defining the CB2 transcript expression during various stages of B cell activation provide clues to therapeutic methods.

The object of this thesis is a traditional and also controversial plant Cannabis sativa. The theoretical part is focused on ubiquitous polyphenolic compounds – flavonoids, also present in the studied plant. All previously researched and confirmed effects on human body are described in detail. The experimental part of this thesis describes the isolation and identification of substances of the chloroform and hexane yields of extract obtained from this plant. Chromatographic methods were used for isolation – thin layer and column chromatography and semipreparative HPLC. Obtained substances were characterized using UV, IR spectrophotometry and NMR. Then the anti-inflammatory activity of obtained substances and also of the ethanolic extract and its yields was observed. The result of the experimental part was the isolation of one pure substance which was identified as the cannabidiolic acid. Using the THP 1 assay, we demonstrated mild anti-inflammatory effect of the non-polar yields and isolated CBDA.

Hemp (Cannabis sativa L.) is a multi-use crop, able to provide fibre celulose a hurds for industrial treatment seeds for oil preparation biomass for energy conversion and produces secondary metabolites useful for pharmaceutical application. For its resistence to stress ability to accumulate high concentration of heavy metals and low cultivations demands, it can also be used for phytoextractions. Current research is focused on establishment of cultivation protocol, which allows transformation of callus cultures, and their regeneration with high efficiency. In this thesis, several varieties of hemp were transferred to in vitro conditions and were tested for their ability to form callus. The best results were achieved using the hypocotyl segments in a nutrient medium supplemented with 1 mg/L of naphtylacetic acid and one of these two synthetic cytokinins 0,5 mg/L of thidiazuron or 5 mg/L of 6-benzylaminopurine. No significant difference in the use of these two cytokinins were observed. None of the explants on four different test media for regeneration of shoots were able to succesfully regenerate. Transformation of hemp was tested using two different methods. Transformed protoplasts from hemp leafs after agroinfiltration were isolated. This method turn out to be unsuitable for use with hemp due to its...