Thesis (PhD.(Plant Science))-University of Pretoria, 2009. / Summary in English. Includes bibliographical references.

African traditional medicine relies largely on the abundant African flora estimated at several tens of thousands of species. These plants, like other living organisms, produce natural products which are organic molecules exhibiting a remarkable wide range of chemical diversity and a multiplicity of biological properties. Over the past 20 years, interest in drugs of plants origin has been reviving and growing steadily. Among the broad spectrum of natural products that are showing promise as possible leads to useful therapeutic agents are the terpenoids. In the present study, selected African medicinal plants were investigated for the presence of extractable and exploitable terpenoids as leads or raw materials for producing more potent bioactive compounds for pre-clinical drugs discovery programme for chemoprotective agents against cancer, HIV/AIDS, diabetes, hypertension, malaria and other chronic diseases. The plants investigated in this study included Callistemon salignus, C. viminalis, Melaleuca bracteata var. revolution gold, M. bracteata var. revolution green, M. trichostachya var. compata, Syzygium aromaticum and Tectona grandis. These plants were subjected to two separate regimes of phytochemical extraction protocols namely volatile and non-volatile-extraction protocol. The Callistemon species and Melaleuca species upon hydrodistillation afforded essential oils. The gas chromatographic and mass spectrometric (GC-MS) analysis of these essential oils reveals that 1,8-cineole was the major constituent of the Callistemon oils. Similarly, 1,8-cineole was the major constituents of the essential oil of M. trichostachya var. compata, while methyl eugenol was the predominant constituent of the oils of Melaleuca bracteata var. revolution gold, and M. bracteata var. revolution green. Antibacterial investigation of the essential oils showed that they possess strong to moderate inhibitory effect against selected bacteria. In the non-volatile extraction protocol, various parts of the plants were sequentially extracted with organic solvents to obtain crude extracts which were subjected to fractionation and purification protocols (chromatographic techniques and re-crystallization). The crude extracts from the leaves of the Callistemon and Melaleuca species gave a crystalline mixture of betulinic acid and oleanolic acid in an appreciable yield. The crude extract from the cloves of Syzygium aromaticum yielded oleanolic acid as the major extractive and maslinic acid as minor extractive. The crude extracts from Tectona grandis afforded betulinic acid in an appreciable yield. The elucidation of the structures of the pure extractives was achieved by extensive 1D and 2D nuclear magnetic resonance (NMR) spectroscopy as well as infra-red spectroscopy (FT-IR) and mass spectrometry (MS). Betulinic acid and oleanolic acid were chosen as seed molecules for making known and unknown derivatives for lead optimization study. The semi-synthesized compounds were 3- acetoxyoleanolic acid, 3-acetoxyoleanolic hydrazide, 3-acetoxyloleanolic hydrazone, 3-succinyl oleanolic acid, 3-acetoxybetulinic acid, 3-succinylbetulinic acid and maslinic acid di-acetate. / Thesis (Ph.D.)-University of KwaZulu-Natal, Westville, 2010.

Thesis (Ph. D.)--Rutgers University, 2010. / "Graduate Program in Medicinal Chemistry." Includes bibliographical references.

Proefschrift--Rijksuniversiteit te Utrecht, 1932. / Includes bibliographical references (p. 74-77).

Thesis (doctoral)--Universität Bern, 1904. / Cover title. Includes bibliographical references.

Thesis (Ph.D.)--Ohio University, November, 2006. / Title from PDF t.p. Includes bibliographical references.

Thesis (M. CM)--Hong Kong Baptist University, 2006. / Dissertation submitted to the School of Chinese Medicine. Includes bibliographical references (leaves 28-30).

Saccharides are the main constituents of many Chinese medicinal materials, they include monosaccharides, oligosaccharides and polysaccharides. They have been proved to have many bioactivities such as immunomodulating effects, anti-cancer effects, anti-viral activities, and anti-fatigue effects. Thereby, it is necessary to establish comprehensive qualitative and quantitative analysis of saccharides. On the other hand, saccharides are the main constituent of water extract of many Chinese medicines, which contains a large amount. Up to now, only two botanical drugs, namely Fulyzaq and Veregen, have been approved by the FDA. Despite their complex chemical profiles, 85%-95% of their chemical components could be determined, enabling effective quality control. Therefore, a higher level of qualitative and quantitative analysis is expected for saccharide-rich Chinese medicines. However, currently available methods are weak in terms of accuracy and specificity. Our study aims to develop a comprehensive qualitative and quantitative analysis of saccharides in saccharide-rich Chinese medicines. In this study, a HPLC-NH2P-ELSD quantification method is developed to determine monosaccharides and oligosaccharides. A high performance gel permeation chromatography method was developed to analyze the apparent molecular weight distribution patterns and a 3-Methyl-1-phenyl-2-pyrazoline-5-one derivatization method was used for monosaccharide composition analysis of polysaccharides. The methods were well validated and were then successfully applied in quality assessment of one Chinese medicine material and three Chinese patent drugs from different manufacturers.

Made available in DSpace on 2014-06-11T19:35:07Z (GMT). No. of bitstreams: 0 Previous issue date: 2001Bitstream added on 2014-06-13T19:24:27Z : No. of bitstreams: 1 lourenco_mv_dr_araiq.pdf: 869109 bytes, checksum: 309d6c297b12fceb302c8b5c561de256 (MD5) / Brosimum gaudichaudii Trécul é uma planta medicinal utilizada na medicina tradicional no tratamento de doenças de pele, assim como o vitiligo. As furanocumarinas lineares psoraleno e bergapteno são os principais compostos responsáveis por esse efeito. O medicamento “V” produzido a partir do vegetal Brosimum gaudichaudii é usado pela população no tratamento do vitiligo, por ser alegado que o mesmo contém bergapteno. Entretanto, nada é conhecido a respeito do conteúdo de bergapteno nesse medicamento. Além disso, o uso de furanocumarinas tem sido relacionado ao aumento na incidência de câncer de pele e outras desordens assim como mutação gênica e aberrações cromossômicas em humanos. Este trabalho reporta análises qualitativas e quantitativas do vegetal B. gaudichaudii e do medicamento “V” por CLAE-DAD e CG-EM. As análises mostraram que as furanocumarinas estão presentes em maiores quantidades no córtex das raízes do vegetal, e que o medicamento “V” contém derivados de ácidos graxos e apenas pequenas quantidades de psoraleno e bergapteno. De acordo com a composição do extrato polar de B. gaudichaudii e do medicamento “V”, os flavonóides 5,7,3’,4’-tetraidroxi-6-C-glucopiranosilflavona e 5,7,3’,4’-tetraidroxi-3-O-β-D-galactopiranosilflavonol foram identificados no extrato metanólico das folhas de B. gaudichaudii, mas estão ausentes no medicamento “V”. Ensaios biológicos para mutagenicidade mostraram que os extratos aquoso e metanólico do córtex das raízes do vegetal são mutagênicos enquanto que os do medicamento não apresentam mutagenicidade... / Brosimum gaudchaudii Trécul is a medicinal plant that has been used in traditional medicine to treat skin diseases such as vitiligo. The linear furanocoumarins psoralen and bergapten are the main compounds responsible for this effect. Drug “V” produced from B. gaudichaudii has been used by the population for the treatment of vitiligo because its alleged content of bergapten. However, nothing is known about the bergapten content in drug V. Furthermore, the use of furanocoumarins has been linked to a higher incidence of skin cancer and other disorders such as gene mutations and chromosomal aberrations in humans. This work reports the qualitative and quantitative analyses of B. gaudichaudii and drug “V” by HPLC and CG-MS. The analyses showed that furanocoumarins are present in large amounts in the root bark of the plant, and that drug “V” contains fatty acid derivatives and only small amounts of psoralen and bergapten. Concerning the polar composition of B. gaudichaudii and drug V, the flavonoids 5,7,3’,4’-tetrahydroxy-6-C-glucopyranosyl flavone and 5,7,3’,4’- tetrahydroxy-3-O-β-galactopyranosyl flavonol were identified in the methanolic extract of B. gaudichaudii leaves, but are absent in drug “V”. . Biological assays showed that the aqueous and methanolic extracts of the root bark of the plant are mutagenic, while drug “V” does not show mutagenicity. Preliminary cytotoxic assays showed that methanolic extracts of the root bark and of drug “V” are more cytotoxic than their respective aqueous extracts. In conclusion, B. gaudichaudii and drug “V” seems to present several differences not only in their chemical composition but also in the biological properties evaluated in this study.

Background and purpose: Angelicae Sinensis Raidx (Chinese Angelica, Dang Gui, DG), the dry root of Angelica sinensis (Oliv.) Diels, is one of the most popular herbs used around the world. It has been named as the “female ginseng and served as an indispensable herb to treat many obstetrical and gynecological diseases. Traditionally, DG was recommended to pregnant women to ease delivery and to eliminate complications. It is believed that the body of DG (Dang Gui Shen, DGS) is superior in nourishing blood, while the tail of DG (Dang Gui Wei, DGW) is commonly used to remove blood stagnation. Clinically, DG is commonly combined with Paeoniae Radix Alba (White Peony Root, Bai Shao, BS) and Rehmanniae Radix (Unprocessed Rehmannia Root, Sheng Di Huang, SDH) to treat disorders during pregnancy as it may not only the strengthen therapeutic effects but also eliminate adverse effects caused by each single herb. However, it is contradictory that DG may increase the risk of miscarriages reported by previous studies: the use of DGS among pregnant women, while avoiding using DGW has always been recommended since ancient times to avoid miscarriage. To date, there is no clear evidence to identify the safety of DG in pregnant women and to support the theory that different pharmaceutical effects are attributed to chemical difference between DGS and DGW. Furthermore, little is known regarding the specific effects of DG on fetal bone while limited research has been done to explore herb-herb interactions between DG and other herbs. The aims of this project are (1) to identify the safety of DG in maternal and fetal health; (2) to compare the chemical composition of DGS and DGW and their cytotoxicity; (3) to analyze the integrated role of herb-pair (DG plus BS or SDH); (4) to investigate the mechanism of specific impact of herb-herb interaction emphasis on embryonic development. Based on the theory of traditional Chinese medicine, our project is believed to provide experimental evidence to rationalize clinical use of DG in pregnant women. Method: (1) For the herbal quality control, aqueous extracts of DG, DGS, DGW, BS and SDH were prepared respectively, and their reference marker compounds were quantitatively authenticated by HPLC. In addition, pesticide residues and heavy metals in DG extract were examined by GC-MS and ICP-MS. Moreover, comparison of composition of DGS and DGW extract in terms of main constituents was performed by GC-MS and LC-MS analysis. (2) In-vivo mouse study (Segment II study), pregnant mice were randomly assigned into different dosage groups: oral administration of either distilled water as negative control, or DG extract of 2, 8, 16, 32 g/kg/day, or BS extract of 2, 16, 32 g/kg/day, or SDH extract of 2, 16, 32 g/kg/day, or DG (32 g/kg/day) plus BS (32 g/kg/day), or DG (32 g/kg/day) plus SDH (32 g/kg/day), respectively from the gestation day (GD) 6 to 15; another group mice were treated with vitamin A (200,000 IU) on the GD7, 9 and 11 as positive control. The mice were sacrificed for assessing parameters on GD18. (3) In-vitro assay using embryonic stem cell (ESC) and fibroblast 3T3 cell was conducted to investigate the cytotoxicity of DG, Z-LIG, FA, DGS, DGW, BS and SDH by MTT test, according to European Centre for the Validation of Alternative Methods. (4) For mechanistic study of DG impacts and herb-herb interactions, the expression of a characteristic set of bone formation/resorption markers, and some site-specific bone regulatory factors in fetal tissues and amniotic fluids on the GD15 were measured by ELISA. Result: (1) In the study to evaluate the safety of DG extract, maternal body weight (BW), gravid uterine weight, corrected BW change, live fetus/litter, mean fetal body weight in the group of DG (32 g/kg/day) were significantly lower than those of the negative control (p < 0.05); while resorption site/litter, post-implantation loss (PIL)/litter, percentage of abnormal skeleton were significantly higher than those of the negative control (p < 0.05). Although there was no statistical difference between IC50 values of ESCs (IC50 ESC) and 3T3 cells (IC50 3T3) after treatment with DG, Z-LIG and FA samples respectively, the IC50 Z-LIG was significantly less than IC50 FA in both ESCs and 3T3 cells (p < 0.05). It was indicated that DG extract (32 g/kg/day) might result in adverse impacts to maternal function and fetal development in mice. Z-LIG in DG extracts might be less safe compared to FA in in-vitro cultured cells and its potential impacts should be further examined its potential impacts in in-vivo studies. (2) In the study to compare the composition of main constituents from DGS and DGW water extract, HPLC quantitative analysis indicated that the ratio of FA and Z-LIG between extract from DGS and DGW is 1:1.83 and 1:1.35, respectively. Sathulenol (1), 3-butylphthalide (2), Z-butylidenephthalide (3), benzeneacetic acid (4), Z-LIG (5) and E-LIG (6) were identified by GC-MS analysis. The peak area of compound 5 in DGW extract was close to 5 times of that in DGS extract. The amounts of compound 2 and 3 in DGW extract were respectively over 20 times and 2 times higher than that in DGS extract, respectively. Except for compound 3, 5, 6, additional three compounds: coniferyl ferulate (7), FA (8), senkyunolide A (9) were identified by LC-MS analysis. The amount of compound 3, 5, 6, 7, 8, and 9 in DGW extract was higher than that in DGS extract. The peak area of compound 3 and 5 in DGW extract was over 2 times of that in the DGS extract. In MTT assay, the effect of DGS and DGW water extract on inhibition of cell viability of cultured ESCs and 3T3 cells was in a dose-dependent manner, respectively. The difference between IC50 ESC and IC50 3T3 after DGS extract treatment was statistically significance (p < 0.05), however no statistical significance was identified in DGW (p > 0.05). Both IC50 ESC and IC50 3T3 values of DGW were much lower than those of DGS (p < 0.05). (3) In the study to evaluate the role of DG plus BS or SDH, expectedly DG extract (32 g/kg/day) resulted in significant abnormalities in maternal and fetal parameters when compared with the negative control. Whereas BS or SDH extracts at a dosage of 2, 16, or 32 g/kg/day did not result in any adverse effect for both maternal health and embryonic development. There was no statistically significant difference between the IC50 ESC and IC50 3T3 value in the cytotoxicity assays of BS or SDH extracts (p > 0.05). It was indicated that the use of BS or SDH extract should be safer than DG extract in pregnant mice. More importantly, the treatment with DG plus BS or DG plus SDH extract could significantly correct abnormalities caused by DG extract alone as seen in the corrected BW change, mean fetal body weight, live fetus/litter (%), resorption site/litter (%), PIL/litter (%), skeletal variation (%), etc. (p < 0.05) in pregnant mice. (4) In the study to analyze the mechanism of herb-herb interactions, the mean values of PICP, ALP-Bone, osteocalcin, BMPs and GDF-5 in fetal tissues were significantly lower in mice treated with DG extract (32g/kg/day) alone when compared with the negative control (p < 0.05); while there was no significant difference among the mice treated respectively with BS, SDH, DG plus BS and DG plus SDH extracts with the same dosage. The outcome was similar to those of the negative control (p > 0.05). In addition, there were no significant differences in the mean value of ICTP in both fetal tissues and amniotic fluids among all mice groups (p > 0.05). Conclusion: (1) High dosage and long-term use of DG water extract may result in adverse effects on embryonic development including fetal bone malformations, hence its use is considered as not safe in pregnant women. As DG extract in this study was not contaminated by pesticide residues and heavy metals, the embryonic toxicity of DG extract can be considered as due to the intrinsic constituents of the herb. (2) As seen in cytotoxicity assay, that water extract of DGW had the lower IC50 value, hence it is believed that the higher phthalides level (3-butylphthalide, Z-butylidenephthalide, senkyunolide A Z-LIG and E-LIG) contributes to a more toxicity on both ESC and 3T3 cells. (3) Herb-pair extract of DG plus BS or SDH could significantly correct abnormalities caused by DG extract alone in pregnant mice. Therefore, herb BS or SDH not only has beneficial effects when used for treating pregnant disorders safety, but also has attenuated effects for DG when used together as herb-pair extract. (4) At the molecular biomarker level, DG extract might significantly affect bone formation rather than bone resorption. However, it could be ameliorated when applied combination with either BS or SDH. These results should be valuable for further analysis on the integrated effects of herb-herb interactions and complex mechanism of formula therapies in Chinese herbal medicine.