No description available.

Proteins secreted or shed by tumors can be found in serum. Detecting these proteins by mass spectrometry (MS) is difficult, due to the wide dynamic range of protein concentrations in serum. To circumvent this issue, we mined the conditioned media of epithelial ovarian cancer (EOC) cell lines which is a less complex fluid to work with. We hypothesize that some of the proteins shed or secreted by EOC cell lines are similar to those secreted or shed by EOC tumors and that some of these proteins can be used as biomarkers. We mined the conditioned medium of four ovarian cancer cell lines (HTB75, TOV-112D, TOV-21G and RMUG-S) by two-dimensional liquid chromatography-mass spectrometry. Our study identified 1208, 1252, 885, and 463 proteins from the HTB-75, TOV-112D, TOV-21G, and RMUG-S cell lines respectively. In all, we identified 2039 proteins from which we focused on 420 extracellular and plasma membrane proteins. High abundance proteins such as albumin and immunoglobulins, which are problematic for serum proteomics, did not interfere with our study. Several known markers of EOC including CA-125, HE4, Mesothelin, and KLK6, were identified in this study. The list of 420 extracellular and membrane proteins was cross-referenced with the proteome of ascites fluid to generate a final list of 51 potential candidates. According to Ingenuity Pathway Analysis, two of the top 10 diseases associated with our list of 51 proteins were cancer and reproductive diseases. Of the 51 candidates, 10 proteins were selected for verification in sera from ovarian cancer patients and healthy individuals. Clusterin showed a significant difference between cancer patients and normal, with sera from cancer patients showing higher levels. Another protein, NPC2, did not show a difference in sera between cancer and normals. Protein expression studies using immunohistochemistry showed that NPC2 is highly expressed in ovarian cancer tissue and absent in normal ovarian surface epithelium. In summary, clusterin and NPC2 appear to play a role in ovarian cancer pathobiology and their role in EOC need to be studied further.

No description available.

Graduation date: 1992

Accurate quantification of phosphorus (P) fractions in manures and agricultural runoff is vital to understanding P dynamics in the environment. Phytate is an organic form of P, with 6 phosphate groups, which is found in manures, but is difficult to quantify due to its affinity for complex formation with aluminum (Al) and iron (Fe). The objective of the first study was to measure if high concentrations of aluminum (Al) and iron (Fe) hinder accurate quantification of phytate in dairy manure and broiler litter when measured by high performance ion chromatography (HPIC). In this study dairy manure and broiler litter samples were spiked with Al, Fe, and phytate. Samples were alkaline extracted, acidified,cleaned up and filtered, and then phytate was analyzed with HPIC. High concentrations of Fe did not hinder phytate recovery in manure or litter samples. While phytate recovery was close to 100% at typical manure and litter Al concentrations, high concentrations of Al inhibited phytate recovery in litter samples and in some manure samples. Overall, alkaline extraction of dairy manure and broiler litter and analysis with HPIC proved to be a relatively accurate, fast and cheap within normal Al and Fe ranges, compared to the commonly used NMR method. The developed method was then used to measure the concentration of phytate P (Pp) rather than the entire phytate molecule (6 moles Pp per mole phytate) in runoff waters and in an adsorption study. Phytate P has not been previously measured in runoff, due to the low concentrations of Pp found in runoff and the complexity Pp extraction from runoff. Runoff treatments were dairy manure with and without added Pp (DM, DM+Pp), broiler litter with and without added Pp (BL, BL+Pp), and control, with and without two levels of Pp (control, control Pp low, control Pp high). Runoff was collected under simulated rainfall and analyzed for total and dissolved reactive P (DRP). Runoff was also separated into sediment (>0.45"m) and liquid (<0.45"m and lyophilized) fractions for Pp analysis by high performance ion chromatography (HPIC). The new method for Pp analysis recovered 70% of Pp spiked into runoff samples. Phytate P was found only on the sediment fraction of runoff and was not detected in the liquid fraction, even when it was lyophilized and extracted. This agreed with the adsorption study, which showed strong adsorption of Pp. Phytate P loss from control Pp high was significantly greater than Pp loss from control. �When control Pp high was removed from analysis, Pp loss from BL, BL+Pp, DM, and DM+Pp was significantly greater than from control. Phytate P in the manures appeared to behave differently than pure Pp salts, likely to do the formation of protein-phytate and cation-phytate complexes in the manures. Phytate P had no effect on DRP in runoff for any treatment, indicating no significant release of inorganic P through competitive binding. The majority of P lost in runoff was in fractions other than DRP and Pp in the sediment fraction. Efforts to control the erosive loss of soil during rainfall events appear to be the best way to reduce total P loss, irrespective of the presence of Pp. / Master of Science

Bibliography: leaves 225-230. / xxi, 235 leaves : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / This thesis investigates the scale-up of an ion-exchange chromatographic system to manufacture whey growth factor extract (WGFE) from cheese whey. Key scale-up problems are identified and discussed. Alternative scale-up methodologies required to overcome several of these problems are considered. / Thesis (Ph.D.)--University of Adelaide, Dept. of Chemical Engineering, 1997?

Vitamin C occurs in relatively high concentrations in fresh and processed fruits and vegetables but is found to a lesser extent in animal tissues and animal-derived products. Nearly 90 % of vitamin C in the human diet is obtained from fruits and vegetables but this can be indirect by way of commercially prepared fruit juices. These juices are often enriched with vitamin C which has been synthetically prepared. There is a wide range of such juices on the New Zealand market, and they are a significant source of dietary vitamin C for many in the population. The focus of this research is on the Keri range of juice products.The present study monitors the fate of vitamin C during storage of Keri juices up to the best-before date, and under a range of other storage and consumption situations. Two methods were adopted for determining ascorbic acid (AA, the chemical identity of vitamin C). These were the titrimetric method, which is based upon the reduction of the dye 2,6-dichlorophenolindophenol by AA in acidic solution, and liquid chromatography, which is used to separate AA from its immediate oxidation product dehydroascorbic acid. In the latter method these two analytes can be measured independently. The liquid chromatography was less successful than the simpler titrimetric method, so most of the work was done by titration. However, the concentration of dehydroascorbic acid, which has vitamin C activity in vivo, remained uncertain. Moreover, the titrimetric method could not be applied to juices with high purple anthocyanin concentrations, like blackcurrant, because the colour change at the titration end point could not be detected. pH adjustment to change colour was ineffective, and decolourisation with charcoal led to the rapid and complete destruction of AA. The concentration of AA in Keri juices at the time of manufacture were always much higher than claimed on the labels. Storage for up to nine months at room temperature resulted in a loss in AA of between 37 and 68 %, depending on the juice and exposure to fluorescent light. However, the time of storage was a much more dominant factor than light exposure. The kinetics of loss, straight lines, were most easily explained by an aerobic model of AA degradation from oxygen diffusing across the polyethylene tetraphthalate bottle wall. Overall, the label claims made were defensible in terms of the best-before date, because it took at least 100 days of storage before the AA concentration in the most susceptible juices fell below the claimed value. This is because these drinks are fast moving consumer goods and storage beyond 100 days is unlikely. (Nonetheless, the supplier (Keri Juice Company) has since adopted its new unitised method of formulating juice. This has resulted in an initially higher concentration of vitamin C as compared to the juices under investigation.) In the nine months storage experiment there was some evidence for the presence of dehydroascorbic acid in blackcurrant drinks, but not in another three juices. Pasteurisation during preparation of these drinks resulted in up to 7 % loss of AA, probably due to oxygen dissolved in water, and accelerated by heat of pasteurisation. Higher temperatures in later storage also accelerated losses. Progressive exposure of juice to air during simulated consumption of 3 L bottles over a week also accelerated losses. Finally, exposure to sunlight in a diurnal temperature environment accelerated losses five-fold higher than in total darkness. Filtration of ultraviolet light approximately halved the loss due to sunlight. Overall however, it can be concluded that AA in the Keri range of juices is very resistant to degradation of AA.

Superoxide Dismutase (SOD) activity in cell-free extracts prepared from healthy mature flowers of Petunia hybrida (variety 'Hurrah') was studied. The SOD activity in the crude extracts was stable for more than one month when stored at -20 oC. It was found that pH 7.8 is optimal for SOD activity. Different flower tissues of petunia (stigma, style and ovary) at various stages of development were extracted and analysed for SOD activity. SOD activity was found to be significantly highest in the ovary tissue of dehiscent petunia flowers. Three SOD isozymes were detected after crude extracts of the different female reproductive tissues of petunia flowers were analysed on a non-denaturing polyacrylamide gel electrophoresis system. Based on a difference in the sensitivity of the SOD isoforms to H2O2 and KCN, it is suggested that Mn-SOD, Fe-SOD and Cu/Zn-SOD were present in the crude extracts of the female reproductive tissues of petunia flowers. The response of the female reproductive parts of petunia flowers was also tested under water deficiency and high temperature (35 oC) stress. The SOD activity seemed to increase more in response to the high temperature than the water deficiency stress. Intense blue staining was observed from developing younger buds, and much lower formazan deposition was detected at the later stage. This indicates the lower O2- produced during later stages mainly due to increasing SOD synthesis. DEAE cellulose chromatography was successfully used to partially purify SOD from the ovaries of petunia flowers. The characteristics of the partially purified enzyme fraction were found to be very similar to those of the crude extracts.

Neuropathic pain is defined as "pain initiated or caused by a primary lesion or dysfunction in the nervous system". In contrast to physiological pain that warns of noxious stimuli likely to result in tissue damage, neuropathic pain serves no protective function. Examples of neuropathic pain states include postherpetic neuralgia (shingles) and phantom limb / stump pain. This pain state also exists in the orofacial region, with the possibility of several variants including atypical odontalgia and burning mouth syndrome. There is a paucity of information on the prevalence of neuropathic pain in the orofacial region. One study assessed patients following endodontic treatment and found that approximately 3 to 6percent of patients reported persistent pain. Patients predisposed to the condition atypical odontalgia (phantom tooth pain) include those suffering from recurrent cluster or migraine headaches. Biochemical and neurobiological processes leading to a neuropathic pain state are complex and involve peripheral sensitisation, and neuronal plasticity of the central and peripheral nervous systems. Subsequent associated pathophysiology includes regional muscle spasm, sympathetic hyperfunction, and centralisation of pain. The relevant clinical features of neuropathic pain are: (i) precipitating factors such as trauma or disease (infection), (ii) pain that is frequently described as having burning, paroxysmal, and lancinating or sharp qualities, and (iii) physical examination may indicate hyperalgesia, allodynia and sympathetic hyperfunction. The typical patient complains of persistent, severe pain, yet there are no clearly identifiable clinical or radiographic abnormalities. Often, due to the chronicity of the problem, afflicted patients exhibit significant distress and are poor pain historians, thus complicating the clinician's task of obtaining a detailed and relevant clinical and psychosocial history. An appropriate analgetic blockade test for intraoral sites of neuropathic pain is mucosal application of topical anaesthetics. Other, more specific, tests include placebo controlled lignocaine infusions for assessing neuropathic pain, and placebo controlled phentolamine infusions for sympathetically maintained pain. The treatment and management of neuropathic pain is multidisciplinary. Medication rationalisation utilises first-line antineuropathic drugs including tricyclic antidepressants, and possibly an anticonvulsant. Topical applications of capsaicin to the gingivae and oral mucosa are a simple and effective treatment. Neuropathic pain responds poorly to opioid medication. Psychological assessment is often crucial in developing strategies for pain management. Psychological variables include distress, depression, expectations of treatment, motivation to improve, and background environmental factors. To enable a greater understanding of neuropathic pain, thereby leading to improved treatments, high-performance liquid chromatography-mass spectrometry is one analytical technique that has the potential to contribute to our knowledge base. This technique allows drugs and endogenous substances to be assayed from one sample in a relatively short time. The technique can identify, confirm, and measure the concentrations of multiple analytes from a single sample.

For pain research, the orofacial region is unique in a number of ways. The region has complex local anatomy, including substantial sensory innervation from neural pathways, and muscles of facial expression that convey important information concerning pain intensity and associated psychological traits. Although chronic orofacial pain conditions appear prevalent, useful documentation on pain intensity ratings using well established instruments is sparse. In particular, two conditions, atypical facial pain and atypical odontalgia, are poorly understood in aetiology so that definitive treatment modalities are severely limited. The region's local biofluid, saliva, has been used to diagnose various local and systemic disease states, and to quantitate drug concentrations. However, recent studies indicate that saliva also contains some of the same peptides, e.g. bradykinin, that are involved in pain mechanisms. It may be that pharmacological-pharmacokinetic studies of these peptides could shed more information on thesignificance of their presence in saliva. This thesis consists of four major sections. Section 1 comprises of three clinical studies investigating orofacial pain. Section 2 deals with clinical laboratory studies of saliva. Section 3 is concerned with the development of chromatographic methods to assay bradykinin and its pharmacokinetics in saliva. Section 4 uses chromatography for the identification of novel salivary peptides. This thesis, then, presents clinical studies of orofacial pain and pharmacological investigations of saliva as the local biofluid.