published_or_final_version / Botany / Master / Master of Philosophy

Graduation date: 1987

Thesis (Ph. D.)--University of Hawaii at Manoa, 1990. / Includes bibliographical references (leaves 171-191) / Microfiche. / x, 191 leaves, bound ill., maps 29 cm

Univ., Diss.--Göteborg, 2005. / Enth. außerdem 5 Zeitschriftenaufsätze.

Thesis (Doctoral)--Göteborg Univ., 2005. / Includes bibliographical references.

Thesis (Ph. D.)--University of Rhode Island, 2005. / Typescript. Includes bibliographical references (leaves 167-174).

Seaweed biomass has been identified as a potential fermentation substrate for third generation biofuel processes due to its high carbohydrate content and its potential for mass cultivation without competing for agricultural land, fresh water and fertilisers. This thesis aimed to develop and advance existing processes to convert brown seaweeds into bioethanol. The main kelp species chosen as biomass candidates were Laminaria digitata, Laminaria hyperborea, Saccharina latissima and Alaria esculenta due to their abundance in Scottish waters and their identified potential for mariculturing. These kelp species were chemically characterised to identify seasonal variations, to recommend suitable seaweed candidates for bioethanol production and predict best harvest times. This has only been demonstrated before on one species - L. digitata. The chemical composition analyses were carried out over a 14 months sampling period, which focused on the storage carbohydrates laminarin and mannitol and the structural carbohydrates alginate, cellulose, fucoidan and xylose. In addition to carbohydrates the protein, nitrogen, carbon, polyphenol, ash and metal content was also profiled. Chemical profiling identified all four kelps as potential fermentation candidates, where maximum carbohydrate contents coincided with lowest ash and polyphenol content, usually seen in autumn. Biomass pre-treatment and saccharification are up-stream processes aimed at enhancing extraction of carbohydrates and converting those into fermentable substrates. Conversion of seaweed biomass into fermentation substrate evaluated acids and enzymes for seaweed pre-treatment and saccharification. Methodologies focused on optimising saccharification yields were developed to identify process critical parameters and develop methods for routine analysis of seaweed biomass. Results demonstrated that dilute acid hydrolysis was were less effective in releasing fermentable sugars, and also resulted in higher salinities compared to enzymatic hydrolysis using hemicellulosic and cellulosic enzymes, which were the preferred method of saccharification. All seaweeds in this thesis were assessed as fermentation substrates using the yeasts S. cerevisiae and P. angophorae, that principally ferment glucose or mannitol, respectively. Small-scale fermentation assays were developed for both yeasts to maximise ethanol yields and achieve process robustness. Both yeasts achieved a maximum ethanol yield of 0.17 g g-1 using Laminaria spp. On the basis of results, S. cerevisiae is recommended as the most useful yeast at this present point for ethanol fermentation from seaweed hydrolysates because of its tolerance to high salinity and ethanol concentrations. As salinity can negatively affect non-halotolerant enzymes, isolation of marine microorganisms was therefore carried out with the aim to highlight their enzymatic potential in seaweed saccharification. This was achieved through the isolation of two members of the genus Pseudoalteromonas, where saccharification yields using crude intracellular enzyme preparations exceeded those of dilute acids. In addition, the fermentative potential of microbial isolates as future ethanologenic strains was also evaluated. Understanding of the metabolic pathways is needed to fully assess the potential of those strains for genetic alteration. In conclusion, this thesis has demonstrated that up to ca. 20 g l-1 of ethanol can be produced from kelp species that grow on the west coast of Scotland. The procedure developed and used to produce ethanol requires further development, specifically the need for ethanol-fermenting microorganisms that can utilize mannitol and alginate; use of marine-adapted enzymes for saccharifiction; and the development of processes to achieve substrate concentration with reduced salinities. Comparison of theoretical ethanol yields from seaweed biomass with ethanol yields from terrestrial crops showed that the complete utilisation of all three major seaweed carbohydrates (laminarin, mannitol and alginate) from kelp species is needed for the process to be able to compete with 1st generation biofuel processes.

Placophora binderi can be described as an "obligate epiphyte" as it does not respond well to any culture conditions and is found growing only on other algae in the natural environment. This habit may have arisen as a response to the best available substrate in a harsh environment (Harlin 1971; Moss 1982). Any nutrient transfer which may occur between Placophora binderi and its basiphyte, usually various species of Codium, is probably by diffusion as rhizoids do not penetrate the basiphyte cells but simply lie between the Codium utricles providing better anchorage. A triphasic life history exists with isomorphic gametophyte, carposporophyte and tetrasporophyte generations. The male and female gametophytes are dioecious. This study confirms Scagel's (1953) observations for the development of the juvenile, mature and reproductive thallus. The juvenile develops as an erect polysiphonous thallus which produces a prostrate lobe as an adventitious branch from the basal segments. This prostrate lobe develops into the dorsiventrally flattened mature thallus. Reproductive structures are produced on erect branches which are initiated at the mature thallus margins. The gametophyte develops on evanescent trichoblasts produced on erect reproductive branches while the tetra sporophyte develops within these erect branches. The female gametophyte has a four-celled carpogonial branch with an auxiliary cell forming after fertilisation from the supporting cell. At the electron microscope level several vesicle types were seen in the reproductive organs. In the male, spermatial vesicles are produced which probably aid in release of the spermatia (Kugrens 1980). These are also visible under the light microscope. In carposporogenesis and tetrasporogenesis, three vesicle types are produced. Striated vesicles appear for a short while during the early stages and probably function as protein stores. Fibrillar vesicles are large and visible under the light microscope. These probably act as carbohydrate storage organelles (Triemer and Vasconcelos 1979; Kugrens and West 1973c; Tripodi 1971). Cored vesicles appear late in sporogenesis and probably aid in adhesion once the spores have settled (Chamberlain and Evans 1973; Wetherbee 1978). Carpospores follow the "serial release" type pattern observed in Polysiphonia (Boney 1978). Tetraspores are released singly via a rupture in the tetrasporangial wall as in Ceramium rubrum (Chamberlain and Evans 1973). Both carpospores and tetraspores germinate in the typical bi-polar Ceramium-type pattern described by Dixon (1973)

This study of the brown algal genus Laminaria Lamour consisted of (1) a critical review of the taxonomy and distribution of taxa of Laminaria occurring in the northeast Pacific, (2) a description of the life histories and growth patterns of long and short stipe forms of L. groenlandica and L. saccharina, (3) an evaluation of the roles of temperature, salinity, and water motion as possible determinants of local distributions of long and short stipe forms of L. groenlandica and L. saccharina, and (4) an evaluation of the roles of temperature, salinity, exposure, and submarine illumination in determining the vertical distribution of L. saccharina. These studies were made from 1961 to 1965. Ten species of Laminaria are recognized for the northeast Pacific: L. groenlandica Rosenv.; L. farlowii Setchell; L. saccharina (L.) Lamour.; L. setchellii Silva; L. dentigera Kjellman; L. longlpes Bory; L. sinclairii (Harvey ex Hooker f. et Harvey) Farlow, Anderson et_ Eaton; L. ephemera Setchell; L. yezoensis Miyabe; and L. complanata (Setchell et Gardner) Setchell. Laminaria cordata Dawson is considered conspecific with L. saccharina, L. personata Setchell and Gardner is regarded conspecific with L. yezoensis, and L. platymeris De la Pyl. (sensu Setchell and Gardner) is considered conspecific with L. groenlandica. Pour forms of L. groenlandica are recognized for the northeast Pacific. These forms are not considered as legitimate taxonomic entities but are distinguished merely to provide a means of facilitating discussion. The known habitat requirements for all ten species were broadened and the known distributions of all species, excepting L. groenlandica, were extended. Laminaria saccharina and L. groenlandica produced sori in the late spring and winter. New sporophytes of L. groenlandica appeared throughout the year, whereas those of L. saccharina appeared in late winter and early fall. Depending upon culture conditions, two morphologically distinct forms of gametophytes were produced by both species: large gametophytes were produced in conditions of high temperature and low salinity; and small gametophytes in conditions of low temperature and high salinity. Abnormal sporophytes were observed under conditions conducive to formation of large gametophytes. Patterns of growth for the blades of the two species were essentially the same.. The growth rate decreased with increase in distance from the blade base, and the position of greatest longitudinal growth coincided with the position of greatest lateral growth. The distributions of L. saccharina and the two forms of L. groenlandica about Vancouver Island were correlated with temperature, salinity, and water motion. The two forms of L. groenlandica were absent from areas of high temperature and low salinity; L. saccharina was absent from areas subjected to surf. These field conclusions were subjected to laboratory and field tests involving gametophytes and sporophytes of both species. The distributions of the two forms of L. groenlandica can be explained on the basis of temperature and salinity distributions. Both forms require low temperature and high salinity for survival. Laminaria saccharina has a wide range of tolerance to temperature and salinity. Surf appears to be the agent controlling the distribution of this species. The upper limits of L. saccharina, as observed in Burrard Inlet, are thought to-be directly related, to air temperature and insolation and indirectly related to tidal characteristics. The lower limits appear to reflect the compensation depth of this species. / Science, Faculty of / Botany, Department of / Graduate

Appropriate sampling and experimental programs resulted in a qualitative and quantitative assessment of seaweed litter biomasses, decomposition rates and concomitant changes in nitrogen content; detritus biomass and decomposition rates; and faunal distribution patterns for the significant species within a successional seaweed community in the Strait of Georgia, British Columbia, Canada. A simulation model incorporating suitable data obtained from these sampling and experimental programs facilitated prediction of detritus formation rates, biomass, nitrogen content and the seasonal availability of detritus as a food resource for fauna. Soluble matter release rates from decomposing seaweed litter and its nitrogen content were also determined. Of the ca 43 taxa identified within the seaweed litter collections, Fucus distichvs L. (41%), Irldaea cordata (Turner) Bory (26%), Nereocystis 1uetkeana (Mertens) Postels and Ruprecht (27%), and Laminaria (4%) (L. saccharina (L.) Lamouroux and L. groenlandica Rosenvinge) accounted for more than 97% of total litter deposition. The mean peak summer biomass of all litter was ca 5 g ash-free dry weight (AFDW)/m² with this figure approaching zero during January and February. Litter distribution was patchy and there was sufficient evidence to conclude that most litter was retained, and underwent decomposition, in the immediate vicinity of its place of deposition. Litter decomposition experiments performed on the 10 most significant contributors to seaweed community structure indicated that decomposition of seaweed litter occurs rapidly compared to vascular plant litter. The time required for seaweed litter to disappear from 2 mm mesh litter bags ranged from six days, for the lamina of Nereocystis luetkeana, to ca 70 days, for Fucus distichus. Some similarity in decomposition rates was observed amongst species displaying taxonomic and/or morphologic affinities. Assessment of nitrogen content of decomposing seaweed litter revealed that nine of the 10 species assayed lost nitrogen less rapidly than total litter biomass. As determined by assaying microbial consumption of particulate material, the time required for detritus (particle size < 1 mm, dry) to fully decompose was short. Of the 10 species tested, Iridaea cordata detritus decomposed most rapidly at a rate of 5.7% per day while rates for Gigartina papillata (C. Agardh) J. Agardh, Laminaria groenlandica, Laminaria saccharina and Nereocystis luetkeana ranged from 2-4% per day. Data for the remaining species were less conclusive although all decomposed at rates less than one percent per day. Variation in specific decomposition rates was shown to be correlated with the structural composition of the detritus. Those species with a relatively small percentage of crude fibre as a component of their particulate fraction decomposed more rapidly than those species with a higher percentage of crude fibre. For the two most rapidly decomposing species, Iridaea cordata and Nereocystis luetkeana, a trend toward a more rapid decomposition rate as mean particle size decreased was evident. Natural detritus (particle size < 2 mm, wet) biomass accumulation within the study site peaked at ca 1.4 g AFDW/m² during the latter half of August 19 76. This value represents 1-5% of the quantity of detritus predicted to have been formed from seaweed litter alone and a lesser percentage of the total quantity of seaweed detritus formed. Exportation out of the seaweed zone is believed to be responsible for this discrepancy. The predicted rates of detritus formation and soluble matter release from decomposing seaweed litter peaked at ca 0.6 and 0.5 g AFDW/m²per day, respectively, in early September 1976 from a low near zero in February. In total, ca 56% of litter biomass formed detritus, the remainder being released as soluble matter. The mean nitrogen contents of the detritus formed and the soluble matter released were 2.48 ± 0.03% and 1.36 ± 0.03% of their dry weights, respectively. The annual contribution of seaweed litter biomass via detritus and soluble matter to local coastal waters is estimated to be in the range of 70-85 g C/m². Detritus formed from seaweed litter was determined to have a C:N ratio of 10-13:1, rendering it suitably nutritious for utilization by fauna as a food resource, however it could not be shown conclusively that the coincidence, en masse, of specific fauna and maximum detritus availability was a response to the availability of detritus as a food resource. The possibility of such a correlation is discussed with reference to two species of caprellids, Caprella alaskana Mayer and Metacaprella anomala Mayer, and the benthic gastropod Lacuna marmorata Dall. / Science, Faculty of / Botany, Department of / Graduate