A systematic study of the mechanical properties of THF-HBSs by Yun et al. [223] has yielded useful insights, but the applicability to HBS systems has yet to be dem­onstrated. No corresponding systematic study for methane HBS has been published, although such studies have been initiated. The mechanical strength of a number of laboratory-formed and natural methane hydrate-bearing samples has also been mea­sured [112,113], and Waite et al. [206] provided a summary of laboratory measure­ments of the mechanical properties of HBS. Studies on mechanical properties of HBS have been hampered by complex equipment needs, and the difficulties of working with methane hydrate. Data are needed particularly for hydrate saturations below 50% and for fine-grain media because much of the hydrate in oceanic depos­its is present in low saturations providing risk to oil and gas operations.

Unlike PHB, which is accumulated intracellularly [91] , IBT is excreted from the cell (Sinskey laboratory, data not shown). Thus, continuous autotrophic IBT pro­duction is possible. A two stage approach could be used, in which the cells are first grown heterotrophically to a high cell density on the porous wall of hollow fibers. Then, the heterotrophic medium would be replaced by a nitrogen-deficient auto­trophic medium to induce the stringent response and trigger rapid IBT production. The gaseous substrates would then be added, with the H2 and O2 being maintained on opposite sides of the fiber wall. A schematic drawing of this hollow fiber reactor setup is shown in Fig. 7.

Since no growth will be supported by the production medium, the cells will need to be periodically regenerated by brief exposure to the heterotrophic growth medium. However, the intervals on growth medium can be short since there will be no require­ment to start the culture anew, thus maximizing the production time.

With regard to the aforementioned restrictions, an increase in PCE of more than a factor two cannot be expected. Process costs need to be drastically reduced by lower investment cost and lower demand for auxiliary energy to target a price range of less than 5 US$/kg biomass.

Aeration with membranes can be one major improvement for future reactor developments [13, 17]. The interfacial area for CO2 input and O2 removal from the culture in case of membrane gas exchange is defined by the surface area of the membrane itself and no longer by the surface of gas bubbles. This concept could drastically reduce the overall input of auxiliary energy since there is no longer the need to generate bubbles, and energy loss when bubbles fuse with the top headspace gas phase is avoided. However, gentle agitation will still be required for gas disper­sion. A proof of concept is missing in photo-biotechnology but implementation of membrane systems are considered by Solix Biofuels [26,47].

If beneficial light/dark cycles cannot be attained with low inputs of auxiliary energy, laminar flow patterns can be accepted if diffusion paths from gas membranes to all volume elements are small enough to supply all cells with CO2 solely by diffusion. In this case, additional power input for dispersion could be spared and the overall energy balance tremendously improved. However, this approach requires large surface areas for membranes and a drastic reduction of diffusion path length and thus simultaneously light path length. Such a short light path length could, by contrast, allow for high cell concentrations which is favorable for DSP. This approach also lacks a proof of concept.

Future development will show if membrane gassing will be established on a large scale.

With regard to the current high demand of auxiliary energy for photobioreactors, reduction of the height of reactors could significantly contribute to increased energy efficiency. “Low ceiling” concepts aim at reducing the hydrostatic pressure and therewith the energy demand for aeration systems. Additional improvements in control engineering can certainly facilitate further energy saving in the future. Requirements of carbon dioxide and therewith removal of oxygen is dependent on cell concentration and on light availability in the culture. Therewith, carbon dioxide supply and energy input for mixing should be adjusted to photosynthetic activity, photon flux-density respectively, and cell concentration. Proviron, for example, claims that the auxiliary energy input can be halved in the future by adapting aera­tion to light availability [31].

Infrared radiation is not photosynthetically convertible into chemical energy but contributes to heating of algae culture. Infrared reflecting materials or coatings could additionally improve the overall energy balance by reducing the energy required for cooling. Transparent materials with selective transmittance are avail­able and were developed for the installation in buildings, cars, and greenhouses [19,38,50].

A different approach to the difficulty of light capturing and distribution aims at harvesting the light in a module that is spatially separated from the reactor itself. The Green Solar Collector [52] harvests light by moving lenses whose orientation is guided by a computer that calculates position and altitude of the sun in order to capture maximum amount of photons. The light is then focused and transported via plastic light guides where light is totally reflected. A change in refraction index releases the photons in the microalgae suspension. It is suggested that light redis­tributing plates are integrated in small distances from each other in airlift-photobio­reactors. According to the authors biomass concentrations of up to 20 g/L could be maintained in such a setting with good light distribution and induction of beneficial high frequency light/dark cycles [20] .

Spatially separated light harvesting and reactor modules can have beneficial advantages because parameters like temperature can be controlled more easily. Furthermore, influences of unfavorable weather conditions, such as hail, will be confined only to the light collector. That will positively influence maintenance costs.

Spatial separation of light harvest and cultivation differs from all other concepts presented here, but the basic principles of other reactor designs can also be retrieved here. In order to optimize light utilization and productivity, light path length is also limited in this reactor, as the reactor contains flat panel compartments with short light path lengths. Moreover, turbulent flow patterns, induced by aeration, ensure rapid circulation of microalgae between dark zones and illuminated volume ele­ments in order to benefit from the intermittent light effect [20] .

3 Conclusion

Many efforts in the field of photo-biotechnology have not brought out the “perfect” photobioreactor, yet. All basic concepts show specific advantages and disadvan­tages that finally lead to the development of more sophisticated reactors. These should be characterized by outstanding light utilization and mass transfer, yet be operated with minimum energy input. Moreover, diverse algae strains show different behavior in terms of light saturation, robustness towards shear stress, and other cul­ture conditions. Therefore, benefits and drawbacks of different reactor concepts should be taken into account in terms of producing biomass or energy rich products for the energy market. At the same time, adjustment of the particular system to the specific algae strain will be unavoidable. Nevertheless, high productivities and photoconversion efficiencies give rise to high expectations in this field of research.

The solar receiver tubing must have a specified length so that photosynthetic growth can be optimised. It has been shown that the maximum tube length relies on three parameters: liquid velocity, dissolved oxygen concentration and the rate of oxygen production by photosynthesis. Generally, a tube run in a photobioreactor should not surpass 80 m. However, the maximum length of tubing is dependent on solar inten­sity, biomass concentration, liquid flow rate and initial oxygen concentration at the tubing entrance [22]. Molina Grima et al. [23] states that “other than “scale up” by multiplication of identical tubular modules, the only way to increase volume is by increasing length and/or diameter”. Ten years on, the debate over scale-up is still prevalent, with no clear solution readily available. A possible solution to scale-up is to make use of current cultivation designs and employ several cultivation units to produce a significant amount of biomass. However, the process must produce enough biomass such that it will offset extensive equipment costs.

Following the process recommended in Peters et al. [26], fixed costs including pip­ing, electrical systems and contractor’s fees were each estimated as a proportion of the total major equipment costs, using the factors adopted in the cost estimation of a similar algae production facility by Molina Grima et al. [21]. This approach in estimating fixed costs as a proportion of the total major equipment costs is consid­ered to typically yield results with accuracy within a ±10%. One major exception to this method was the estimation of land costs, which was examined in the cultivation stage of production. The system is assumed to be co-located with a power genera­tion station. This location is selected to ensure adequate supply of free carbon diox­ide, readily available in the form of flue gas. Taking this into consideration, the land cost of each system was modelled based on the cost of two large agricultural proper­ties situated in the area; a 32 ha property in Moe South and a 36 ha property in Tyers (REA Group, 2009) in Australia. Based on the total reactor volume required and the area required for each reactor type (Table 1), the overall land requirement and cost for each system was estimated. The FCI was obtained by summing all fixed capital costs and the total major equipment costs.

Most of the research on algae-to-energy systems carried out to date has been at the bench or demonstration scale [18]. This makes it difficult to say with much certainty what a full-scale algae-to-energy industrial facility would look like and herein lies one of the fundamental challenges of developing reliable LC estimates for algae production. Using best engineering judgment, it is possible to design hypothetical algae-to-energy facilities, but naturally, there is variability among these designs (Fig. 1). For example, one modeler might assume that algae should be cultivated in ponds, while another could assume photobioreactors [6]. Similarly, a belt filter press could be modeled as means to separate algae from the growth medium, whereas self-cleaning bowl centrifuges might be a viable alternative [24]. Both unit opera­tions carry out the same dewatering function but with different requirements in terms of inlet and outlet concentration, demand for chemical flocculants used to accelerate the settling of the algae out of solution, and energy use profiles. Similarly, there are several technically viable options for extraction of oil from algae biomass, namely: sonication [28], bead mills [7], and enzymatic processes [11]. For conver­sion of algae biomass into biodiesel, one might choose decarboxylation of fatty acids [29] and digestion of non-fatty acid fraction [27] or the conventional transesterification route. Finally, the end-product of the algae-to-energy facility can

ICIarcns 2010|

— solar drying

I Sander20I0|

— autoflocculation (Lardon 2009]

Fig. 1 In selected system boundaries for an algae LCA study, one must typically select from (a) or capture all of (b) a large number of possible unit operations also vary, because biodiesel is not the only energy carrier that can be produced from alga biomass [3]. It can be dried and combusted directly to generate electricity or it can be separated such that the carbohydrate fraction may be fermented to produce ethanol [24]. Naturally these two systems would have very different impacts.

As an example of the way in which systems boundaries selection can impact LCA results and conclusions, it’s informative to consider two of the more thoroughly documented algae LCA studies that have been published to date: Clarens et al. [8] and Stephenson et al. [31]. Clarens et al. [8] used an energy-basis functional unit and only modeled cultivation-phase burdens for open pond systems. They did not account for the possibility that energy production from algae might also create valuable coproducts since they argue that it is still unclear whether there will be tenable mar­kets for these coproducts. In contrast, Stephenson et al. [31] utilized a functional unit of 1 ton algae biodiesel to compare between open pond cultivation systems and pho­tobioreactor cultivation systems. These authors included two types of valuable coproducts: electricity, as produced via combustion of natural gas generated during anaerobic digestion of residual (non-lipid) algae biomass, and glycerin. In light of these dramatically different sets of systems inputs, it’s not surprising that each study reached different types of conclusions. Clarens et al. found algae-derived biomass energy to be generally more environmentally burdensome than corn, canola, or switchgrass alternatives. In contrast, Stephenson et al. found algae-derived biodiesel to be more environmentally beneficial than fossil-derived diesel.

Once an algae-to-energy process has been specified there is the additional uncer­tainty associated with setting system boundaries. LCA is typically intended to cap­ture all of the environmental impacts of an engineered system. Naturally, in a highly interconnected technical world, system expansion results in models that become impossibly large and complex. For example, to produce carbon dioxide for use in industrial processes, it is necessary to model ammonia production since most of the carbon dioxide in this country comes from the steam reforming of hydrocarbons to produce hydrogen, most of which is used to produce ammonia via the Haber-Bosch process [21]. This in turn requires that we understand something about the way
natural gas is produced and transported in this country and the countless unit operations that allow us to purchase a canister of relatively pure carbon dioxide for the factory. To cope with this complexity, many LCA practitioners have set arbitrary boundaries around their processes of interest. For example, one study might state that any process contributing less that 5% of the total mass or energy or other impact to the final total is neglected. In this way the problem can be distilled down to some­thing that is not computationally expensive and still yields good approximations of a process’ impact.

Beyond system design and boundary setting, LCA analysts may chose to focus on specific pieces of a larger system to provide a desired level of resolution. For example, in their work, Clarens et al. considered only the cultivation of algae argu­ing that the uncertainties with that first step in the algae-to-energy life cycle should be addressed [8]. By focusing only on cultivation, the authors were able to explore the full implications of that important LC stage including crucial upstream impacts such as fertilizer production and carbon dioxide generation and delivery. In fact, a sensitivity analysis included in this chapter suggests that these two impacts are among the most important factors driving the overall life cycle burdens of algae production. Many of the other studies assume that the upstream impacts of deliver­ing fertilizers and carbon dioxide should not be included. In Sander and Murthy, a cut off of 5% was assigned to LC contributions that would be neglected in the analy­sis [24] (Fig. 2). This represented the most rigorous treatment of boundaries from any of the studies published to date. However, this study also made certain assump­tions, notably, that the effluent from a secondary wastewater treatment plant would contain enough nutrients to sustain a community of algae [4] . This assumption is not supported by stoichiometry or by the bench-scale research and as a result their estimates for algae life cycle impacts are most likely low.

The work described here was carried out in the Department of Chemical Engineering, Monash University, in 2010 and has not been published elsewhere. The study describes the kinetics of chlorophyll extraction from T. suecica using acetone or methanol as an extractant. The three parameters investigated in order to optimize the extraction process were storage temperature of the biomass prior to chlorophyll extraction, level of intracellular water in the biomass during chlorophyll extraction, and average temperature during chlorophyll extraction.

3.2 Materials and Methods

3.2.1 Chemicals and Reagents

Chlorophyll a and chlorophyll b standards were purchased from Sigma-Aldrich Pty. Ltd (Australia). Organic solvents (100% acetone and 100% methanol) were analyti­cal grade.

Table 4 Previous studies on HPLC fractionation of chlorophylls extracted from phytoplankton

Study Mobile phase Stationary phase

Jeffrey [23] First dimension: 0.8% «-propanol in light petroleum (by volume) Sucrose

Second dimension: 20% chloroform in light petroleum (by volume)

Jeffrey et al. [24] 90:10 (v/v) methanol:acetone for 8 min at a flow rate 1 mL/min 3 pm C18 Pecosphere

Pre-injection mix of sample 3:1 (v/v) sample: 0.5 M ammonium acetate Jeffrey et al. [24] Solvent A is 80:20 (v/v) methanol:0.5 M ammonium acetate 3 pm C18 Pecosphere

Solvent В is 90:10 methanol: acetone

Elution order: 0-3 min: solvent A; 3-17 min: solvent B. flow rates: 1 mL/min Pre-injection mix of sample 3:1 (v/v) sample: 0.5 M ammonium acetate

Jeffrey et al. [24] Solvent A is 80:20 (v/v) methanol:0.5 M ammonium acetate 3 pm C18 Pecosphere

Solvent В is 90:10 (v/v) acetonitrile:water Solvent C is ethyl acetate Elution order:

0-4 min: linear gradient from 100% A to 100% В 4-18 min: linear gradient to 20% В and 80% C 18-21 min: linear gradient to 100% В 21-24 min: linear gradient to 100% A 24-29 min: isocratic flow of 100% A

Lynn Co and Three different solvent systems were experimented Silica Gel

Schanderl [27] Solvent system 1 (modified Bauer solvents):

First dimension is benzene: petroleum ether: acetone (10:2.5:2 v/v/v).

Second dimension is benzene: petroleum ether: acetone: methanol (10:2.5:1:0.25 v/v/v)

Solvent system 2:

First dimension is benzene: petroleum ether: acetone: methanol (10:2.5:1:0.25 v/v/v).

Second dimension is petroleum ether: acetone: «-propanol

(8:2:0.5 v/v/v)

Solvent system 3:

First dimension is benzene: petroleum ether: acetone (10:2.5:2 v/v/v).

Second dimension is petroleum ether: acetone: «-propanol

(8:2:0.5 v/v/v)

Seaweeds are known to be high in mineral content. More than 30% of the dry weight of marine algae is ash which contains various kinds of minerals, as they are bathed in the rich seawater. Some of the minerals are necessary for our health while some are toxic in varying degrees. Most of the macroalgae have high Ca, Mg, P, K, Na, and Fe contents [116], as can be seen in Table 4.

In comparison with higher plants, their outstanding feature is their high iodine content [89]. Seaweeds are the best natural sources of biomolecular dietary I. Some seaweeds contain 1,000 times as much iodine as found in a marine fish like cod. Seaweeds provide di-iodotyrosin (I2T) which is precursor to essential thyroid hor­mones thyroxine (T4) and triiodothyronine (T3) [14] .

The mineral content in general is highly dependant on the environmental growing conditions (season, temperature, physiological state, geographic varia­tions…). For example, in a recent study of Porphyra and Laminaria from France, Spain, Korea, and Japan [152] it was found, by using ICP-MS, that seaweeds from Korea and Japan tended to display the highest concentrations of Pb and Cd. In contrast, Spanish and French samples showed the highest levels of some microelements essential to human nutrition. Moreover, Porphyra presented

higher concentrations of most elements (Cd, Co, Cr, Mo, Ni, Pb, Sb, Se, and V), except for As, than Laminaria.

However, the linkage of certain minerals with anionic polysaccharides (alginate, agar, or carrageenan) might limit the absorption and extraction of these minerals. In such cases, mineral availability is a function of the type of linkage between the polysaccharide and the mineral. For instance, the weakness of the linkages between polysaccharides and iodine allows rapid release of this element. In contrast, the strong affinity of divalent cations (particularly Ca2+) for carboxylic polysaccharides (alginates) probably limits the availability of associated minerals. From a nutri­tional standpoint, this high affinity might be compensated by the high mineral con­tents of seaweeds [95].

Other compounds with proven bioactivity are those related with photosynthesis, mainly pigments such as chlorophylls, carotenoids or proteins like opsins. Among them chlorophylls are the most wide spread compounds. Chlorophylls and their intermediate metabolites have proved its contribution to antioxidant and antimicro­bial activities. For example, in supercritical CO2 extracts of S. platensis, chloro — phyll-a, pheophytin-a, pheophytin-a O-allomer, and pyropheophytin-a were detected by LC-MS/MS among the contributors to antioxidant activity measured by DPPH radical scavenging method [107]. On the other hand, phytol was detected by GC-MS among the bactericidal compounds present in D. salina extracts [111], as can be observed in Fig. 2, being all of them secondary metabolites of chlorophylls.

Certain alkaloids have been isolated from seaweeds. Among the many chemical classes present in plant species, alkaloids stand out as one of major importance in the development of new drugs, because they possess a wide variety of chemical structures and have been identified as responsible for many of the pharmacological properties of medicinal plants. Caulerpin, a bisindole alkaloid, was isolated from the green alga Caulerpa racemosa in 2009 [24]. This alkaloid showed low toxicity and a variety of important biological activities already described in the literature, among which it is important to mention the antitumor, growth regulator and the plant root growth stimulant properties. De Souza et al. isolated caulerpin from lipoid extract of C. racemosa and its structure was identified by spectroscopic methods, including IR and NMR techniques and demonstrated in vivo and in vitro its anti­nociceptive and anti-inflammatory activities [24].

Microalgae have been also studied in the search of alkaloids, in this sense most of this research has been conducted to identify toxins [78]. The non-sulfated alka­loid toxins of freshwater cyanobacteria (anatoxins and saxitoxin) are all neurotox­ins. The sulfated polysaccharides, C-toxins and gonyautoxins are also neurotoxins, but the sulfated alkaloid cylindrospermopsin blocks protein synthesis with a major impact on liver cells. Some marine cyanobacteria also contain alkaloids (lyngbya — toxins, aplysiatoxins) which are dermatoxins (skin irritants), but have also been associated with gastroenteritis and more general symptoms such as fever [ 78] . Several freshwater bloom forming cyanobacterial genera, including Anabaena, Aphanizomenon, Oscillatoria, and Cylindrospermum, produce the neurotoxin, ana — toxin-a, an alkaloid with a high toxicity to animals [117].

Hydrolysis of particulate organic matter is frequently the rate-limiting step during AD [227, 228]. Mechanical pretreatment of lignocellulose materials increased the hydrolysis and methane yield by 5-25% [229] and reduced the digestion time by 23-59% [230] compared to nontreated samples. Disintegration of WAS in an agita­tor ball mill Model LME 50 K with fine sand balls (0.5-0.9 mm of diameter and

2.7 kg/L of density) increased chemical oxygen demand (COD) solubilization by 25-31% and enhanced biogas yield by 38% compared to nontreated WAS [231]. Mechanical pretreatment of WAS in a high-pressure homogenizer (pressure up to 600 bar) increased biogas production by 18% [195].

Ultrasonic treatment has been widely studied as a method for enhancement of WAS solubilization and methane yield. The minimum energy level required to dis­rupt the activated sludge cell wall is in the range of 1,000-3,000 kJ/kg TS [197, 203] or 20-30 kJ/L [197, 232]. Ultrasonic pretreatment (£=40,000 kJ/kg SS) of WAS resulted in a fivefold increase in the amount of organic matter solubilization but had no influence on biogas yield [199]. Another study showed the same beneficial trend as solubilization increased by 127% at 42 kHz for 120 min [201]. Biogas and meth­ane yields were observed to increase by 20% [201] and by 50% (for biogas) at £s = 6,950 kJ/kg TS [197].

Mechanical pretreatment is required prior to AD of macroalgae and includes chopping (>5 mm), milling (1-5 mm), or homogenization (<1 mm). Grinding of the

Ulva did not influence the total methane yield but it increased the kinetics of hydro­lysis and methane production rate [129] . The grinding of the feedstock is likely more important for continuous stirred-tank reactor (CSTR) or semi-continuous reactors where the HRT is an important operating parameter.

One study examined the influence of mechanical disintegration plus ultrasonic treatments on A. maxima organic matter solubilization, VS reduction, and methane yield [233]. Ultrasonic treatment (Polytron generator PT20ST, from Brinkmann Instruments) increased the soluble COD (sCOD) 3.8-fold compared to freshly har­vested cyanobacteria. A. maxima COD solubilization increased from 21.3 to 76.7%. Surprisingly, VS reduction and methane yield were 0.9 and 0.85 times the values from fresh (not pretreated) biomass, respectively. Only hydrolytic bacteria and aci — dogens benefited from the larger amount of readily degradable substrate. The VFA concentration increased from 12 g/L in the digester with fresh algae to 46 g/L in the digester with pretreated algae. The lack of improvement in the methane yield might be due to the inhibition of methanogenic organisms from the high VFA concentration. Ultrasound pretreatment (19 kHz, treatment energy 1-5 Wh/L) had no influence on the methane yield from homogenized red macroalga Polysiphonia [123]. While the amount of biogas increased by 25-28%, the methane fraction dropped from 42-49 to 33-37%.

An attractive algal cultivation strategy is to use carbon dioxide emitted from power plants for autotrophic biomass assimilation. Such a carbon capture system with microalgae has the following advantages compared to other carbon sequestration technologies [484]:

— Algal system does not require high purity carbon dioxide.

— Algal system produces biofuels that can be used for electric power generation.

— Some flue gas impurities (nitrogen and sulfur oxides) can be removed as well and be used by algae.

A conceptual diagram for algal production and ADP integrated into a scheme with carbon dioxide mitigation and waste treatment processes is shown in Fig. 21.