G.J. Leonard,
Bureau of Sugar Experiment Stations
Brief reference is made to the various options relating to distillery waste disposal. As a more basic approach to the waste problem, investigations into the major organic components of various types of yeast-fermentation wastes have been pursued. From the viewpoint of analytical procedures, waste material resulting from yeast- fermentation of sugar cane molasses presents substantially more problems from
colorant interference than do the residues from the fermentation of mixed or clari- fied juice. Analytical systems have been developed for the assay in waste materials
or polysaccharides, oligomonosaccharides, and non-nitrogenous organic acids. A comparison of distillery wastes of different origins will be presented in terms of per- centage organic matter, saccharide profiles, organic nitrogen content, and non- nitrogenous organic acids.
Introduction
A variety of methods have been proposed for solving the distillery waste problem and there is little doubt that different countries will finally adopt different methods.
For example, one scheme involves concentration of the distillery waste in a retort furnace followed by incineration.1 Other methods involve: aerobic bioconversion of the organic waste into fungal biomass as a source of single cell protein;2 3 anaerobic fermentation for the production of methane.4
As a more basic approach to the waste problem, we have aimed our initial research program at developing analytical systems to allow a fast and accurate monitor of the major organic components. In addition, we are interested in measuring the rate of change of major organic components when waste materials are undergoing microbiological transformation. With this approach we expect to predict, more quickly and more precisely at the specific molecular level, the per- formance of a relevant microbiological process.
Methodology for the assay of major organic components in distillery waste It is our objective to apply suitably developed analytical systems to a number of different types of distillery waste materials, reflecting different substrates (e.g.
based on molasses or sugar cane mixed or clarified juice) a n d / o r significantly dif- ferent fermentation conditions during the production of ethanol.
It is important, of course, that adequate reproducibility of analysis of any specific type of waste material can be obtained over a period of time covering a number of production runs. In this regard some knowledge of the variability of the substrate (molasses, mixed or clarified juice) composition is also necessary.
In our experimental work we have given a lot of attention to colorants, but mainly as interferences in various analytical procedures. Colorants which occur in sugar cane products, particularly molasses, can be classified according to the following origins:
• plant pigments and polyphenolic compounds
• caramels which are produced by thermal degradation and condensation reactions of sugars
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• alkaline degradation and condensation products from reducing sugars
• melanoidins formed from sugar-amino acid reactions via the maillard reaction.
Such colorants thus exhibit a variety of chemical characteristics, e.g. some can be separated on a molecular size basis, dialysis or gel permeation chromatography;
many can be adsorbed onto a variety of adsorbants, such as Amberlite XAD poly- styrene or activated charcoal.
In future times we expect to develop suitable analytical systems to allow fast and accurate quantitation of colorants in distillery waste materials.
Analytical systems for the assay of various saccharides in distillery waste
Our investigations thus far have related to the water-soluble fraction of distillery residue materials. The following procedures have been applied to yeast-fermentation residues resulting from commercial distilleries utilising molasses and from a lab- oratory fermenter utilising mixed and clarified juices.
An outline of a clean-up procedure prior to the application of any chromato- graphic fractionations is shown below:
• removal of insoluble material by centrifugation
• salt removal by ion-exchange resins
• colour removal by a combination of ion-exchange and adsorption
In the case of the laboratory fermenter wastes resulting from mixed or clarified juices, complete decolourisation was achieved by use of a macroporous anion ex- changer and Amberlite XAD polystyrene. However, distillery wastes (resulting from molasses) could not be completely decolourised by the above technique. The residual colorant was found to be predominantly low molecular weight and thus could be re- moved by charcoal adsorption. Since charcoal is known to irreversibly adsorb some polysaccharides, such polymeric components were first separated from the residual colourants and low molecular weight carbohydrates by dialysis.
Chromatographic fractionations: mostly a high performance liquid chromato- graphy (HPLC) system was used. Two different HPLC columns were found to be satisfactory for this particular application:
• Aminex Q15S (CA+ + form) stainless steel column run at an elevated tempera- ture. This column gave adequate separations of polymer, oligo- and mono- saccharides; many of the monosaccharides could be separated from each other.
• 'Dextrose'-column (developed by Waters Associates in Australia) was used for the sub-fractination of the oligosaccharide component, when necessary. This column is a radial compression type, and was run in a radial compression module at room temperature in aqueous medium.
For most of the HPLC work a differential RI detector and a flow-through carbohydrate detector (an autoanalyser version of the phenol-sulphuric colorimetric assay) were used.
When a sub-fractination of the polysaccharides was required, an appropriate porosity gel permeation column was used in conjunction with the flow-through carbohydrate detector. As a rough guide to the total carbohydrate content of a dis- tillery waste sample, two additional techniques were used: gas chromatography (of aldononitrile acetates) was performed after acid hydrolysis of the sample; the soluble fraction was also analysed for total carbohydrate by use of an autoanalyser (phenol-sulphuric colorimetric assay).
An analytical system for the assay of non-nitrogenous organic acids in distillery wastes
The water-soluble fractions of the same types of distillery residues (referred to in 2.1) were analysed for organic acids.
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Sample clean-up: In this case we sought to remove colorants without the use of ion exchangers and therefore without desalting the sample. Polystyrene XAD resin alone was not adequate while activated charcoal could not be used because of losses of organic acids onto the charcoal. Sep-Pak C18 (Waters Associates) however, proved adequate when subsequent chromatographic fractionations were monitored and quantified by use of the RI detector.
Chromatographic fractionation by H P L C on u-bondpak C18 column, using experimental conditions, similar to those described by Coppola, Conrad and Cotter,5 was adequate for our purposes.
A comparison of different types of distillery wastes in terms of major organic components
The methodology described above has been applied to one set of yeast-fermentation residue types. However, an examination of reproducibility aspects has not been completed and hence a set of preliminary figures only will be presented at this work- shop.
REFERENCES
1. Anon., Sugary Azucar 72, 1977, 10.
2. Araujo, N., Visconti, A.S., etal, Brazil Acucareiro 88, 1976, 479-489.
3. Chuang, Y-T, Hwang, P-T, etal, Taiwan Sugar Res. Instit. Rpt # 81, 1978, 35-45.
4. Chuang, C.L., Sang, S.L., etal, Taiwan Sugar24, 1917, 268-272.
5. Coppol, E.D., Conrad, E.C., Cotter, R., J.A.O.A.C. 61, 1978, 1490-1492.
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CONVERSION OF ETHANOL TO HIGH OCTANE MOTOR SPIRIT
T. Mole and J.A. Whiteside,
Catalysis and Surface Science Laboratory, Division of Materials Science, CSIRO
ZSM5 zeolite (the Mobil catalyst) has a unique capacity to catalyse the conversion of a wide range of gaseous and liquid organic materials to liquid hydrocarbons.
Particular interest has been attached to the conversion of synthesis-gas-derived methanol to motor spirit.
The possibility of applying ZSM5 catalyst to ethanol and aqueous ethanol has been examined. Ethanol is a less suitable reactant than methanol, and gives mainly ethylene under mild reaction conditions. Under more drastic conditions some 40 per cent of the carbon content of ethanol is converted to aromatic hydrocarbons, which represent high octane components of motor spirit. Most of the residual carbon con- tent is converted into light-hydrocarbons (C2-C4), which can in turn be converted into aromatic hydrocarbons under comparable conditions using 'dry' ZSM5 catalyst.
The conditions for ZSM5-catalysed ethanol conversion are outlined. The relative merits of ethanol and synthesis gas as petrochemical feedstocks will be discussed briefly.
The provision of substitute liquid fuels is highly dependent upon the production and use of catalysts. Where the substitute liquid fuel is ethanol, the catalytic processes are enzymic. More commonly the catalysts and catalytic processes are artificial rather than natural, and of a heterogeneous nature.
CSIRO's Division of Materials Science has a substantial and wide ranging program of research into the application of heterogeneous catalysts to the produc- tion of substitute liquid fuels. The Department of National Development and Energy contributes to the support of some parts of this research program, including the work presented in the present paper.
Such an important problem as the need for substitute liquid fuels presents oppor- tunities for innovative research and discovery. The outstanding innovation of the last thirty years has been the discovery by Mobil Oil of ZSM5 zeolite.
ZSM5 zeolite is a most remarkable material and may well have as great an in- fluence on the production of liquid fuels as the introduction of catalytic cracking or catalytic reforming. It has the capacity to convert a wide range of gaseous and liquid organic materials into hydrocarbons. Under suitable circumstances much of the hydrocarbon lies in the boiling range of motor spirit and much consists of branched- chain and benzenoid-aromatic compounds.
Thus methanol can be converted quite simply to motor spirit of ca. 90 octane number in a yield which is about 80 per cent of the theoretical. Methanol can be pro- duced from synthesis gas by well established and relatively simple technology, and synthesis gas is readily obtained by steam reforming of methane or by the action of steam and oxygen on coal or coke. Thus the Mobil catalyst opens up a most attrac- tive route to substitute motor spirit.
Methanol is a relatively undesirable fuel for direct use because of its low calorific value (of 32g. methanol only the 14g CH2 group burns) and because it is poorly mis- cible with petrol at low temperatures and under moist conditions. Direct use of ethanol is not greatly limited by these factors, but does conflict with the normal 142
practice of restriction and taxation of the production, sale and consumption of ethanol.
Little information is available on the applicability of ZSM5 zeolite to ethanol con- version. Initial CSIRO experiments by Drs Anderson and Rajadhyaksha showed that ethanol is a much more difficult reactant than methanol. Unless the catalyst is adequately activated, the ethanol largely undergoes simple dehydration to ethylene;
furthermore the catalyst life is very limited. Fortunately catalyst activation can be achieved by simple treatment with hot dilute mineral acid.
We have now made a preliminary assessment of the possibility of converting ethanol and aqueous ethanol to hydrocarbons over ZSM5 catalyst. The experimental method used in the present work is extremely simple. Ethanol or aqueous ethanol, fed by motorised syringe into a slow stream of nitrogen, is carried through a bed of 3/20g. of catalyst in a tubular microreactor, which can be main- tained at temperatures in the range 300-500°C.
The nitrogen flow rate is 2.2 mL/min; thus the apparent contact time is <5 sec.
The ethanol feed rate is 0.8 g/hour; that is the catalyst is fed with five times its own weight of ethanol per hour to produce three times its own weight of hydrocarbons.
The conversion of ethanol is complete in all our experiments.
The hydrocarbons produced are diluted with nitrogen and analysed by a gas chromatograph. The results obtained are summarized in the accompanying charts, which show the weight percentage composition of the reaction products as a func- tion of on-stream time. The data is not extensive and no attempt has been made to optimise the catalyst or the conversion conditions. The data should nevertheless indicate the technical possibilities. Our results show that at temperatures of 380°C and above the main products are C6-9 benzenoid hydrocarbons of the series: ben- zene, toluene, xylenes, and trimethylbenzenes. Of these, toluene and xylenes are dominant. The other important products are C3 hydrocarbons (mainly propane) and C4+ aliphatic hydrocarbons (mainly isobutane and n-butane, with smaller amounts of C5 and C6 hydrocarbons).
The C4+ aliphatic hydrocarbons and the C3 hydrocarbons can both be converted to C6-9 aromatic hydrocarbons over ZSM5 catalyst. I am not at liberty to discuss with you our work on this topic, but I can assure you that dry C3 gases and particu- larly dry C4 gases will give yields of around 50 per cent of C6.9 aromatics. Thus an overall 65-70 per cent conversion of ethanol to C6-9 aromatic hydrocarbons is feasible.
Turning back to the primary ethanol conversion, the results presented suggest a conversion temperature of 380-420° and an on-stream time of up to an hour at an ethanol feed rate of about 5 grams per gram of catalyst per hour. Under these circumstances, a fixed bed of catalyst could not be practical. A fluidised catalyst bed would be used. Spent catalyst would be withdrawn, regenerated by oxidation in air at about 500 °C, and then returned to the bed (as in catalytic cracking).
The undesirable C10+ hydrocarbons make up only about 5 per cent of the ethanol conversion product, as does ethylene. The C6-9 aromatic hydrocarbons which are the main products of conversion are prime high-octane components of gasoline and so would be a very acceptable blending stock.
Since ethanol is produced by fermentation as an aqueous solution, particular interest attaches to the possibility of conversion of aqueous ethanol over ZSM5 catalyst. We have examined the products obtained by the action of ZSM5 on 1:1 and 3:1 aqueous ethanol at our standard rate of 5 grams feed per gram catalyst per hour.
The results obtained at 420-460° are quite encouraging and suggest that the catalyst may be able to work on an aqueous ethanol feed.
Notwithstanding the positive results obtained it is clear that the conversion of ethanol to high-octane motor spirit is technically inferior to the conversion of 143
methanol. This must be seen in the perspective that the manufacture of methanol from synthesis gas (derived from natural gas or coal) and Fischer- Tropsch synthesis are amongst the most technically simple and economic (money-wise) sources of sub- stitute liquid fuels in Australia in the short to medium term. On this basis it seems likely that utilisation of ethanol is a prospect for the longer term in which we are concerned about the depletion of our fossil fuel reserves generally rather than the crude oil particularly.
Under these circumstances it is worth remembering that ethanol is a prime source of pure ethylene for petrochemical purposes, and has been used as such in Australia in the past. By contrast synthesis gas and methanol cannot be converted to ethylene selectively. Thus it may be that as natural ethane resources are depleted, we may again turn to ethanol as an ethylene source. Under these circumstances our observa- tions that ZSM5 catalyst will handle aqueous ethanol and that ZSM5 catalyst con- verts ethanol mainly to ethylene at lower. temperatures may be particularly signifi- cant.
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hours ' ' hours
Figure 3. Products from Ethanol at 420 °C Figure 4. Ethylene Formation
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Figure 5. Formation of C6.9 Aromatics Figure 6. C3 Hydrocarbon Formation
Figure 7. Formation of C4 + Aliphatics Figure 8. Products from Aqueous Ethanol (i) 146
Figure 9. Products from Aqueous Ethanol (ii) hours ' Figure 10. Products from Aqueous Ethanol (iii)
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TRANSCRIPT OF DISCUSSIONS, SESSION TWO:
LARGE SCALE PRODUCTION OF FUEL ETHANOL
The aims of the second session of the workshop were to review the prospects for a major fuel ethanol industry by considering current technology, prospective develop- ments in technology, the possibilities for cost and energy savings, efficient treatment and waste disposal. It has not always been possible to identify speakers. Any errors or omissions are regretted.
The session was chaired by Dr H. G. Higgins, Chief of the Division of Chemical Technology, CSIRO.
Mr McNeil, Sugar Research Institute: I am seeking evidence from Dr MacLennan to support his claims that the removal of protein from yeasts for wheat stillage allows the effluent treatment cost component in his proposal to be ignored. I assure you that the removal of these components from molasses or cane stillage results only in a small improvement in the waste quality.
Dr MacLennan, Biotechnology (Aust): The economics that I quoted do in fact include an effluent treatment system. The process certainly doesn't remove all of the effluent, but it's a great deal less than you would get if you left all the yeast plus wheat protein in there. The economics do include a capital cost component for an effluent treatment process.
Mr Strong, Bioenergy (Aust): I'd like to ask a question of Dr Rogers. We saw in Dr MacLennan's paper this afternoon the value of the protein in stillage in terms of viability in the production of ethanol, particularly from grain crops. In the bacterial fermentation process as opposed to yeast fermentation, I'm just wondering if you've been able to determine the suitability, given the bacterium involved, of the stillage protein as an animal food, as opposed to its value in the yeast process?
Dr Rogers, University of N.S.W.: Thanks for that question. At the moment we haven't assessed the nutritional value of Zymomonas. We've done a little bit of work on it, and the protein is fairly high, but we haven't got quantitative data on nucleic acids, proteins, polysaccharides and so on. The only point that's worth making though is that Zymomonas is the causative agent in the production of tequila and palm wines and wines from Indonesia — it certainly doesn't have any problems in terms of pathogenic properties. But it is something that we're working on at the moment.
Dr Doelle, University of Queensland: First a comment on a previous question. I'm pretty sure Zymomonas does have a pathogenic effect. We know this from Mexico:
one has to be very careful with this organism. I would not dispute that eventually we may be able to use it — say when it is killed or dried. You referred to tequila. Of course tequila does not actually contain any of the organism. There can be no doubt that Zymomonas is more dangerous than yeast for human consumption. My question is to Dr Rogers. What about the BOD levels after fermentation? When you ferment and produce alcohol from sucrose using Zymomonas, what sort of effluent would you expect — is it worse or better than with yeast? And also may I add an additional point. Do I understand you right when you said that Zymomonas was equivalent to yeast in alcohol tolerance? Surely not. Zymomonas has only a tolerance at the maximum of around about 10 or 11 per cent alcohol and we have yeasts of 16 per cent to 20 per cent — I even worked with one of 25 per cent alcohol tolerance.
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Dr Rogers, University ofN.S. W.: With the latest Zymomonas we have been working with we can get up to values of 125 — 130 grams per litre, which is about 15-6 per cent volume for volume, and is as good as most yeasts. Some yeasts do have very high alcohol tolerance — the sake yeasts and so on go up to 20 per cent volume for volume alcohol — but they do it very, very slowly. You're talking of a two or three month fermentation period. Remember also that the alcohol tolerance we have at the moment is without mutations to try to get further alcohol tolerance from Zymomonas, I think it's as least as good as yeast if not better.
Dr Doelle, University of Queensland: I don't see why it should not be possible to get to 25 per cent with yeast without genetic engineering.
Dr Rogers, University of N.S.W.: I haven't seen any published data on yeasts showing 25 per cent tolerance achieved in a reasonable period of time. I think most yeast fermentations work to about 10-12 per cent — in fact, I think CSR works with about 9-10 per cent alcohol in fermentation.
Dr Doelle, University of Queensland: One yeast that reaches 16 per cent is known in the German literature.
Dr Rogers, University of N.S. W.: Well without getting into detailed argument on this, I think Zymomonas is comparable to yeast at this point, and has the potential for being better. People have spent decades working on yeast genetics, while Zymomonas is relatively unexplored at the moment. Coming back to your original question on BOD levels — again this is more relevant with commercial substrates, and is an area we are moving into at present. Certainly we found very high efficien- cies of glucose conversion to alcohol, with very few side products. So the BOD would be very low.
Dr Rowlings, CSIRO: A question of clarification addressed to Dr MacLennan. In Table 1 in his paper he has estimated the economics of producing ethanol from wheat. Is he quoting cents per litre ethanol or cents per litre petrol equivalent? How does he establish this?
Dr MacLennan, Biotechnology Australia: Cents per litre of ethanol. I have some difficulty in understanding fully the factor of 1.4 for converting ethanol into petrol equivalent. As I understand it there are disadvantages to ethanol and there are advantages to ethanol. I mentioned this to my colleague from Ampol, Mr Tony Rapson, and I'm quite sure he would like to carry this one for me.
Mr Rapson, Ampol: I wouldn't like to go too much further than to say that this will no doubt be discussed tomorrow when we come to the engine session. While in heat values you have a great discrepancy, you do pick up a little bit in terms of efficiency of use of ethanol in an engine. To pursue that further I suggest that you should attend the session tomorrow on engines, when speakers who are no doubt more expert than I will cover that subject.
Question: The wholesale price you quoted, is that the wholesale price of motor spirits?
Mr Rapson, Ampol: Yes, the wholesale price of motor spirit including excise.
Mr Michael Playne, CSIRO Chemical Technology: My question concerns sweet sorghum and is directed to Dr Harris and Mr Ferguson. Firstly to Dr Harris; have you considered sweet sorghum instead of maize in New Zealand, both being warm temperate subtropical crops; and the question to Mr Ferguson; have you considered 149