WO2015092338A1 - Method for producing ethanol, using sugars with 5 and 6 carbon atoms - Google Patents

Abstract

The invention relates to a method for producing ethanol, using sugars with 5 and 6 carbon atoms.

Description

 PROCESS FOR PRODUCTION OF ETHANOL USING SUGARS OF 5 AND 6 CARBON ATOMS

The present invention relates to a process for the production of ethanol using sugars with 5 and 6 carbon atoms.

The use of fossil resources, especially oil since the advent of the industrial era in the early twentieth century, is accompanied by a continuous and increasing release of many elements in the atmosphere. The direct consequence of these discharges is in particular the increase of the carbon dioxide (CO ₂ ) content in the atmosphere. This rise in the C0 ₂ rate contributes to global warming, which has a clear and lasting impact on the environment.

 The share of oil used in the means of transport is in majority compared to the other uses, that is why many actors of the economic world sought to substitute a part, or even all, of the oil and its derivatives in the fuels intended for the vehicles. Ethanol produced by fermentation of sugars is already a molecule widely used as fuel in many countries and especially in Brazil.

The first-generation fuel ethanol produced today comes almost exclusively from the fermentation of sugars, glucose and sucrose, derived from the agricultural exploitation of the soil. Succulent plants such as sugar beet or sugar cane and starchy plants such as wheat or maize, the main sources of sugars, are plants grown mainly to feed the populations. The main consequence of the boom in ethanol ^era of generation is the competition between fuel and the food industry regarding the destiny of products and sugars in turn competition on land use.

 To overcome this duality, emerged the idea of the use of plants or agricultural co-products not intended for human consumption for the production of biofuels. While first-generation ethanol comes from the fermentation of sucrose and glucose from starch, these "new" raw materials are characterized by high productivity per hectare and a high total sugar content. These sugars consist mainly of glucose derived from cellulose and sugars from hemicelluloses.

Cellulose and hemicelluloses are therefore sources of sugars alternatives to starch and sucrose. This is called second generation ethanol. However, while the strains used for the fermentation of the ^era generation can use glucose from cellulose, they are unable to produce ethanol from hemicellulose composed mainly of xylose and arabinose, five carbon sugars carbon. The proportion of sugars from hemicelluloses is variable from one plant to another but can represent up to 40% of the sugars present in the plant. These are so many sugars not used for the production of ethanol.

 At the industrial level, the processes of producing ethanol using glucose or sucrose as a carbon source almost exclusively use Saccharomyces cerevisiae, the latter being unable to use naturally occurring sugars with five carbon atoms.

 The search for strains capable of transforming pentoses into ethanol began in the 1970s. Scientists mainly focus on xylose because it constitutes between 10 and 40% of the sugars present in lignocellulosic biomass, while arabinose is at most 3% of the lignocellulosic biomass. A strain capable of fermenting xylose and glucose would increase the overall yield of alcoholic fermentation by about 25%.

 There are microorganisms that naturally ferment five-carbon sugars into ethanol. Among these microorganisms there are bacteria that ferment C5 and yeasts that ferment C5.

et une productivité de 0,5 g.L^_1.h^_1 ont été rapportés pour Thermoanaerobacter ethanolicus sur un milieu complexe à 4 g.L^"1 de xylose. Several species of thermophilic bacteria such as Thermoanaerobacter ethanolicus, Clostridium thermohydrosulfuricum and Clostridium thermocellum are capable of carrying out naturally alcoholic fermentation on xylose. A return of

and a productivity of 0.5 gL ^-1 .h ^-1 have been reported for Thermoanaerobacter ethanolicus on a complex medium at 4 gL ^"1 xylose.

 Their use has the advantage of reducing the risk of contamination due to the high temperature used, but the performance is too low for an industrial application.

Other bacteria such as Escherichia coli, Klebsiella oxytoca, Erwinia chrysanthemi can be used for the production of ethanol, but a major disadvantage of bacteria is the high risk of contamination of the medium during fermentation. Bacterial contaminations are common and difficult to control in non-sterile conditions and the implementation of sterile conditions is not always applicable in industrial settings. By using yeasts for fermentation, exogenous bacterial contaminations are more easily controllable. Indeed, high alcoholic titles are unfavorable to bacteria and it is easier, thanks to the fermentation conditions, in particular the pH, to prevent the appearance of a bacterial contamination.

 The alternative to the use of bacteria for the production of ethanol at the industrial stage is therefore the implementation of fungal organisms: yeasts.

 Several yeast species, naturally fermenting xylose, have been identified as the most effective: Pichia stipitis, Candida shehatae, Candida tropicalis and Pachylosen tannophilus.

 a: yield based on the consumed xylose C. sh. - Candida sheliaiae

 b: maximum productivity P. st. = Pichia stipitis

 c: maximum concentration P. ta. - Paehysalen tannophilus

 Table 1: Fermentative performance of three yeasts on xylose Today many strains metabolizing pentoses exist, but these laboratory yeasts are not adapted to industrial conditions, because they do not tolerate the presence of inhibitors present in lignocellulose hydrolysates . In addition, the construction of genetically modified yeast strains capable of effectively converting xylose and arabinose into ethanol is a difficult goal, as it involves modifying yeast metabolic fluxes and energy equilibria.

 In addition, when the metabolizing strains, in addition to the hexoses, the pentoses, ferment said hexoses and pentoses to give ethanol, said strains, in the presence of both types of sugars, always consume the C6 sugars first and then only the C5 sugars, whether natural or genetically modified.

This mechanism, called diauxie, results in a C5 / C6 co-fermentation with the presence of C5 at the end of fermentation which, because of the presence of alcohol and the conditions rather unfavorable culture for the strain (long residence time in the presence of alcohol, possibility of presence of organic acids produced during fermentation by a contaminating flora), will not be consumed by the strain. This results in a loss of sugar in the wine going to the ethanol isolation stage, especially by distillation.

 Thus, an object of the present invention is to provide a method for optimal use of sugars from cellulose and hemicelluloses from lignocellulosic biomasses or agro-industrial co-products to produce ethanol.

 The subject of the invention is therefore the use of a propagation must comprising a microbial population for preparing ethanol from a raw material generating at least one pentose, in particular xylose and / or arabinose. , and at least one hexose, in particular glucose, by fermentation of said pentose and hexose by said microbial population,

 said raw material being extracted to give an extraction juice containing said pentose, in particular xylose and / or arabinose, and a pomace, said pomace giving, after being liquefied, a liquefied, said liquefied being a source of the above hexose, in particular glucose,

 said propagation must being obtained by proliferation of an inoculum of said microbial population with the aid of said extraction juice, in the presence of air,

 said propagation must containing no ethanol,

 said microbial population being used at a time:

for the preparation of a first source of ethanol, by fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice by said microbial population and

 for the preparation of a second source of ethanol, by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied by said microbial population, said ethanol being isolated from the first and / or second sources of ethanol, particularly by distillation of said first and / or second sources of ethanol.

 Said microbial population is therefore used to ferment pentose and then hexose, obtained by hydrolysis of a lignocellulosic biomass.

In addition, said microbial population is obtained by growth on pentose, and is used to ferment the rest of pentose and hexose. Fermenting the pentose of a stream A before fermenting the hexose of a stream B makes it possible to integrate the fermented stream A into the stage of preparation of the stream B, which is of economic interest.

 On the other hand, fermenting the hexose of a stream B before fermenting the pentose of a stream A simply amounts to mixing the streams A and B after their fermentation, which does not allow any particular savings.

 Thus, as illustrated by FIG. 2 relating to a use according to the present invention, the raw material, in particular a lignocellulosic biomass, is extracted after having been in particular impregnated and pretreated.

 Said extraction makes it possible to obtain a liquid rich in pentoses (juice rich in sugars

C5) and a solid rich in hexoses (marc rich in C6 sugars).

 Said liquid allows the propagation of microorganisms, in particular yeasts: it is a step of proliferation of said microorganisms, mainly on the C5 sugars contained in said liquid.

 After propagation, said microorganisms ferment the pentoses of said liquid into alcohol, obtained in the form of wine (derived from C5 sugars) corresponding to a first source of ethanol.

 Said solid is subjected to liquefaction (to obtain a liquefied) and to a fermentation using the microorganisms obtained after the propagation and / or those isolated after the fermentation of the pentoses, to obtain the second source of ethanol (final wine) .

 The second source of ethanol, and possibly the first source of ethanol, is (are) processed, in particular distilled to give ethanol. Indeed, said first source of ethanol can be distilled with the second source of ethanol and / or used during liquefaction.

 The liquid (clarified vinasse) isolated by clarification of the distillation residue (raw vinasse) is optionally joined to said raw material during the extraction.

 Interestingly, the sugars with five carbon atoms that are notably xylose and arabinose are used for the production of yeast cells in the presence of air, then for the production of ethanol, thus preserving the easily fermentable sugars that are sugars with six carbon atoms and in particular glucose for the production of ethanol.

The separation of the C5 and C6 streams and their use in two distinct stages make it possible to maximize the effectiveness of the sugars for the production of ethanol, in particular by limiting the diauxis phenomena. In addition, with the use of a single strain of microorganism capable of both growth on C5, ferment C5 to ethanol and then ferment C6 ethanol, it is no longer necessary to have two fermental infrastructures (one for the fermentation of C5 using a first strain, the other for the fermentation of C6, with the aid of a second strain) and as a result, the cross contamination of two C5 ethanol and C6 ethanol production lines are non-existent.

 By "propagation must" is meant a composition comprising a proliferated microbial population and a culture medium appropriate to said microbial population.

 "Appropriate culture medium" means an aqueous solution containing nutrients, in particular a source of carbon, a source of nitrogen, a source of trace elements and a source of growth factors necessary for metabolism and proliferation. of said microbial population.

 Said culture medium comprises said extraction juice containing pentose, in particular xylose and / or arabinose, as well as a residual glucose content of less than 10 g / l.

 The propagation wort thus comprises the microbial population having proliferated and the culture medium at the end of the growth step of said microbial population, that is to say an aqueous solution in which the nutrients not consumed are present. by said microbial population and metabolites possibly excreted by the strain, said solution comprising said extraction juice.

 By "microbial population" is meant a set of microorganisms of the same strain, the mass of said set being likely to change during its use.

 By "raw material generating at least one pentose and at least one hexose" is meant a raw material capable of providing, after transformation (s), said pentose and said hexose in a form permitting their assimilation by said microbial population.

 By "extraction" is meant the action of mixing said raw material with an extraction solvent, in particular water, said mixture being followed by a liquid / solid separation: this is called liquid / solid extraction

 By "extraction juice" is meant the liquid obtained at the end of said liquid / solid separation relative to said extraction. From 50 to 98% of the sugars with five carbon atoms that can be generated by said raw material are in said extraction juice. Indeed, the majority of pentoses, resulting from the destructuration of hemicellulose chains, migrate into the extraction solvent, in particular water.

Said extraction juice contains from 1 to 50% by weight of C5. By "marc" is meant the solid obtained at the end of said liquid / solid separation relative to said extraction. From 80 to 99% of the six-carbon sugars likely to be generated by said raw material are in said pomace, mainly in the form of cellulose present in said pomace. In fact, cellulose remains largely insoluble and is concentrated in the solid fraction that is marc.

 Said liquid-solid separation is for example carried out on packet press or on a belt press.

By "liquefaction" is meant an operation of first hydrolysis of the marc making it possible to reduce the viscosity of said marc with a value of between 40,000 and 25,000 Pa.s at a value of less than 100 Pa.s. (AR Instrumentation Rheometer) , 33 ° C, speed ls ^" geometry: 40 mm plane, 1 mm gap", desirable for the subsequent fermentation of hexose, in particular glucose, generated by said liquefied. less than or equal to this value allows a better availability of water for the microbial population, to avoid the formation of foam, and to limit the need for agitation.

 Said liquefaction is in particular carried out in the presence of lignocellulolytic enzymes, said enzymes allowing a first hydrolysis of said marc, said enzymes being more particularly ligno-cellulolytic enzymes contained in commercial products such as Celic Cetec 2 (Genencor®) or Celluclast (Novozymes®).

 By "inoculum" is meant a preculture medium comprising a microbial population and a culture medium appropriate to said microbial population, said preculture medium for inoculating a culture medium as described above to obtain said fermentation must.

By "the wort of propagation does not contain ethanol" is meant that said wort of propagation contains less than about 1 gl ^"1 of ethanol, or about 1 g.kg ^{" 1} of ethanol, in particular less than about 0.5 g.kg- ¹ of ethanol.

In particular, the "propagation must contains no ethanol" means a propagation must containing less than about 1 g.kg- ¹ of ethanol, in particular less than about 0.5 g.kg ^"1 ethanol when said wort is obtained by growth of a microbial inoculum of said population in which the partial pressure of 0 ₂ is maintained at more than 20%, in particular more than 30%>.

Even more particularly, "propagation wort does not contain ethanol" means a wort containing less than about 1 g.kg- ¹ ethanol, particularly less than about 0.5 g. kg ^"1 of ethanol, where the said must is obtained by proliferation, during 48 hours, of an inoculum of said microbial population, at a temperature of between 20 and 40 ° C., in particular at a temperature of approximately 30 ° C. and at a pH of between 4 and 6, in particular about 5.5, under partial pressure at 0 ₂ maintained at more than 20, in particular more than 30%.

 By "ethanol source" is meant a liquid or solid-liquid suspension containing ethanol.

 By "first source of ethanol" is meant a source of ethanol obtained by fermentation of pentoses, that is to say sugars with 5 carbon atoms.

 By "second source of ethanol" is meant a source of ethanol obtained by fermentation of hexoses, that is to say sugars with 6 carbon atoms.

 In particular, said first and / or second sources of ethanol are substantially free of micro-organisms from said microbial population.

 According to an advantageous embodiment, the invention relates to the use of a propagation wort as described above, in which said raw material is a lignocellulosic biomass, in particular chosen from:

 - cereal straw, said cereals being in particular chosen from wheat, barley, triticale, sorghum and maize,

 - cereal bran, said cereals being in particular chosen from wheat, barley, triticale, sorghum and maize,

 energy crops, chosen in particular from switchgrass, miscanthus and arundo donax, and

 - forest crops, chosen in particular from wood, short rotation coppices, very short rotation coppices, and forest chips.

 According to an advantageous embodiment, said raw material is, prior to said extraction, pretreated. Advantageously it is impregnated and then pretreated. Any means of pretreatment, in particular impregnation and pretreatment, known to those skilled in the art may be suitable, insofar as it can provide a liquid and a separable solid, the liquid containing most of the C5 and the solid the most of the C6.

The impregnation consists in preparing said raw material with a view to its pretreatment by, for example, penetrating into its matrix, in particular lignocellulosic, an acidic or basic chemical catalytic agent, in particular an acidic catalyst, in particular sulfuric acid. The pretreatment of said raw material consists, for example, of steam cooking or explosion in the presence of said acidic or basic catalyst, in particular the acid catalyst, in particular sulfuric acid, having previously impregnated said raw material.

 Said pretreatment allows in particular the hydrolysis of the hemicelluloses contained in said raw material as well as the preparation for the hydrolysis of the cellulose also contained in said raw material.

 The pentoses, resulting in particular from the destructuring of the hemicellulose chains of said raw material, are rendered soluble by the pretreatment, and are thus capable of migrating into the extraction solvent, in particular water, during said extraction.

 According to an advantageous embodiment, the invention relates to the use of a propagation wort as described above, wherein said ethanol is isolated either from the first source of ethanol, or from the second source of ethanol, or first and second sources of ethanol, by distillation of said first and / or second sources of ethanol.

 According to an advantageous embodiment, the invention relates to the use of a propagation must as described above, wherein said liquefaction of said grounds is carried out in the presence of all or part of said first source of ethanol.

 According to a particularly advantageous embodiment, the invention relates to the use of a propagation wort as described above, in which said liquefaction of said marc is carried out in the presence of all or part of said first source of ethanol, said first ethanol source being substantially free of microorganisms from said microbial population.

 Indeed, the yeasts can be separated from said first source of ethanol before recycling, because of their intolerance to the temperatures used in liquefaction.

 The fact of devoting part of said first source of ethanol to liquefaction makes it possible in particular to reduce the total volume of the vat room required for the second fermentation.

According to an advantageous embodiment, the invention relates to the use of a propagation wort as described above, wherein said distillation, in addition to said ethanol, generates a residue, which contains substantially no ethanol, all or part of the soluble elements, water and insolubles is added to said raw material during said extraction of said raw material. The recycling of all or part of said soluble elements during said extraction makes it possible in particular to minimize the amount of extraction solvent, in particular water, used during said extraction.

 Said residue comprises in particular the first and / or second sources of ethanol, exhausted in alcohol, non-sugars and the microbial population at the end of the second fermentation.

 According to an advantageous embodiment, the invention relates to the use of a propagation must as described above, in which:

 said liquefaction of said grounds is carried out in the presence of all or part of said first source of ethanol,

 said distillation, in addition to said ethanol, generates a residue, which contains substantially no ethanol, all or part of the soluble elements, water and insolubles is added to said raw material during said extraction of said raw material .

 The flows described hereinafter have been numbered according to Scheme 3, having use according to the present invention.

 According to an advantageous embodiment, the invention relates to the use of a propagation wort (13) as described above, comprising a microbial population for preparing ethanol from a raw material generating at least one pentose , in particular xylose and / or arabinose, and at least one hexose, in particular glucose, by fermentation of said pentose and hexose by said microbial population,

 said raw material being extracted to give an extraction juice (9) containing the aforesaid pentose, in particular xylose and / or arabinose, and a pomace (10), said pomace giving, after being liquefied, a liquefied (23), said liquefier being a source of the aforesaid hexose, in particular glucose,

 said propagation must (13) being obtained by proliferation of an inoculum of said microbial population with the aid of said extraction juice (11), in the presence of air,

 said propagation must containing no ethanol,

 said propagation must (13) is subjected to solid / liquid separation to give a propagation juice (16) and a fresh microbial population (17),

 said fresh microbial population (17) is separated into two parts, to obtain a first portion (17a) and a second portion (17b) of said fresh microbial population (17),

said first portion of said fresh microbial population (17a) is used for the preparation of a fermentation must (18) by fermentation of the aforesaid pentose, in particular of xylose and / or arabinose, contained in said extraction juice (12), by said first portion of the fresh microbial population (17a), said fermentation wort (18) giving after liquid / solid separation said first source of ethanol (19) and all or part of the recyclable microbial population (20),

 said second source of ethanol (24) is obtained by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied (23), by all or part of said recyclable microbial population (20) and said second part of the population fresh microbial (17b).

 Said liquid / solid separation making it possible to obtain the first source of ethanol and a recyclable microbial population is such that said first source of ethanol is substantially free of microorganisms originating from said microbial population.

 By "substantially free of microorganisms" is meant that the microorganism concentration of said first source of ethanol (19) is less than 10% of that of said fermentation must (18).

 According to an advantageous embodiment, the invention relates to the use of a propagation wort as described above, said wort of propagation containing no ethanol, wherein said microbial population comprises yeasts of Candida shehatae strain, Pichia stipitis, Pichia guilliermondii, Candia tropicalis or Pachysolen tannophilus, said microbial population including in particular yeasts of Candida shehatae strain.

 According to an advantageous embodiment, the invention relates to the use of a propagation wort (13) as described above, comprising a microbial population comprising yeasts of Candida shehatae, Pichia stipitis, Pichia guilliermondii, Candia tropicalis or Pachysolen strain. tannophilus, in particular Candida shehatae, for preparing ethanol from a lignocellulosic biomass generating at least one pentose, in particular xylose and / or arabinose, and at least one hexose, in particular glucose, by fermentation said pentose and hexose by said microbial population,

 said lignocellulosic biomass being extracted to give an extraction juice (9) containing the aforesaid pentose, in particular xylose and / or arabinose, and a pomace (10), said pomace giving, after being liquefied, a liquefied (23), said liquefier being a source of the aforesaid hexose, in particular glucose,

 said propagation must (13) being obtained by proliferation of an inoculum of said microbial population with the aid of said extraction juice (11), in the presence of air,

said propagation must containing no ethanol, said microbial population being used at a time:

 for the preparation of a first source of ethanol (19), by fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12) by said microbial population and

 for the preparation of a second source of ethanol (24), by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied by said microbial population,

 said ethanol being isolated from the first (19) and / or second (23) sources of ethanol by distillation of said first and / or second sources of ethanol,

 said liquefaction of said grounds (10) being carried out in the presence of all or part of said first source of ethanol (19),

 said propagation mash (13) being subjected to solid / liquid separation to give a propagation juice (16) and a fresh microbial population (17),

 said fresh microbial population (17) being separated into two parts, to obtain a first portion (17a) and a second portion (17b) of said fresh microbial population,

 said first part of said fresh microbial population (17a) being used for the preparation of a fermentation broth (18) by fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12), by said first portion of the fresh microbial population (17a), said fermentation wort (18) giving after said liquid / solid separation said first source of ethanol (19) and all or part of the recyclable microbial population (20). )

 said second source of ethanol (24) is obtained by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied, by all or part of said recyclable microbial population (20) and said second part of the fresh microbial population ( 17b).

According to an advantageous embodiment, the invention relates to the use of a propagation wort (13) as described above, comprising a microbial population comprising yeasts of Candida shehatae, Pichia stipitis, Pichia guilliermondii, Candia tropicalis or Pachysolen strain. tannophilus, in particular Candida shehatae, for preparing ethanol from a lignocellulosic biomass generating at least one pentose, in particular xylose and / or arabinose, and at least one hexose, in particular glucose, by fermentation said pentose and hexose by said microbial population, said lignocellulosic biomass being extracted to give an extraction juice (9) containing the aforesaid pentose, in particular xylose and / or arabinose, and a pomace (10), said pomace giving, after being liquefied, a liquefied (23), said liquefier being a source of the aforesaid hexose, in particular glucose,

 said propagation must (13) being obtained by proliferation of an inoculum of said microbial population with the aid of said extraction juice (1 1), in the presence of air,

 said propagation must containing no ethanol,

 said microbial population being used at a time:

 for the preparation of a first source of ethanol (19), by fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12) by said microbial population and

 for the preparation of a second source of ethanol (24), by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied by said microbial population,

 said ethanol being isolated from the first (19) and / or second (23) sources of ethanol by distillation of said first and / or second sources of ethanol,

 said distillation, in addition to said ethanol, generating a residue (26), which contains substantially no ethanol, all or part of the soluble elements, water and insolubles is added to said bio lignocellulosic mass during said extraction of said lignocellulosic biomass,

 said liquefaction of said grounds (10) being carried out in the presence of all or part of said first source of ethanol (19),

 said propagation mash (13) being subjected to solid / liquid separation to give a propagation juice (16) and a fresh microbial population (17),

 said fresh microbial population (17) being separated into two parts, to obtain a first portion (17a) and a second portion (17b) of said fresh microbial population,

said first part of said fresh microbial population (17a) being used for the preparation of a fermentation broth (18) by fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12), by said first portion of the fresh microbial population (17a), said fermentation wort (18) giving after said liquid / solid separation said first source of ethanol (19) and all or part of the recyclable microbial population (20). ) said second source of ethanol (24) is obtained by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied, by all or part of said recyclable microbial population (20) and said second part of the fresh microbial population ( 17b).

 Since said first source of ethanol is substantially free of microorganisms from said microbial population, said liquefaction of said pomace is carried out in the absence of microorganisms from said microbial population.

 According to an advantageous embodiment, the invention relates to the use of a propagation wort as described above, wherein said extraction juice containing said pentose, in particular xylose and / or arabinose, is detoxified prior to said proliferation and the preparation of said first source of ethanol.

 The detoxification treatment allows a reduction of the fermentation inhibitors that are in particular 5-HMF, furfural and especially the degradation products of lignin.

 Said detoxification is in particular:

 an alkaline treatment, more particularly with calcium hydroxide or with ammonia,

 the addition of acetaldehyde, to reduce the lag phase and the yeast mortality rate,

 an extraction of the inhibitors on ion exchange resin or on activated carbon,

 biological and enzymatic treatments,

 extractions with ether or ether acetate, or

 a treatment by adding gas or vapor into the product ("stripping").

According to an advantageous embodiment, the invention relates to the use of a propagation wort as described above, in which said raw material is subjected, after said impregnation and said pretreatment, to an additional hydrolysis treatment. .

 Said additional hydrolysis treatment follows the pretreatment allowing in particular the hydrolysis of the hemicelluloses contained in said raw material.

 Said additional hydrolysis treatment is in particular thermal or enzymatic, and allows the hydrolysis of C5 oligomers to monomeric sugars.

According to an advantageous embodiment, the invention relates to the use of a propagation wort as described above, in which said liquefier is hydrolyzed before said fermentation of the aforesaid hexose, in particular glucose, generated by the hydrolysis of said liquefied material.

 Said hydrolysis of said liquefied thus constitutes a second hydrolysis, the first being the liquefaction, that is to say a first hydrolysis, of said grounds, to give said liquefied.

 The microbial population is thus brought into contact with the aforesaid hexose, generated by the hydrolysis of said liquefied: this type of fermentation decoupled from hydrolysis is called: hydrolysis and separate fermentation (SHF: separate saccharification and fermentation).

 Said hydrolysis is in particular carried out in the presence of lignocellulolytic enzymes, more particularly ligno-cellulolytic enzymes contained in commercial products such as Celic Cetec 2 (Genencor®) or Celluclast (Novozymes®).

 In particular, said hydrolysis makes it possible both to convert the cellulose of said liquefier into cellobiose, then into hexose, more particularly into glucose.

 According to an advantageous embodiment, the invention relates to the use of a propagation must as described above, in which said liquefied acid is hydrolysed, said hydrolysis and said fermentation of the aforesaid hexose, in particular glucose, generated by the hydrolysis of said liquéfiât being concomitant. This type of fermentation coupled with hydrolysis is called Simultaneous Saccharification and Fermentation (SSF: Simultaneous Saccharification and Fermentation).

 Thus, said microbial population is brought into contact with the liquefied, during the hydrolysis of said liquefied.

 According to an advantageous embodiment, the liquefaction of said grounds (that is to say a first hydrolysis of said grounds) to obtain said liquefied and the hydrolysis of said liquefied (that is to say a second hydrolysis) are carried out by the action of the same enzymes, in particular with cellulolytic action.

 According to another advantageous embodiment, the liquefaction of said grounds (that is to say a first hydrolysis of said grounds) to obtain said liquefied and the hydrolysis of said liquefied (that is to say a second hydrolysis) are carried out by the action of the same enzymes, in particular cellulolytic action, said microbial population being brought into contact with the liquefied, during the hydrolysis of said liquefied.

The invention also relates to a process for preparing ethanol from a raw material generating at least one pentose, in particular xylose and / or arabinose, and at least one hexose, in particular glucose, comprising: a) a step of extracting said material, to obtain an extraction juice containing said pentose, in particular xylose and / or arabinose, and a pomace,

b) a step of liquefying said coffee obtained in the preceding step, to obtain a liquefied, said liquefied being a source of the aforesaid hexose, in particular glucose,

c) a step of proliferation of an inoculum of a microbial population using said extraction juice containing said pentose, in particular xylose and / or arabinose, in the presence of air, to obtain a propagation must comprising said microbial population, said propagation wort containing no ethanol,

d) a first fermentation step of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice, by said microbial population, to obtain a first source of ethanol,

e) a second fermentation step of the aforesaid hexose, in particular glucose, generated by said liquefied by said microbial population, to obtain a second source of ethanol, f) a step of isolating said ethanol from the first and / or second sources of ethanol, in particular by distillation of said first and / or second sources of ethanol.

During said extraction, the liquid-solid separation is for example carried out on a package press, in particular when the mass of raw material to be treated is of the order of about ten kg, or on a belt press, in particular when the mass of raw material to be treated is of the order of a hundred kg.

 The extraction provides an extraction juice whose composition varies according to the raw material treated and pretreatment. An example of composition is given in the example below. These modes of preparation and the juices obtained are widely described in the literature.

 The liquid-solid separation is preferably carried out on a belt press.

 According to an advantageous embodiment, the temperature of the wort varies from about 20 ° C to about 40 ° C, in particular from about 28 to about 32 ° C.

 In another advantageous embodiment, the pH of the wort is controlled to vary from about 4 to about 7, in particular from about 5 to about 6.

 According to another advantageous embodiment, the wort of propagation is such that the oxygen partial pressure in said wort is greater than or equal to about 20%, in particular greater than or equal to about 30%.

By oxygen partial pressure is meant the oxygen concentration (0 ₂ ) dissolved in the liquid phase. This partial pressure is expressed as a% of the saturating concentration in oxygen in the working conditions considered, such as, but not limited to, temperature, pressure, agitation.

 According to an advantageous embodiment, the temperature during said first fermentation varies from about 20 ° C to about 40 ° C, in particular from about 28 to about 32 ° C.

 In another advantageous embodiment, the pH at said first fermentation is controlled to vary from about 4 to about 7, in particular from about 5 to about 6.

 According to another advantageous embodiment, the oxygen partial pressure during said first fermentation is about 0%.

 This oxygen partial pressure of about 0% can in particular be obtained despite an aeration flow of about 0.2 VVM.

 According to an advantageous embodiment, the temperature during said liquefaction ranges from about 25 to about 75 ° C., in particular from about 40 to about 60 ° C., the temperature being more particularly about 50 ° C.

 According to an advantageous embodiment, the temperature during said second fermentation varies from about 20 ° C. to about 40 ° C., in particular from about 28 ° to about 35 ° C.

 According to another advantageous embodiment, the pH during said second fermentation is regulated to vary from about 4 to about 7, in particular from about 5 to about 6.

 According to another advantageous embodiment, the oxygen partial pressure during said second fermentation is about 0%.

 This oxygen partial pressure of about 0% can in particular be obtained despite a ventilation rate of about 0.4 VVM.

 According to an advantageous embodiment, said liquefaction is carried out in the presence of lignocellulolytic enzymes, at a concentration varying from approximately 5 to approximately 500 mg / g of cellulose, in particular from approximately 10 to approximately 80 mg / g of cellulose, more particularly at about 40 mg / g of cellulose.

 According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, wherein said raw material is, prior to said extraction, impregnated and pretreated.

According to a particularly advantageous embodiment, said impregnation is carried out in the presence of an acidic or basic chemical catalytic agent, in particular a acid catalyst, especially sulfuric acid, said catalytic agent impregnating said raw material.

 According to another particularly advantageous embodiment, said impregnation is carried out at a temperature of approximately 50 to approximately 80 ° C, in particular of approximately 60 to approximately 70 ° C, more particularly to approximately 65 ° C.

 According to an advantageous embodiment, said pretreatment is carried out by injecting steam at a temperature of about 120 to about 250 ° C., in particular about 170 to about 190 ° C., more particularly to about 180 ° C. and at a pressure of about 5 to about 15 bar, particularly about 8 to about 10 bar, more preferably about 9 bar.

 At the end of this pretreatment step, the lignocellulosic matrix impacted by said pretreatment is then reactive to the liquefaction enzymes. The combination of thermal, chemical and possibly mechanical treatment implies a strong increase in the digestibility of the cellulose.

 According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, in which the ethanol is isolated either from the first source of ethanol, or from the second source of ethanol, or from first and second sources of ethanol, by distillation of said first and / or second sources of ethanol.

 According to an advantageous embodiment, the invention relates to a process for the preparation of ethanol as described above, in which said step of liquefying said grounds is carried out in the presence of all or part of said first source of ethanol obtained from the from the first fermentation step of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice.

 According to an advantageous embodiment, the invention relates to a process for the preparation of ethanol as described above, in which said step of liquefying said grounds is carried out in the presence of all or part of said first source of ethanol obtained from the from the first fermentation step of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice, said first source of ethanol being substantially free of microorganisms from said microbial population.

Indeed, the yeasts can be separated from said first source of ethanol before recycling, because of their intolerance to the temperatures used in liquefaction. According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, wherein said distillation generates, in addition to said ethanol, a residue, which contains substantially no ethanol, all or part of which soluble elements, water and insoluble elements, is added to said raw material during said extraction of said raw material.

 According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, in which:

 said liquefaction of said grounds is carried out in the presence of all or part of said first source of ethanol,

 said distillation, in addition to said ethanol, generates a residue, which contains substantially no ethanol, all or part of the soluble elements, water and insoluble elements, is added to said raw material during said extraction of said raw material.

 According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, wherein said raw material is a lignocellulosic biomass, in particular chosen from:

 - cereal straw, said cereals being in particular chosen from wheat, barley, triticale, sorghum and maize,

 - cereal bran, said cereals being in particular chosen from wheat, barley, triticale, sorghum and maize,

 energy crops, chosen in particular from switchgrass, miscanthus and arundo donax, and

 - forest crops, chosen in particular from wood, short rotation coppices, very short rotation coppices, and forest chips.

The flows described hereinafter were numbered according to Scheme 3, showing a method according to the present invention.

 According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, comprising a microbial population for preparing ethanol from a raw material generating at least one pentose, in particular xylose and / or arabinose, and at least one hexose, in particular glucose, by fermentation of said pentose and hexose by said microbial population,

said raw material being extracted to give an extraction juice (9) containing the aforesaid pentose, in particular xylose and / or arabinose, and a marc (10), said marc giving, after being liquefied, a liquefied (23), said liquefied being a source of the aforesaid hexose, in particular glucose,

 said propagation must (13) being obtained by proliferation of an inoculum of said microbial population with the aid of said extraction juice (11), in the presence of air,

 said propagation must containing no ethanol,

 said propagation must (13) being subjected to a solid / liquid separation to give a propagation juice (16) and a fresh microbial population (17), said propagation wort containing no ethanol,

 said fresh microbial population (17) being separated into two parts, to obtain a first portion (17a) and a second portion (17b) of said fresh microbial population (17),

 said first part of said fresh microbial population (17a) being used for the preparation of a fermentation broth (18) by fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12), by said fresh microbial population, said fermentation wort (18) giving after said liquid / solid separation said first source of ethanol (19) and all or part of the recyclable microbial population (20),

 said second source of ethanol (24) being obtained by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied (23), by all or part of said recyclable microbial population (20) and said second part of the population fresh microbial (17b).

 Said liquid / solid separation making it possible to obtain the first source of ethanol and a recyclable microbial population is such that said first source of ethanol is substantially free of microorganisms originating from said microbial population.

 According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, in which said microbial population comprises yeasts of the Candida shehatae, Pichia stipitis, Pichia guilliermondii, Candia tropicalis or Pachysolen tannophilus strain, said population microbial including in particular yeasts of Candida shehatae strain.

 According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, comprising:

a) a step of extracting a lignocellulosic biomass, to obtain an extraction juice (9) containing the aforesaid pentose, in particular xylose and / or arabinose, and a pomace (10), b) a liquefaction step of said pomace (10) obtained in the preceding step in the presence of all or part of said first source of ethanol (19) obtained at the end of the first fermentation step f), to obtain a liquefied (23), said liquefier being a source of the aforesaid hexose, in particular glucose,

c) a step of proliferation of an inoculat comprising yeasts of Candida shehatae, Pichia stipitis, Pichia guilliermondii, Candia tropicalis or Pachysolen tannophilus strain, in particular Candida shehatae, with the aid of said extraction juice (11) containing the aforesaid pentose, in particular xylose and / or arabinose, in the presence of air, to obtain a propagation must (13) comprising a population of said yeasts, said propagation wort containing no ethanol,

d) a solid / liquid separation step relative to said propagation wort (13), to give a propagation juice (16) and a population of said fresh yeasts (17),

e) a step of separating said fresh yeast population into two parts, to obtain a first portion (17a) and a second portion (17b) of said population of fresh yeasts,

f) a first step of fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12), by said first portion of the population of said fresh yeasts (17a), to obtain after liquid / solid separation a first source of ethanol (19) and a population of said yeasts, recyclable (20), g) a second fermentation step of the aforesaid hexose, in particular glucose, generated by said liquefied (23), by said population of said recyclable yeasts (20) and said second portion of said population of said fresh yeasts (17b), to obtain a second source of ethanol (24),

h) a step of isolating said ethanol from the first (19) and / or second (24) sources of ethanol by distillation of said first and / or second sources of ethanol.

 According to an advantageous embodiment, the invention relates to a process for preparing ethanol as described above, comprising:

a) a step of extracting a lignocellulosic biomass and all or part of the soluble elements, water and insoluble elements, of the residue (26) obtained at the end of the distillation step h), for obtaining an extraction juice (9) containing the aforesaid pentose, in particular xylose and / or arabinose, and a pomace (10),

b) a liquefaction step of said pomace (10) obtained in the preceding step in the presence of all or part of said first source of ethanol (19) obtained at the end of the first step of fermentation f), to obtain a liquéfiât (23), said liquefied being a source of the aforesaid hexose, in particular glucose,

c) a step of proliferation of an inoculat comprising yeasts of Candida shehatae, Pichia stipitis, Pichia guilliermondii, Candia tropicalis or Pachysolen tannophilus strain, in particular Candida shehatae, with the aid of said extraction juice (11) containing the aforesaid pentose, in particular xylose and / or arabinose, in the presence of air, to obtain a propagation must (13) comprising a population of said yeasts, said propagation wort containing no ethanol,

d) a solid / liquid separation step relative to said propagation wort (13), to give a propagation juice (16) and a population of said fresh yeasts (17),

e) a step of separating said fresh yeast population into two parts, to obtain a first portion (17a) and a second portion (17b) of said population of fresh yeasts,

f) a first step of fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12), by said first portion of the population of said fresh yeasts (17a), to obtain after liquid / solid separation a first source of ethanol (19) and a population of said yeasts, recyclable (20), g) a second fermentation step of the aforesaid hexose, in particular glucose, generated by said liquefied (23), by said population of said recyclable yeasts (20) and said second portion of said population of said fresh yeasts (17b), to obtain a second source of ethanol (24),

h) a step of isolating said ethanol from the first (19) and / or second (24) sources of ethanol by distillation of said first and / or second sources of ethanol, said distillation generating, in addition to said ethanol, a residue (26); ), which contains substantially no ethanol.

 Since said first source of ethanol is substantially free of microorganisms from said microbial population, said liquefaction of said pomace is carried out in the absence of microorganisms from said microbial population.

 According to an advantageous embodiment, the invention relates to a process for the preparation of ethanol as described above, in which said extraction juice containing said pentose, in particular xylose and / or arabinose, is detoxified before said proliferation and the preparation of said first source of ethanol.

According to an advantageous embodiment, the invention relates to a process for the preparation of ethanol as described above, in which said raw material makes the object, after said impregnation and said pretreatment, of an additional hydrolysis treatment.

 According to an advantageous embodiment, the invention relates to a process for the preparation of ethanol as described above, in which said liquefier is hydrolysed before said fermentation of the aforesaid hexose, in particular glucose, generated by the hydrolysis of said liquefied material.

 According to an advantageous embodiment, the invention relates to a process for the preparation of ethanol as described above, in which said liquefied acid is hydrolysed, said hydrolysis and said fermentation of the aforesaid hexose, in particular glucose, generated by the hydrolysis of said liquefying being concomitant.

 According to an advantageous embodiment, liquefaction of said grounds (that is to say a first hydrolysis of said grounds) to obtain said liquefied and hydrolysis of said liquefied (that is to say a second hydrolysis) are carried out by the action of the same enzymes, in particular with cellulolytic action.

 According to another advantageous embodiment, the liquefaction of said grounds (that is to say a first hydrolysis of said grounds) to obtain said liquefied and the hydrolysis of said liquefied (that is to say a second hydrolysis) are carried out by the action of the same enzymes, in particular cellulolytic action, said microbial population being brought into contact with the liquefied, during the hydrolysis of said liquefied.

DESCRIPTION OF THE FIGURES

Figure 1 shows the metabolic pathways involved in the use of glucose and xylose by microorganisms.

Figure 2 shows the general diagram of a use or a method according to the invention. Figure 3 shows the detailed diagram of a use or a method of the invention.

In Figure 3, the flows and operations are as follows:

La figure 4 présente la cinétique d'hydrolyse de la cellulose de la paille imprégnée et prétraitée versus paille imprégnée sans acide et prétraitée.

FIG. 4 shows the kinetics of hydrolysis of the cellulose of the impregnated and pretreated straw versus straw treated without acid and pretreated.

EXAMPLE

Unless otherwise indicated, the flows described hereinafter have been numbered according to Scheme 3, showing a method according to the present invention.

Example 1: Application of the process with a strain not producing ethanol in the propagation phase: Candida shehatae:

The substrate used is a wheat straw whose composition is as follows:

 Table 2: Composition of an Experte straw 2011 Impregnation in dilute phase / batch process:

An expert straw of composition as described in Table 2 is impregnated by soaking in a reactor 10001. The soaking is carried out by complete immersion of the straw for 12 hours, after contacting the sulfuric acid solution previously brought to 65 ° C. The formula of the straw mixture and sulfuric acid solution is such that the final dry matter reaches 5%. For the preparation of the sulfuric acid solution, the ratio sulfuric acid (dry weight) per 100 g of final solution is 0.51%. After one night of tempering, the temperature that has not been regulated is only 35 ° C.

The reactor is then emptied of its liquid content, this draining is achieved by the opening of the bottom valve, the output is filled with a selection gate.

At the end of this dripping, the impregnated straw is introduced into the pre-treatment reactor. Pretreatment:

 The impregnated straw is placed in a reactor heated by steam injection at 180 ° C. and kept under pressure at 9 bar for 5 minutes.

 i

The internal temperature measured at the beginning of the reaction is 167 ° C., and 174 ° C. at the end of the reaction.

At the end of this pretreatment step, the reactor is brought back to atmospheric pressure. The lignocellulosic matrix impacted by this pretreatment, is then reactive to liquefaction enzymes. The combination of thermal, chemical and mechanical treatments implies a strong increase in the digestibility of cellulose. Thus, at the end of the liquefaction, 90% of the constituent glucose of the cellulose is available for conversion to ethanol.

FIG. 4 shows the kinetics of hydrolysis of the cellulose of the impregnated and pretreated straw versus straw treated without acid and pretreated.

Extraction of C5:

At the end of the pretreatment, the product is mixed with water (extraction solvent in this example) to allow diffusion and extraction of C5. Once hydrated, the pretreated straw (stream 8) can undergo a liquid-solid extraction. It aims to separate sugars into 5 carbons (mostly solubilized) sugars into 6 carbons (mostly insoluble).

 The pentoses, resulting from the destructuration of the hemicellulose chains, are rendered soluble by the pretreatment, and migrate into the extraction solvent. As for cellulose, although hydrolysable, it remains mostly insoluble, and is concentrated in the solid fraction.

For a large quantity of straw to be treated (250kg / h), the liquid-solid separation is carried out on a belt press. The pretreated straw (stream 7, FIG. 3) is distributed uniformly over the lower fabric and progressively advances from a drainage zone to a pressing zone. In this area, pretreated straw is under increasing pressure and shear forces. Under the action of these two factors, the liquid fraction is extracted from the solid residue which sees its dry matter is raised. Example of a balance sheet resulting from the pretreatment: 1000 T of straw at 88% DM (stream 1) makes it possible to obtain 1 754 T of crude marc at 50% of MS (stream 7) containing 372 T of potential glucose and 177 T of xylose. C5 extraction is maximized to recover 95% of the C5 in the C5 juice stream (stream 9). The remaining 5% are found in the washed C6 marc (stream 10). In the example, the 1,754 T of crude marc (stream 7) are washed with 2,335 T of diluent (stream 29). This stream 29 can be composed of make-up water (stream 31) and / or clarified clarified stream 34. The separation leads to the production of 2643 T of C5 juice (stream 9) containing 95% of the C5 ( xylose) and having glucose and xylose concentrations of 1.2% and 6.4% (w / w), respectively. The other flow corresponds to 1 446 T of marc washed at 44% MS (stream 10) and contains 5% of the C5 as well as the majority of potential C6 (355 T versus 33 T in the C5 juice).

Preculture stage

The use of yeasts in fermentors requires the establishment of a chain of precultures. The first preculture consists in inoculating with the desired strain (here Candida shehatae) a tube containing the following medium:

 Yeast extract 10 g / kg

 - Bacto peptone: 10 g / kg

 - Xylose: 30g / kg

 - NaCl: 9g / kg

 The incubation conditions of this first preculture are as follows:

 Temperature: 30 ° C

 - pH: 5

 - agitation: 130 rpm

 After 12 hours of incubation under the above conditions, the first preculture serves as an inoculum for the second preculture performed in an Erlenmeyer flask.

The middle of the second preculture is shown in Table 3 below:

Le milieu contient également 30g/kg de xylose. Les conditions et la durée d'incubation sont identiques à celles suivies lors de la première préculture (30°C, pH 5, 130 RPM pendant 12h).

The medium also contains 30g / kg of xylose. The conditions and the duration of incubation are identical to those followed during the first preculture (30 ° C, pH 5, 130 RPM for 12 hours).

Stage of propagation of yeasts on C5 juice

 The precultures carried out previously make it possible to inoculate the yeast propagation fermenter. The propagation medium corresponds to stream 11. The latter is complemented in various elements listed in Table 3 above.

 The respective glucose and xylose concentrations of stream 11 are 7 to 12 g / kg and 45 to 64 g / kg. The mass fraction of the flow 1 1 dedicated to the propagation is here of 5%. The propagation culture conditions are as follows:

 Temperature 30 ° C

 pH 5.5 regulated with potash (KOH) and ortho-phoshoric acid agitation and aeration sufficient to maintain the P02 at more than 30% and this without dome pressure

 After 48 hours of culture under the above conditions, the culture makes it possible to obtain 22 g / kg of yeasts from 48.2 g / kg of xylose. The recovery yield of the substrate (xylose) in yeast (Yx / s) reached 46% / g). Co-products such as ethanol, glycerol, xylitol are also present at the following concentrations:

 - Ethanol: 0.38 g / kg

 - Glycerol: 0.4 g / kg

 - Xylitol: 0.15 g / kg

The conversion efficiency of the substrate in ethanol is therefore 1.8%. Step of fermentation of juice C5

 The juice of C5 (stream 9) is sent to about 80% towards the alcoholic fermentation of juice C5 (stream 12). The medium is complemented by following the same formula:

 Yeast extract at 5 g / kg,

 - Urea at 0.4g / kg

 The culture conditions for the fermentation of C5 juice are as follows:

 Temperature: 30 ° C

 - pH: 5.5

Aeration: oxygen limitation. The culture conditions described above (oxygen limitation), allow the yeast population to produce ethanol with a yield of 28% to 33%. The ethanol concentration reaches 18.3 g / l (glucose: 7.2 g / l, xylose: 52.3 g / l) after 46 to 63 hours of culture. At the same time, there was yeast production with a yield of 6 to 8%. Finally, co-products were generated with the following returns:

 Yp (glycerol) / s) = 0.021 g / g

 Yp (xylitol) / s) = 0.11 g / g

 Yp (ribitol) / s) = 0.039 g / g

L0 Monitoring the yeast population:

 The ethanol workshop of the C5 juice is inoculated with 1 g yeast per kg of medium. In addition to this population added during inoculation, yeast yield is between 6 and 8% and allows yeasts to be produced in parallel with ethanol production. At the end of the ethanol fermentation workshop of C5 juice, the

The yeast concentration thus reaches 0.6%.

 The wort corresponds to the stream 18 can then be sent directly in dilution water of the liquefaction stage (hydrolysis 1) or undergo a separation step S3 to remove yeasts from the wine (stream 18). This separation operation S3 makes it possible to obtain a wine free from the majority of yeasts (stream 19) and yeast creams (stream 20). In

In the example, half of these yeast creams are recycled in the ethanolic fermentation step of the C 5 juice (stream 21). The other half can be sent in fermentation of the C6 marc after the liquefaction stage (hydrolysis 1). This recycling of the yeasts makes it possible to substantially increase the concentration of yeasts in the stream 18. In fact, when the steady state is established, the concentration of yeasts at the outlet of the fermentation plant of the C5s

It reaches 1.2% against 0.6% without this recycling.

Monitoring of ethanol production:

Stream 12 (2075 T of C5 juice) contains 26 T glucose and 132 T xylose. The Yp (EtOH) / s yield of 28% to 33% thus makes it possible to produce 53.9 T of ethanol. In the example, the wine is stripped of these yeasts and sent directly to the liquefaction plant. This separation operation S 3 is carried out without washing the creams, which generates a flow 19 containing 49.6 T of ethanol (the difference in the amount of ethanol corresponds to the ethanol contained in the recycled creams: flow 21 and flow 22). The mass concentration of ethanol flux 33 (C5 wine cleared yeasts) is 2.5%, which corresponds to a TAV of 3.2 °. C6 fermentation step by hydrolysis and simultaneous fermentation or not

The washing of the marc made it possible to obtain a first stream rich in sugars with 5 carbon atoms (stream 9) and a second stream rich in sugars with 6 carbon atoms (stream 10). This latter stream contains an equivalent of 25% potential glucose. However, this glucose equivalent is in the vast majority in the form of cellulose. It is not usable as such by yeasts. It must be subjected to hydrolysis steps by action of enzymatic cocktails with cellulolytic actions, which allows both to reduce the viscosity of the product (liquefaction or hydrolysis step 1) and both to transform cellulose into cellobiose then glucose (hydrolysis 2). These hydrolysis steps can be carried out either totally separate from the fermentation (we speak of SHF), or simultaneously with the action of yeasts (we speak of SSF). In the latter case, the yeasts can be introduced either directly during the hydrolysis 1 (liquefaction), or during the hydrolysis 2. In the example, it has been chosen to carry out a hydrolysis step 1 (liquefaction) in the absence of yeasts and a hydrolysis step 2 in the presence of yeasts (SSF step).

Liquefaction stage

 The liquefaction stage consists in bringing together the washed marc containing the cellulose and the

enzymatic cocktail with cellulolytic actions. The dilution of the flow of C6 marc is allowed by the integration of the entire stream 19 (ex-C5 wine free of yeasts). This implementation makes it possible to obtain a cellulose concentration of 9.4%. The conditions of hydrolysis 1 (liquefaction) are as follows:

 Temperature: 50 ° C

 - pH: 4.8

 - enzyme concentration: 40mg / g cellulose

 duration from 2 to 12 h

 From a mass balance point of view, therefore, 1 446 T of washed marc (stream 10) diluted with 1 956 T of ex-C5 wine freed from yeasts (stream 19). The enzymatic cocktail flow is 14.2 T protein for 355 T of potential glucose.

Step of fermentation of marc C6

To facilitate the fermentation, the stream 23 is enriched in nutrients (yeast extract 5g / kg and urea 0.4g / kg). The potential glucose and xylose concentrations are 10-12% and 0.4-0.7%, respectively. After the liquefaction stage (hydrolysis 1), the liquefied stream (stream 23) is inoculated by the yeast cream corresponding to stream 17b (fresh yeasts from the propagation workshop) and by the yeast cream corresponding to the stream 22 (yeast cream obtained at the end of the C5 juice fermentation workshop: recycled yeasts). The crop conditions implemented are as follows:

 Temperature: 30 ° C

 - pH 5.5

 aeration: oxygen limitation

 The culture conditions described above allow the yeast population to produce L0 ethanol with a yield of 28% -33%. At the same time, there was yeast production with Yx / s yield of 8.2%.

Monitoring of the yeast population:

 According to the mass balance, the ethanol fermentation plant of the C6 marc corresponds to an L5 flow of 3 402 T of liquefied 10.4% potential glucose. These 3 402 T of juice are inoculated with 2.3 T MS of fresh yeasts obtained from the propagation of yeasts on C5 juice, but also from the yeasts resulting from the ethanol fermentation of C5 juice (stream 22) to height of 12.8 T MS. This amounts to inoculating the fermentation of the marc C6 with 4.3 g of yeast per kg of medium (including cream mass).

 10

 Monitoring of ethanol production:

 Stream 23 (3,402 T) contains 355 T of potential glucose and 9 T of xylose. The yield of hydrolysis of cellulose to glucose is 87%) and the conversion efficiency of glucose to ethanol of 33.4%. The residual xylose (9 T) is also fermented to ethanol with the same yield. The ethanol production during the SSF stage (or during the SHF) is therefore 106 T. Moreover, the stream 24 also contains the ethanol that was produced during the fermentation of the C5 juice (49.6 T d). ethanol). All this makes it possible to reach a mass concentration of ethanol in wine (stream 24) of 4.66%, a TAV of 5.9 °.

30 distillation stage

In the example, distillation only recovers a single flow that corresponds to stream 24. The title of this wine entering the distillation column is 5.9 °. The vapor consumption associated with a wine having a TAV of 5.9 ° is about 2.67 kg of steam per kg of ethanol produced. Step of treatment of the vinasses

 The vinasses at the outlet of the distillation column are clarified (stream 26) by an S4 separation plant. The lignocellulosic residues (stream 27) are treated so as to recover energy by carrying out their combustion (steam generation). The fraction of clarified vinasse (stream 28) is 100% recycled to the C5 extraction plant (stream 34). This recycling rate makes it possible to limit as much as possible the volumes of water introduced into the system (stream 31 = 239T). Overall performance step of the example

 The overall performance of this configuration is as follows: For 1000 tons of straw at 87.7% MS (or 877 kg of MS), the system produces 158.4 tons of ethanol, two thirds of which comes from the fermentation of C6 and the remaining third of the C5 fermentation. The overall yield of the recovery of dry matter ethanol is 18.06% (m / m). This configuration also allows to produce 4.7 tons of fresh yeasts divided into two equal streams towards the fermentation of C5 or fermentation of C6. The average TAV of the wines supplying the distillation workshop is 5.9 °.

Claims

Use of a propagation mash comprising a microbial population for preparing ethanol from a raw material generating at least one pentose, in particular xylose and / or arabinose, and at least one hexose, glucose, by fermentation of said pentose and hexose by said microbial population,  said raw material being extracted to give an extraction juice containing said pentose, in particular xylose and / or arabinose, and a pomace, said pomace giving, after being liquefied, a liquefied, said liquefied being a source of the above hexose, in particular glucose,  said propagation must being obtained by proliferation of an inoculum of said microbial population with the aid of said extraction juice, in the presence of air,  said propagation must containing no ethanol,  said microbial population being used at a time: for the preparation of a first source of ethanol, by fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice by said microbial population and  for the preparation of a second source of ethanol, by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied by said microbial population,  said ethanol being isolated from the first and / or second sources of ethanol, in particular by distillation of said first and / or second sources of ethanol,  said liquefaction of said grounds being carried out in the presence of all or part of said first source of ethanol, said first source of ethanol being, in particular, substantially free of micro-organisms from said microbial population. A process for preparing ethanol from a raw material generating at least one pentose, in particular xylose and / or arabinose, and at least one hexose, in particular glucose, comprising: a) a step of extracting said material, to obtain an extraction juice containing said pentose, in particular xylose and / or arabinose, and a pomace, b) a step of liquefying said coffee obtained in the preceding step, to obtain a liquefied, said liquefied being a source of the aforesaid hexose, in particular glucose, c) a step of proliferation of an inoculum of a microbial population using said extraction juice containing said pentose, in particular xylose and / or arabinose, in the presence of air, to obtain a propagation must comprising said microbial population, said propagation wort containing no ethanol, d) a first fermentation step of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice, by said microbial population, to obtain a first source of ethanol, e) a second fermentation step of the aforesaid hexose, in particular glucose, generated by said liquefied by said microbial population, to obtain a second source of ethanol, f) a step of isolating said ethanol from the first and / or second sources of ethanol, in particular by distillation of said first and / or second sources of ethanol, said liquefaction step of said grounds being carried out in the presence of all or part of said first source of ethanol obtained at the end of the first stage fermenting the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice, said first source of ethanol being, in particular, substantially free of microorganisms from said microbial population. A process for the preparation of ethanol according to claim 2, wherein said distillation, in addition to said ethanol, generates a residue, which contains substantially no ethanol, all or part of the soluble elements, water and elements. insoluble, is added to said raw material during said extraction of said raw material. 4. A process for the preparation of ethanol according to any one of claims 2 to 3, wherein said microbial population comprises yeasts of Candida shehatae strain, Pichia stipitis, Pichia guilliermondii, Candia tropicalis or Pachysolen tannophilus, said microbial population comprising in particular yeasts of Candida shehatae strain. A process for the preparation of ethanol according to any one of claims 2 to 4, comprising a microbial population for preparing ethanol from a raw material generating at least one pentose, in particular xylose and / or arabinose, and at least one hexose, in particular glucose, by fermentation of said pentose and hexose by said microbial population, said raw material being extracted to give an extraction juice (9) containing the aforesaid pentose, in particular xylose and / or arabinose, and a pomace (10), said pomace giving, after being liquefied, a liquefied (23), said liquefier being a source of the aforesaid hexose, in particular glucose,  said propagation must (13) being obtained by proliferation of an inoculum of said microbial population with the aid of said extraction juice (11), in the presence of air,  said propagation must containing no ethanol,  said propagation must (13) being subjected to a solid / liquid separation to give a propagation juice (16) and a fresh microbial population (17), said propagation wort containing no ethanol,  said fresh microbial population (17) being separated into two parts, to obtain a first portion (17a) and a second portion (17b) of said fresh microbial population (17),  said first part of said fresh microbial population (17a) being used for the preparation of a fermentation broth (18) by fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12), by said fresh microbial population, said fermentation wort (18) giving after said liquid / solid separation said first source of ethanol (19) and all or part of the recyclable microbial population (20),  said second source of ethanol (24) being obtained by fermentation of the aforesaid hexose, in particular glucose, generated by said liquefied (23), by all or part of said recyclable microbial population (20) and said second portion fresh microbial population (17b). The process for preparing ethanol according to any one of claims 2 to 5, comprising: a) a step of extracting a lignocellulosic biomass and all or part of the soluble elements, water and insoluble elements, of the residue (26) obtained at the end of the distillation step h), for obtaining an extraction juice (9) containing the aforesaid pentose, in particular xylose and / or arabinose, and a pomace (10), b) a liquefaction step of said pomace (10) obtained in the preceding step in the presence of all or part of said first source of ethanol (19) obtained at the end of the first fermentation step f), to obtain a liquefied (23), said liquefier being a source of the aforesaid hexose, in particular glucose, c) a step of proliferation of an inoculat comprising yeasts of Candida shehatae, Pichia stipitis, Pichia guilliermondii, Candia tropicalis or Pachysolen tannophilus strain, in particular Candida shehatae, with the aid of said extraction juice (11) containing the aforesaid pentose, in particular xylose and / or arabinose, in the presence of air, to obtain a propagation must (13) comprising a population of said yeasts, said propagation wort containing no ethanol, d) a solid / liquid separation step relative to said propagation wort (13), to give a propagation juice (16) and a population of said fresh yeasts (17), e) a step of separating said fresh yeast population into two parts, to obtain a first portion (17a) and a second portion (17b) of said population of fresh yeasts, f) a first step of fermentation of the aforesaid pentose, in particular xylose and / or arabinose, contained in said extraction juice (12), by said first portion of the population of said fresh yeasts (17a), to obtain after liquid / solid separation a first source of ethanol (19) and a population of said yeasts, recyclable (20), g) a second fermentation step of the aforesaid hexose, in particular glucose, generated by said liquefied (23), by said population of said recyclable yeasts (20) and said second portion of said population of said fresh yeasts (17b), to obtain a second source of ethanol (24), h) a step of isolating said ethanol from the first (19) and / or second (24) sources of ethanol by distillation of said first and / or second sources of ethanol, said distillation generating, in addition to said ethanol, a residue (26); ), which contains substantially no ethanol. 7. Process for the preparation of ethanol according to any one of claims 2 to 6, wherein said raw material is, prior to said extraction, impregnated and pretreated, said raw material being particularly subject, after said impregnation and said pretreatment, additional hydrolysis treatment. 8. Process for the preparation of ethanol according to any one of claims 2 to 7, wherein the raw material is a lignocellulosic biomass, in particular chosen from: - cereal straw, said cereals being in particular chosen from wheat, barley, triticale, sorghum and maize,  - cereal bran, said cereals being in particular chosen from wheat, barley, triticale, sorghum and maize,  energy crops, chosen in particular from switchgrass, miscanthus and arundo donax, and  - forest crops, chosen in particular from wood, short rotation coppices, very short rotation coppices, and forest chips. A process for preparing ethanol according to any one of claims 2 to 8, wherein said extraction juice containing said pentose, in particular xylose and / or arabinose, is detoxified prior to said proliferation and the preparing said first source of ethanol. The process for preparing ethanol according to any one of claims 2 to 9, wherein said liquefied is:  hydrolyzed before said fermentation of the aforesaid hexose, in particular glucose, generated by the hydrolysis of said liquefied material, or  hydrolysed, said hydrolysis and said fermentation of the aforesaid hexose, in particular glucose, generated by the hydrolysis of said liquefying being concomitant.