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Description:

Pretreatment and Fermentation of Fibrous Feedstocks for Feeding and Energy: a "green" process.

By D. A. Flores

SB Internet

Port Coquitlam, BC

Canada V3B 1G3

The Problem and the Potential.

As research proceeds to utilize residual fibre-based feedstocks for both animal feeding and for energy feedstock, described here are the "green" pretreatment processes discussed as to the nature of technological adaptation within the milieu of bioreactor and animal rumen stomach fermentation processes together with their biomolecular-level process and manipulations adapted for fermentation of microbes.

We will discuss the e. g. of pretreatment by solid state fermentation (SSF) with fungal inoculants to selectively breakdown the ligneous components of lignocellulose and at the same time supplement with single cell protein (SCP) the nitrogen (N) for improved feedstock digestibility with animal feeding or for batch or continuous fermentors for energy alcohols and what is introduced here as Immersive Cold Energy (ICE) Fermentation under much colder ambient conditions adapted for more northern climes with the accessibility to lignocellulosic biomass including agroforestry pulp and farm waste residual biomass, as examples.

Describing Microbial Bioreactor and Rumen Fermentation.

New enzyme technology will be used to process the feedstock to sugars and oligosaccharides, a so-called "super energy" for fermentation, and also enzymic repolymerization to new substrates, also a "super energy" for fermentation.

It is a known theory that oligopeptides are a major driving force in microbial cell protein synthesis, for e. g., in the ruminant's rumen ecosystem, rather than as would be thought conventionally as individual free amino acids (FAA), and further, as conjectured, a role for oligosaccharides including water-soluble carbohydrates (WSC), e. g. fructans as oligomeric sugars, as part of the complex mixtures of sugars and carbohydrates (CHO), also present in Spring grass fodders serving "catalytically" to boost microbial cell growth hypothesized in part to be 'mitogenic' in cellular metabolism.

It should be averred to that the current problem of drug discovery starting with our proprietary Lanosteral precursor for bioreactor synthesis using the microbial enzyme regulated process to the VitD2 agonist (active form) drug lead target (alternatively is the VitD3 agonist, active form) is a schema in the same realm of problem solving in a more complex sense for modifying/selecting clones with the ideal enzyme forms and producing batches of precursor substrate before pursuing the next step down the line with "banking" as another strategy to supply copious enough amounts of substrate as reserve for this elaborated ~30-step typical drug synthetic sequence, without reiterating back each time as needed. The enzymology of the bioreactor in the method provides for host generated substrates or intermediates that are exact to the desired specifications as to their oligomeric or polymeric structure. This method can theoretically be used to derive tailor-made enzymes for producing potentially very valuable substrates for oligomers of amino acids and sugars in the future as functional feed additives including for cell fermentation endproducts and microbial cell protein (MCP) applicable to both batch or continuous fermentors for both feeding supplements and for rumen digestion. This could yet be an important field in proteomics research and enzyme engineering.

There is also new technology proposed here using PNA-B12 biologics for activating or dampening regulatory pathways in microbial cell metabolism, the hosts, labelled as genetically regulatory organisms (GRO), for e. g., those being manipulated for transport across the microbial cell membranes for both major substrates in sugars and proteins, as oligomers, and manipulated synthetic rates of protein synthesis from these imported oligopeptides and from ATP production with fermentative catabolism.

The Processing of Solid-State Fermentation (SSF) Yeast Residual Feeds.

The beginnings of this technology type were with the bagasse process by a Japanese concern in the Philippine Islands to feed dairy cows (viz. Holstein bred-type cattle) for milk production. It therefore comes to mind that year processing, however modified to be enhanced, refined and made cost-effective and animal tested, can be extended to cover also other byproduct farm wastes such as sugarcane pith from bagasse, corn stover and sorghum stover, haulms from vines or legume spp., rice and wheat straw, the latter occupying the largest proportion of byproduct farm waste and potentially the most productive in terms of volume processed and consumed by animal livestock. The outline of major stages wherein the milled substrate is processed into "yeast byproduct" feed is now given.

Making a Semi-Refined Consistent Particulate Based.

First is threshing the byproduct biomass feedstock from the field's harvest on-farm and then fine-chopping as in silage making, washing thoroughly the substrate followed by air-drying over 48 hrs, with the option to forced air-drying the biomass. This is followed after drying with fine grinding to a semi-consistent powder. Adaptation is made per substrate type or feedstock.

Mixing Reaction Components Prior to Live Yeast Culture (LYC) Incubation.

The dried particulate substrate or biomass is first bleached with added dilute aqueous acid [H2SO4, (aq)] over a 48 hr period and then mass centrifuged (industrial type) and washed clear of any residuals and then the pH adjusted for enzymatic conditions involving aerobic lacasse treatment via bioreactor over a few days depending on the batch weight used.

Addition of LYC and Incubation.

The yeast inoculum or live yeast culture (LYC) is applied as follows. The remixed or reconstituted LYC is counter-sprayed onto the spray-lofted particulate feedstock and further auger mixed into a mixer together with a biologic-added solution to the LYC just before it is stored for incubation with the feedstock. This will be used to hyper boost or produce high enough levels of HIS, ARG, LEU, MET and LYS eventually in the incubating biomass. These are so-called GROs in yeast. As for the temperature, pH and time of incubation under open air conditions (S. cerevisiae, the spp. of choice, is facultatively anaerobic), these are yet to be determined experimentally for the yeast fermentation processes.

Air-Spray Drying and Storage.

The biomass substrate for feeding is kept in the holding tank for incubation and is effectively stored in a compacted, cool, and dry place after thorough drying by spraying onto an industrial chamber wall and collecting at the base. It is then collected with the biologic already 'spent' and not in any measurable concentration, i. e., it is strictly labile after a defined period of storage and mechanically packaged in square, rectangular bags for feed dispensing at the farm feedlot for dairy and beef cattle. The matter of the on-surface film LYC is apparently flash-dried with forced air with, for e. g. , N2 (g) for drying, which instantly kills and renders it a DYC (as non-sporulated it is hypothesized) on the particulate surface and fully dried away after N2(g) sealed packing to be kept inert until dispensed with feeding.

Feeding Trials Regards Nutritive Value (N. V.) of Protein and Energy Supplements.

It is open to question at this time what levels of amino acids can be attained and what are optimal for functional use such as lean body mass (LBM) accretion, milk solids output, and in general, the body condition of finishing steers and lactating cattle. There is a need to test various basal rations such as hay or silage from herbage plus the addition of highly digestible fibre from herbage (grasses and legumes) and boost protein-amino acid (AA)/energy concentrates derived from farm feedstock. What remains as the most limiting amino acids for optimum nutrition is to be demonstrated by nutritionists and what should be supplemented. Here used as an example is the biological rate of addition of a theoretical PNA-B12 biologic to a live yeast culture (LYC) innoculant and the volume of pretreated LYC biomass prior to and during the mixing process and the incubation conditions, to effectively elevate microbial cell protein (MCP) levels and so with incremental increase in critical essential amino acids that are functional over and above their nutritional value as measured by feeds analysis and predicted to livestock product. It is expected that "spiking" the amino acids of the supplemental feed(s) will incur different results in nutritive value (N. V.) when manipulated in addition to its conventional role in fulfilling nutritional amino acid daily allowances for requirements.

Proposals here are preliminary leading one to speculate what specifically is needed to be learned from further research regards amino acid nutrition in various ruminant class livestock, their dairy allowances to optimize hay or silage feeding, and of course more data obtained to study specifics of rumen digestion and the supply of protein and its amino acids from it to compare and determine which are most limiting. Amino acids of interest here are: histidine (HIS), arginine (ARG), lysine (LYS) and leucine (LEU), in particular.

Semi-Automated Mechanical Field with Solid-State Fermentation (SSF) Processing.

Proposed here is the amalgamation of enzyme technology including fibrolytics anaerobic enzymes (non-O2 sensitive) called lignases that can work with "dry" solid-state fermentation (SSF) that is fungal-driven, in the field as it speculated via process "flocculation" onto the biomass under field conditions, slow-released by dissolution onto the field ensiling biomass and anoxygenically excluded of air with semi-automated field operations in mechanical-driven process flow.

Mechanically, the field is to be longitudinally "striated" by semi-automated cutters and combines which can air spray cultures onto the precision-cut biomass, while segmented bagging occurs with sealing and the air flow excluded with CO2/N2 and field incubated until completion by microbial and enzymatic action when biomass nutrifies with MCP or protein from this fungal-driven process.

Farming conditions will require more sophisticated metereological forecasting with use of advanced technologies such as WiFi, GPS and weather satellites overhead to manage during harvesting times annually the temperature and air humidity and to avoid or minimize losses due to weathering in the field.

Harvesting will then take place by opening the locked bag longitudinally by semi-automated collected baling in closed coil cylindrical rolls and bag sealed again for storage in the field before transport to downstream plants as feedstock for both animal feed and biofuel fermentation.

Feeding trials will have to be carried out for both balance in nutritional requirements and oropharyngeal qualities for any changes in feed intake under controlled animal field trial conditions.

Immersive Cold Energy (ICE) Partitioned Fermentation for Fibre-based Feedstocks: Process Conception and Biomolecular Mechanisms.

The conception for fibre-based processed feedstock conversion to high concentrate energy alcohols as in iso-pentenol and iso-butanol, the latter from ABE or acetone-butanol-ethanol fermentation, is discussed here at length utilizing psychrophilic microbial cultures from the sub-Arctic (e. g. sea or tundra) which would put an upper cap on maintaining energy needs during the frigid Winter season, and even during the "lighter" seasons, using cold saline or freshwater from lakes or rivers targeted for locations in northern geographical areas in Canada and even for Siberia in Soviet Russia.

It is believed as well that with the advent of heat exchanger technology which is to be deployed in ICE fermentation plants in the near-Arctic regions, that during the warmer months of the year, cultures can be cooled down through maintaining a seamless performance in energy extraction/cooling them naturally at great energy savings.

Deploying plants in the true North of Canada will also save on transportation of fibre biomass inputs from further south including any available waste pulp from the silviculture industry requiring transportation.

The Model's Process Conception and Molecular Mechanisms.

The fundamental idea here is to partition the cell's surroundings and interior milieu, between the output in the outer milieu of the bioreactor where product can be partitioned out and cumulated to be downstream processed in the continuous reactor system using microbial immobilization onto a column, while the inside milieu of the cell is revved up metabolically to produce upregulated rates of product through the new technology of PNA-B12 (there is also a hypothetical carriered PNA-K1, K2 yet to be tested that is being proposed) readily entering the cell through its molecular mechanisms with the ATP generating mitochondria and the enzymatic catabolic network to form iso-pentenol or iso-butanol in the bioreactor's outer milieu together with cell membrane, outer membrane and mitochondrial membrane, with modification using mutational changes in the transport channeling porins of both mitochondria for the export of energy-yielding currency of ATP and so-called dual-transporter carrier systems, at equilibrium, using facilitated diffusion (preferable to active transport that would utilize too much ATP energy by the cell) allowing the inner cell milieu to "vent" of what would otherwise be the "overproduction" of alcohols and leading to toxification of cell hosts.

Changes in enzymatic terms of transport proteins (or porins) that somehow mutate fortuitously to accelerate rates of processing of substrate can be studied with proteomics and then "superposed" to genomic data for studying gene mutations and then the genes edited onto the original genome with gene editing (GEditing) with exact precision.

There is also analysis using X-ray diffraction techniques to visualize the atomic centres and their bond lengths, and the orientations with respect to each other, shedding light on the fine-structural mechanisms of the reaction of transport processes. When it comes to enzymic acceleration usually the matter centres around feedback inhibition (FBI) which can be antagonized in mechanism as well as altering the Michaelis-Menten reaction describing receptor-substrate's binding on the outer and inner membrane's interphase in the cell.

Immersive Cold Energy (ICE) fermentation marries together the ideas of membrane biology and PNA-carrier based genetically regulated organismal (GRO) type manipulations for metabolism, thus partitioning the process between making "something on the spot" and on the other hand taking it out of the "manufacturing floor" rather than having if "back up", thus killing the cell host, when in fact a faster rate of equilibrium can be achieved and maintained over the continuous, immobilized fermentor's reactive process. And, at that, at a lower temperature of maintenance at the reactor's floor and also within its insulated interior including its filtered solvent milieu.

It is presumed that distances between plant location where it is conveniently cold (in the Winter) or cool (the rest of the year), water sources, either saline or fresh, transport into the location of inputs including processed fibre feedstock and transport of downstream processed materials to their depots for distribution be strategically located for cost and convenience.

There is now a belief that colder countries can utilize fibre-base feedstocks from farmlands for bioenergy fuel generation, and together with conservation of oil reserves and with other energy unconventional energy sources like wind, solar and hydro, survive independently from their more southernly neighbors in the Americas who already plant considerably fibre-based plant feedstocks from cropping farm wastes such as straws, stover and bagasse utilizing them eventually in future in like with oil reserves as planned up North (U.S.A and Canada). Without complicating each other's trade ties to utilize farm waste milled feedstocks from pulp and sugarcane bagasse, as examples, that they may opt for like their northernly neighbours.

//END

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