Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Enzymatic production of a protein or polypeptide
Reexamination Certificate
2001-02-01
2002-12-17
Weber, Jon P. (Department: 1651)
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Enzymatic production of a protein or polypeptide
C530S343000, C530S350000, C530S372000, C530S376000, C530S374000
Reexamination Certificate
active
06495342
ABSTRACT:
TECHNICAL FIELD
The present invention provides a nitrogenous composition resulting from the enzymatic hydrolysis of maize gluten.
The present invention also relates to a process for the preparation of such a nitrogenous composition and to the application thereof in culture media for the fermentation industries.
This nitrogenous composition may also be used as a feedingstuff or food additive in compositions for animals.
BACKGROUND OF THE INVENTION
Maize gluten is known to be a source of nitrogen in the fermentation industries and a constituent of certain foodstuffs.
However, due to the viscoelastic properties of maize gluten, it must be solubilised if it is to be used as a source of nitrogen in bioengineering processes or as a feedingstuff or food additive.
This solubilisation of gluten may be carried out by chemical means (alkaline hydrolysis or acid hydrolysis) or by enzymatic means.
For example, patent application WO 95.28850 describes a method of solubilising plant proteins by using enzymes with phytase activity in combination with one or more enzymes with protease activity, one or more enzymes with lipase activity and one or more enzymes with glycosidase activity. These enzymes are extracted from microorganisms of the Bacillus, Pseudomonas, Kluveromyces, Saccharomyces, Aspergillus type.
The mixture thus hydrolysed is then used in animal feed.
The patent EP 518.999 describes the preparation of partially hydrolysed products by virtue of a specific treatment using a particular serine protease produced by
Bacillus licheniformis.
These products are of interest for the preparation of foods for human consumption and particularly in infant foods as an agent which limits the allergenic capacity of the proteins of full cream milk, and on account of their properties of better digestibility.
Patent application EP 461.261 describes the preparation of particular oligopeptides by enzymatic hydrolysis of plant proteins in order to obtain a high proportion of branched amino acids and the use thereof in food preparations intended for patients suffering from liver diseases.
A process which allows the preparation of an aromatiser by enzymatic hydrolysis of a substance rich in plant proteins without having to use purified exopeptidases to take away the bitterness, whilst giving it attractive organoleptic properties, has been described in patent EP 429.760.
The possibility of using an hydrolysed of plant proteins in a fermentation medium is also mentioned in patent application WO 94.25580.
However, none of these documents indicates or suggests that a hydrolysed of maize gluten having a particular composition in terms of its phosphorus and nitrogen content, its concentration of soluble and insoluble proteins and its distribution spectrum of amino acids, may be particularly suitable for fermentation applications, particularly in the field of fermentations of microorganisms which produce organic acids and/or in animal feed applications.
DESCRIPTION OF THE INVENTION
The invention relates primarily, therefore, to a nitrogenous composition resulting from the hydrolysis of maize gluten, having a ratio of the concentrations of inorganic phosphorus to total phosphorus (Pi/Pt)greater than or equal to 0.05, preferably from 0.05 to 0.5, and a ratio of the concentrations of amine nitrogen to total nitrogen (Na/Nt) greater than or equal to 0.025.
The concentrations of inorganic phosphorus and total phosphorus are measured by methods known on the whole by the skilled person, such as those described below.
With regard to inorganic phosphorus, the reference method consists in extracting the mineral or inorganic phosphorus by a solution of trichloroacetic acid, forming a phosphomolybdic complex by reaction between inorganic phosphorus and ammonium molybdate, and measuring the absorbance of this complex with a spectrophotometer at a wave length of 360 nm.
The determination of total phosphorus is carried out by a method which is based on the detection of phosphorus on a plasma emission spectrometer (JY83 type from JOBIN YVON following the constructor's specifications). A preliminary step is used, however, which consists in destroying the organic materials of the products to be determined in order to convert all the organic phosphorus to inorganic phosphorus (the total phosphorus is therefore the organic phosphorus plus the inorganic phosphorus), by mineralisation using a sulfonitric mixture.
The ratio of the inorganic phosphorus concentration to the total phosphorus concentration (Pi/Pt) is preferably greater than or equal to 0.05, preferably from 0.05 to 0.5.
The concentrations of amine nitrogen and total nitrogen are measured by known methods such as those described below.
The determination of amine nitrogen is carried out by allowing the amine function of the product to be determined to react with trinitrobenzenesulfonic acid (TNBS) in order to produce a chromophore group, the absorbance of which is measured at 340 nm.
The determination of total nitrogen is carried out according to the ISO 3188 method which involves mineralisation of the product to be determined with sulfuric acid in the presence of a catalyst, then alkalinisation of the reaction products and distillation of the ammonia liberated and collected in a solution of boric acid which is titrated with a solution of sulfuric acid.
The total protein content and the soluble protein content of the composition according to the invention are, respectively, from 20% to 80%, based on dry matter basis, and from 30% to 80%, on dry matter basis.
The term total protein content means the total nitrogen multiplied by a factor of 6.25.
The term soluble protein content means the total nitrogen multiplied by a factor of 6.25 contained in the soluble fraction which corresponds to the supernatant obtained after dispersion of the sample in distilled water and centrifugation.
Moreover, the nitrogenous composition according to the invention has a free alanine content greater than or equal to 100 mg/100 g on a dry matter basis, preferably 500 mg/100 g on a dry matter basis, a free leucine content greater than or equal to 200 mg/100 g on a dry matter basis, preferably 1000 mg/100 g on a dry matter basis, and a free phenylalanine content greater than or equal to 100 mg/100 g on a dry matter basis, preferably 500 mg/100 g on a dry matter basis.
The invention relates secondly to a process for the preparation of a nitrogenous composition resulting from the enzymatic hydrolysis of maize gluten by a particular combination of enzymes.
The maize gluten used is preferably a slurry of maize proteins obtained after grinding grains of steeped maize and after separating the germs and starch by a physical process based on the difference in density of the compounds.
Maize gluten may also be extracted from the slurry of proteins by physical separation of the centrifugation or filtration type and then resuspended in water in order to carry out the enzyme treatment according to the invention.
The process according to the invention consists in subjecting an aqueous solution of maize gluten to an enzyme treatment using at least one protease and optionally at least one enzyme selected from the group comprising a complex of endo- and exopeptidases, an enzyme complex which hydrolyses the polysaccharides of maize other than starch, a phytase and a glucoamylase.
In a first method of preparing the nitrogenous composition according to the invention, an aqueous solution of maize gluten is treated in a series of steps comprising:
introducing successively into the aqueous solution of maize gluten at least one protease then optionally a complex of endo- and exopeptidases,
allowing the enzymes to act, with stirring,
inactivating the enzymes,
optionally separating the insoluble matter,
recovering the nitrogenous composition thus obtained.
The proteases are preferably papain (for example, COROLASE® L10 sold by the company RÖHM ENZYME) and/or an alkaline protease (such as the alcalase isolated from
B. licheniformis
). This first hydrolysis reaction will be performed advantageously at a pH of 8.
The co
Dubois Eric
Fouache Catherine
Seigueilha Laurent
Henderson & Sturm LLP
Roquette Freres
Srivastava Kailash C.
Weber Jon P.
LandOfFree
Nitrogenous composition resulting from the hydrolysis of... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Nitrogenous composition resulting from the hydrolysis of..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Nitrogenous composition resulting from the hydrolysis of... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2985622