# US 2014/0154231 A1 · Process for separating proteins fibrinogen, factor xiii and biological glue from a solubilized plasma fraction and for preparing lyophilised concentrates of said proteins > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2014/0154231 A1 - Document ID: US20140154231A1 - Status: Published application - Kind: A1 - Application number: 14/056,903 - Filed: 2013-10-17 - Published: 2014-06-05 - Earliest claimed: 2005-06-29 - Assignees: Laboratoire Francais Du Fractionnement Et Des Biotechnologies - Inventors: Michel NOGRE; Pierre PORTE; Michel TELLIER - Web page: https://patentgrep.com/patent/US20140154231A1 ## CPC - A61K 38/45: Transferases (2) - A61P 7/04: Antihaemorrhagics; Procoagulants; Haemostatic agents; Antifibrinolytic agents - A61P 17/02: for treating wounds, ulcers, burns, scars, keloids, or the like - C07K 14/75: Fibrinogen - C12N 9/1044: {Protein-glutamine gamma-glutamyltransferase (2.3.2.13), i.e. transglutaminase or factor XIII} - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) - A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) ## Abstract Process for separating proteins fibrinogen, Factor XIII and biological glue from a solubilized plasma fraction and for preparing lyophilised concentrates of said proteins. ## Claims ### Claim 1 A freeze-dried biological glue concentrate, obtained by a process comprising the steps of: i) equilibrating an anion. exchanger of weak base type with a first buffer having a predetermined ionic strength of an alkaline pH, ii) loading said anion exchanger of weak base type with a Cohn fraction I, thus allowing the retention of the biological glue, iii) eluting the biological glue with a second buffer having an alkaline pH and an ionic strength greater than said first buffer, iv) concentrating the biological glue by ultrafiltration, and v) diafiltering the biological glue solution, followed by freeze-drying of said solution. ### Claim 2 A freeze-dried Factor XIII (FXIII) concentrate, obtained by a process comprising the steps of: i) equilibrating an anion exchanger of weak base type with a first buffer having a predetermined ionic strength of an alkaline ph, ii) loading said anion exchanger of weak base type with a Cohn fraction I, thus allowing the retention of the biological glue, iii) eluting the biological glue with a second buffer having an alkaline pH and an ionic strength greater than said first buffer, iv) concentrating the biological glue by ultrafiltration, and v) separating FXIII from fibrinogen by adding to at least a part of the biological glue eluate at least one chemical agent precipitating the FXIII, wherein the chemical agent precipitating the FXIII is present in the form of an aqueous solution based on sodium citrate and resolubilizing the precipitated FXIII, and vi) diafiltering the resolubilized FXIII solution, followed by freeze-drying said solution. ### Claim 3 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein said first buffer has an ionic strength of less than 0.2. ### Claim 4 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein the pH of said first buffer is in the range of values higher than 7 up to 9. ### Claim 5 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein the process includes, prior to the biological glue elution step, a washing step with said first buffer of the anion exchanger until not retained proteins and contaminants are removed. ### Claim 6 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein the biological glue elution is carried out with said second buffer having an ionic strength in a range between 0.5 and 1.3, the pH of which is set to a value of 7.4-7.6. ### Claim 7 (depends on 5) The freeze-dried concentrate according to claim 5, wherein said second buffer further contains a mixture from 10 to 12 g/l of trisodium citrate, from 1 to 5 g/l of lysine, from 1 to 5 g/l of glycine, from 2 to 5 g/l of Tris, from 25 to 50 g/l of arginine and from 5 to 15 g/l of isoleucine. ### Claim 8 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein the chemical agent precipitating the FXIII is present in the form of an aqueous solution based on citrate salts 1 M. ### Claim 9 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein the precipitate of FXIII is resolubilized in water or in a buffer containing a mixture from 10 to 12 g/l of trisodium citrate, from 1 to 5 g/l of lysine, from 1 to 5 g/l of glycine, from 2 to 5 g/l of Tris, from 25 to 50 g/l of arginine and from 5 to 15 g/l. isoleucine, at a pH between 6.9 and 7.1. ### Claim 10 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein the diafiltration is carried out against the buffer containing a mixture of trisodium citrate, lysine, glycine, arginine and isoleucine. ### Claim 11 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, the process by which the concentrate is obtained further comprising at least one step of viral inactivation and/or viral and contaminants removal treatment, selected from the group consisting of a chemical viral inactivation treatment, a nanofiltration and a dry heat viral inactivation treatment. ### Claim 12 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein the nanofiltration is carried out with the eluate obtained in step a) or with the diafiltered fibrinogen, biological glue and resolubilized XIII solutions, prior to freeze-drying. ### Claim 13 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, wherein a concentration step by ultrafiltration is carried out prior to the step of diafiltration or following the said step, prior to freeze-drying. ### Claim 14 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2, including, prior to step a), an initial prepurification step of the solubilized plasma fraction by a classical pretreatment with aluminium hydroxide and/or by a precipitation at low temperature. ### Claim 15 (depends on 1, 2) The freeze-dried concentrate according to claim 1 or 2 of therapeutic quality, obtained by carrying out of a process including at least one step of viral inactivation and/or viral and contaminants removal treatment, selected from the group consisting of a chemical viral inactivation. treatment, a nanofiltration and a dry heat viral inactivation treatment.