ITMI960227A1

ITMI960227A1 - SEMI-FLUORINATED ALKANES AS OXYGEN CARRIERS IN BIOLOGICAL SYSTEMS

Info

Publication number: ITMI960227A1
Application number: IT96MI000227A
Authority: IT
Inventors: Hasso Meinert; Wolfgang Rohlke; Peter Reuter; Ursula Geister; Joachim Bar; Costanze Lenz; Haike-Henrike Medenblik; Aime Cambon; Heidi Trabelsi; Kenneth Charles Lowe; Carl Martin Edwards; Giampaolo Gambaretto; Massimo Napoli; Lino Conte
Original assignee: Miteni Spa
Priority date: 1996-02-08
Filing date: 1996-02-08
Publication date: 1997-08-08
Also published as: ITMI960227A0; IT1282579B1

Abstract

Alcani semifluorurati a blocchi (fluoroalcani) aventi le formule seguenti: (FORMULA I e II) dove R, R1 ed R2 sono uguali o diversi tra loro e scelti tra idrogeno ed alchile C1-C8, n è compreso tra 1 e 20 ed m, q e p sono uguali o diversi tra loro e sono compresi tra 1 e 8, come trasportatori/mediatori di ossigeno in sistemi biologici.Detti fluoroalcani consentono di realizzare emulsioni stabili atte ad essere impiegate in sostituzione di fluidi biologici, ad esempio come sostituti del sangue o come fluidi "resuscitativi" per il trasporto e/o la conservazione di organi, e nelle "banche" di organi.Block semifluorinated alkanes (fluoroalkanes) having the following formulas: (FORMULA I and II) where R, R1 and R2 are the same or different from each other and selected from hydrogen and alkyl C1-C8, n is between 1 and 20 and m, q and p are the same or different from each other and are between 1 and 8, as oxygen transporters / mediators in biological systems. These fluoroalkanes allow to create stable emulsions suitable to be used in replacement of biological fluids, for example as blood substitutes or as "resuscitative" fluids for the transport and / or conservation of organs, and in the "banks" of organs.

Description

OGGETTO DELL'INVENZIONE OBJECT OF THE INVENTION

La presente invenzione ha per oggetto l'impiego di alcani semifluorurati (fluoroalcani) come mediatori trasportatori di ossigeno in sistemi biologici. The present invention relates to the use of semifluorinated alkanes (fluoroalkanes) as oxygen-carrying mediators in biological systems.

Pure oggetto della presente invenzione e' un procedimento per la purificazione di detti fluoroalcani. Also object of the present invention is a process for the purification of said fluoroalkanes.

Sempre oggetto del presente trovato sono emulsioni a base di fluoroalcani e impiego di dette emulsioni come sostituti/integratori di fluidi biologici, quali ad esempio sangue o suoi derivati. The present invention also relates to emulsions based on fluoroalkanes and the use of said emulsions as substitutes / integrators of biological fluids, such as for example blood or its derivatives.

STATO DELLA TECNICA STATE OF THE TECHNIQUE

Come e' noto, le emulsioni a base di perfluorocarburi sono impiegate come fluidi resuscitativi o come sostituti del sangue. I perfluorocarburi agiscono come mezzi trasportatori di ossigeno e svolgono un'azione sostitutiva delle funzioni dell'emoglobina nel sangue naturale umano ed animale. As is known, perfluorocarbon-based emulsions are used as resuscitative fluids or as blood substitutes. Perfluorocarbons act as oxygen transporters and perform a substitute action for the functions of hemoglobin in natural human and animal blood.

La dimensione delle particelle dei perfluorocarburi e' ritenuta un fattore importante in relazione ai problemi di tossicità' e agli effetti collaterali che si riscontrano impiegando fluidi biologici modificati; in particolare, quando le particelle hanno una dimensione di 500 nm o quando hanno dimensioni medie superiori a 200 nm, risulta problematico per una efficace alimentazione degli animali-tests. The particle size of perfluorocarbons is considered an important factor in relation to the toxicity problems and side effects encountered when using modified biological fluids; in particular, when the particles have a size of 500 nm or when they have an average size greater than 200 nm, it is problematic for an effective feeding of the animals-tests.

Esiste quindi la necessita' di impiegare perfluorocarburi caratterizzati da particelle di dimensioni estremamente ridotte. Come e' noto, i perfluorocarburi per essere utilizzati come trasportatori di ossigeno e come sostituti del sangue, devono essere portati in forma di emulsioni stabili. Quando vengono utilizzati perfluorocarburi caratterizzati da particelle di piccole dimensioni, si riscontrano alcune difficolta' nel realizzare emulsioni adatte che siano stabili nel tempo e quindi conservabili con facilita'. Infatti, i perfluorocarburi a particelle di piccole dimensioni, se da un lato non provocano problemi di tossicità' e non danno luogo ad effetti collaterali significativi, dall'altro non consentono di realizzare emulsioni stabili nelle condizioni ordinarie di conservazione, a causa della loro immiscibilità' nei mezzi acquosi nei quali sono emulsionati, con conseguenti problemi di biocompatibilita'. Come trasportatori di ossigeno nei mammiferi sono stati utilizzati a livello sperimentale numerosi perfluorocarburi, quali ad esempio: perfluorotripropilammina, perfluorooctilbromuro, perfluoroadamantano, perfluorometiladamantano, perfluoro -1,3- dimetiladamantano, perfluorotrimetilbiciclononano, perfluoro -4- metiloctaidrochinolizina, perfluoro -n- metildecaidroisochinolina, perfluorocicloesilmorfolina, perfluoro -n- esilmorfolina, perfluorodimorfolino -n- alcani e perfluorodipiperidino -n- alcani. There is therefore a need to use perfluorocarbons characterized by extremely small particles. As is known, perfluorocarbons must be brought in the form of stable emulsions to be used as oxygen carriers and as blood substitutes. When perfluorocarbons characterized by small particles are used, some difficulties are encountered in making suitable emulsions that are stable over time and therefore can be easily stored. In fact, the perfluorocarbons with small particles, if on the one hand they do not cause toxicity problems 'and do not give rise to significant side effects, on the other hand they do not allow the creation of stable emulsions in ordinary storage conditions, due to their immiscibility' in the aqueous media in which they are emulsified, with consequent problems of biocompatibility. Numerous perfluorocarbons have been used experimentally as oxygen transporters in mammals, such as, for example: perfluorotripropylamine, perfluorooctylbromide, perfluoroadamantane, perfluoromethyladamantane, perfluoro -1,3- dimethyladamanthane, perfluorotrimethylbicyclonine- methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl methyl chlorine , perfluoro -n- hexylmorpholine, perfluorodimorpholino -n- alkanes and perfluorodipiperidino -n- alkanes.

Sono stati anche provati numerosi emulsionanti per emulsionare i perfluorocarburi in fase acquosa; alcuni di questi sono ad esempio i tensioattivi "Pluronic", che hanno la truttura chimica di un copolimero poliossietilene-poliossipropilene, i lipidi, in particolare la lecitina dal tuorlo d'uovo, i fosfolipidi dal rosso d'uovo e fosfolipidi derivati dalla soia; pure sono stati provati i monogliceridi di acidi grassi scelti dal gruppo comprendente: acido caprilico, acido decanoico, acido laurilico, acido dodecanoico, acido tetradecanoico, acido esadecanoico, acido stearico, acido docosanoico, acido palmitoleico, acido oleico, acido linoleico e acido arachidonico. Numerous emulsifiers have also been tried to emulsify perfluorocarbons in the aqueous phase; some of these are for example the "Pluronic" surfactants, which have the chemical structure of a polyoxyethylene-polyoxypropylene copolymer, lipids, in particular lecithin from egg yolk, phospholipids from egg yolk and phospholipids derived from soy; the monoglycerides of fatty acids selected from the group comprising: caprylic acid, decanoic acid, lauryl acid, dodecanoic acid, tetradecanoic acid, hexadecanoic acid, stearic acid, docosanoic acid, palmitoleic acid, oleic acid, linoleic acid and arachidonic acid were also tested.

Questi perfluorocarburi ed emulsionanti vengono dispersi in mezzo acquoso fisiologicamente accettabile che comprende anche soluzioni isotoniche quali la soluzione di Tyrode, la soluzione di Ringer, soluzione di Ringer addizionata con lattato o ad esempio contenente glucosio, la soluzione Bretschneider. In alcuni casi il mezzo acquoso comprende adiuvanti dell'agente emulsionanate, quali ad esempio tracce di acidi grassi. These perfluorocarbons and emulsifiers are dispersed in a physiologically acceptable aqueous medium which also includes isotonic solutions such as Tyrode's solution, Ringer's solution, Ringer's solution added with lactate or for example containing glucose, Bretschneider's solution. In some cases the aqueous medium comprises emulsified agent adjuvants, such as, for example, traces of fatty acids.

US 3,962,439 descrive un sangue artificiale comprendente un perfluorocarburo costituito da 9-11 atomi di carbonio, avente una dimensione di particelle di 50-300 nm e comprendente anche un fosfolipide come emulsionante ed un acido grasso, un suo sale o un estere monoglicerico di detto acido grasso come adiuvante dell'agente emulsionante. Il perfluorocarburo e' presente ad una concentrazione del 10-40% (peso/volume) in una emulsione comprendente il 2-6% (peso/volume) di un fosfolipide e minori quantità' di un acido grasso. US 3,962,439 describes an artificial blood comprising a perfluorocarbon consisting of 9-11 carbon atoms, having a particle size of 50-300 nm and also comprising a phospholipid as an emulsifier and a fatty acid, a salt thereof or a monoglycerine ester of said acid fat as an adjuvant to the emulsifying agent. The perfluorocarbon is present at a concentration of 10-40% (weight / volume) in an emulsion comprising 2-6% (weight / volume) of a phospholipid and smaller quantities of a fatty acid.

US 4,397,870 descrive un procedimento per prolungare la stabilita' di composti perfluorurati in animali e nell'uomo, che comprende l'iniezione di un agente emulsionante nell'individuo perfuso. Il brevetto riporta una concentrazione di composto perfluorurato del 15-40% in volume rispetto al volume totale della miscela, che corrisponde al 30-75% in peso rispetto al volume totale della miscela, ed una concentrazione del 7-9% in peso di lecitina rispetto al volume totale. US 4,397,870 discloses a process for prolonging the stability of perfluorinated compounds in animals and humans, which comprises the injection of an emulsifying agent into the perfused individual. The patent reports a concentration of perfluorinated compound of 15-40% by volume with respect to the total volume of the mixture, which corresponds to 30-75% by weight with respect to the total volume of the mixture, and a concentration of 7-9% by weight of lecithin. compared to the total volume.

US 4,423,077 descrive una emulsione stabile in un mezzo acquoso fisiologicamente accettabile; l'emulsione ha un contenuto del 30-75% in peso di composti fluorurati per volume totale di emulsione ed una concentrazione di 7-9% in peso per volume di emulsione di un lipide che riveste il composto perfluorurato. Questa emulsione ha una dimensione di particelle di circa 100 nm ed il 95% delle particelle ha un diametro inferiore a 200 nm. WO 9100110 descrive emulsioni che comprendono il 25-125% in peso rispetto al volume dell'emulsione di perfluorocarburi in fase acquosa ed un fosfolipide con legami carbonio-carbonio saturi come emulsionante. US 4,423,077 describes a stable emulsion in a physiologically acceptable aqueous medium; the emulsion has a content of 30-75% by weight of fluorinated compounds by total volume of emulsion and a concentration of 7-9% by weight by volume of emulsion of a lipid which coats the perfluorinated compound. This emulsion has a particle size of approximately 100 nm and 95% of the particles have a diameter of less than 200 nm. WO 9100110 discloses emulsions comprising 25-125% by weight with respect to the volume of the aqueous phase perfluorocarbon emulsion and a phospholipid with saturated carbon-carbon bonds as emulsifier.

EP 415263 si riferisce ad una emulsione stabile e conservabile, stabilizzata per aggiunta di composti eterociclici periluorurati aggiunti all'emulsione contenente perfluorocarburi. EP 415263 refers to a stable and conservable emulsion, stabilized by the addition of perylurinated heterocyclic compounds added to the emulsion containing perfluorocarbons.

EP 415264 rivendica composti eterociclici cicloalchilsostituiti perfluorurati e loro miscele come trasportatori di ossigeno nei sostituti del sangue, in quanto la cicloesilmorfolina e' adatta a generare emulsioni di seconda generazione per quanto riguarda la velocita' di eliminazione, la solubilità' di ossigeno e la stabilita' delle dimensioni delle particelle. EP 415264 claims perfluorinated cycloalkyl substituted heterocyclic compounds and their mixtures as oxygen carriers in blood substitutes, as cyclohexylmorpholine is suitable for generating second generation emulsions as regards the elimination rate, oxygen solubility and stability. particle size.

WO 9409625 riguarda emulsioni di fluorocarburi stabili allo stoccaggio che contengono una minore quantità' di un composto perfluorocarburico stabilizzante il cui peso molecolare e' maggiore di quello del perfluorocarburo principale e comprende una parte lipofila per garantire la stabilita' delle dimensioni delle particelle (ad esempio perfluorodecilbromuro/perfluorodecalina). WO 9409625 relates to storage stable fluorocarbon emulsions which contain a smaller amount of a stabilizing perfluorocarbon compound whose molecular weight is greater than that of the main perfluorocarbon and includes a lipophilic part to ensure the stability of the particle size (e.g. perfluorodecylbromide / perfluorodecaline).

WO 9414415 si riferisce ad emulsioni per la somministrazione di farmaci che consistono in un perfluorocarburo, perfluorooctilbromuro, lecitina, olio di cartamina, glicerolo ed il farmaco desiderato. WO 9414415 relates to drug delivery emulsions which consist of a perfluorocarbon, perfluorooctylbromide, lecithin, cartamine oil, glycerol and the desired drug.

FR 2700696 descrive emulsioni a base di perfluorocarburi contenenti iodio come mezzo di contrasto. Queste contengono uno iodioperfluoroalchilalcano come trasportatore di ossigeno, lecitina come emulsionante e una soluzione salina tamponata fisiologicamente accettabile. FR 2700696 describes perfluorocarbon based emulsions containing iodine as a contrast medium. These contain an iodine perfluoroalkylalkane as an oxygen carrier, lecithin as an emulsifier and a physiologically acceptable buffered saline solution.

SCOPI DELL'INVENZIONE AIMS OF THE INVENTION

Scopo del presente trovato e quello di mettere a disposizione dei composti che consentano il trasporto di ossigeno in sistemi biologici. The purpose of the present invention is to provide compounds which allow the transport of oxygen in biological systems.

Ancora scopo del presente trovato e quello di mettere a disposizione dei composti che consentano la realizzazione di fluidi sostitutivi di fluidi biologici. Another object of the present invention is to provide compounds which allow the production of fluids which replace biological fluids.

Sempre scopo del presente trovato e quello di mettere a disposizione dei composti impiegabili come trasportatori/mediatori di ossigeno in sistemi biologici che diano luogo ad emulsioni sostitutive di fluidi biologici che siano stabili anche allo stoccaggio per lunghi periodi. Another object of the present invention is to provide compounds that can be used as oxygen carriers / mediators in biological systems which give rise to substitute emulsions of biological fluids which are also stable in storage for long periods.

Pure scopo del presente trovato e’ quello di mettere a disposizione dei composti che non diano luogo ad un significativo accrescimento delle dimensioni delle particelle nel tempo. Another purpose of the present invention is to make available compounds that do not give rise to a significant increase in the size of the particles over time.

Ancora scopo del presente trovato e quello di mettere a disposizione dei composti che diano luogo ad emulsioni stabili anche in condizioni di sterilizzazione. Another object of the present invention is to provide compounds which give rise to stable emulsions even in sterilization conditions.

Sempre scopo del presente trovato e' quello di mettere a disposizione dei composti che siano chimicamente, fisicamente e fisiologicamente inerti. Another object of the present invention is to provide compounds which are chemically, physically and physiologically inert.

Ancora, scopo del presente trovato e' quello di mettere a disposizione dei composti che non siano tossici, che non siano inquinanti e che risultino compatibili con l'ecosistema . Still another object of the present invention is to make available compounds which are not toxic, which are not polluting and which are compatible with the ecosystem.

Sempre scopo del presente trovato e' quello di mettere a disposizione dei composti che non siano metabolizzabili o catabolizzabili. Another object of the present invention is to provide compounds which are not metabolizable or catabolizable.

Pure scopo del presente trovato e' quello di mettere a disposizione mezzi sostitutivi di fluidi biologici. Another object of the present invention is to provide replacement means for biological fluids.

DESCRIZIONE DELL'INVENZIONE DESCRIPTION OF THE INVENTION

Questi ed altri scopi ancora e relativi vantaggi che meglio saranno chiariti dalla descrizione che segue, vengono raggiunti dall'impiego di fluoroalcani aventi le seguenti formule: These and other objects and related advantages which will be better clarified by the following description, are achieved by the use of fluoroalkanes having the following formulas:

dove R, R^ ed R2 sono uguali o diversi tra loro e scelti tra idrogeno ed alchile - Cg, n e' compreso tra 1 e 20 ed m, q e p sono uguali o diversi tra loro e compresi tra 1 e 8, come trasportatori/mediatori di ossigeno in sistemi biologici. where R, R ^ and R2 are the same or different from each other and selected from hydrogen and alkyl - Cg, n is between 1 and 20 and m, q and p are the same or different from each other and between 1 and 8, as transporters / mediators of oxygen in biological systems.

Detti fluoroalcani di formula (I) e (II) possono essere impiegati anche in miscela tra loro. Said fluoroalkanes of formula (I) and (II) can also be used in admixture with each other.

Inoltre, detti composti (I) e (II) consentono di realizzare emulsioni adatte ad essere impiegate in sostituzione di fluidi biologici, dette emulsioni essendo stabili anche ad elevate temperature ed allo stoccaggio anche per lunghi periodi. Moreover, said compounds (I) and (II) allow to realize emulsions suitable for being used in substitution of biological fluids, said emulsions being stable even at high temperatures and for storage even for long periods.

Detti fluoroalcani sono preparati mediante reazione di un perfluoroalchilioduro con un alfa-alchene o un alfa,omega-alcadiene, seguita da eliminazione di acido iodidrico in mezzo alcalino ed idrogenazione finale mediante H2/Nichel Raney o idrogeno su Platino o tributilstagno idruro , come descritto da H.Meinert in DE 19536504.6. Said fluoroalkanes are prepared by reaction of a perfluoroalkyldide with an alpha-alkene or an alpha, omega-alkadiene, followed by elimination of hydrogen iodide in an alkaline medium and final hydrogenation by H2 / Nickel Raney or hydrogen on Platinum or tributyltin hydride, as described by H. Meinert in DE 19536504.6.

La preparazione di detti fluoroalcani e' semplice da realizzare, garantisce alte rese ed e' economicamente vantaggiosa. Dal momento che i fluoroalcani non contengono atomi di alogeni diversi dal fluoro e non contengono gruppi distruttori di ozono, sono considerati non inquinanti, e sono ritenuti avere una azione ecologicamente benefica, al contrario dei perfluoroalchil alogenuri come, ad esempio, il perfluoroottilbromuro. The preparation of said fluoroalkanes is simple to carry out, guarantees high yields and is economically advantageous. Since fluoroalkanes do not contain halogen atoms other than fluorine and do not contain ozone-destroying groups, they are considered non-polluting, and are believed to have an ecologically beneficial action, unlike perfluoroalkyl halides such as, for example, perfluorooctyl bromide.

Detti fluoroalcani sono vantaggiosamente impiegati, secondo il presente trovato, come trasportatori/mediatori di ossigeno nei sistemi biologici, ad esempio nei sostituti del sangue; detti alcani consentono inoltre di realizzare emulsioni particolarmente adatte a tali scopi . Said fluoroalkanes are advantageously used, according to the present invention, as oxygen transporters / mediators in biological systems, for example in blood substitutes; said alkanes also make it possible to produce emulsions particularly suitable for these purposes.

Si e' visto che le suddette emulsioni che comprendono detti fluoroalcani come componenti principali, sono sterilizzabili a 121 °C per 15 minuti in presenza di ingredienti osmotici senza che si riscontri alcun aumento delle dimensioni delle particelle; inoltre, dette emulsioni, pur comprendendo fino al 100% in peso di detti fluoroalcani per volume di emulsione, si conservano a temperatura ambiente per lunghi periodi, anche piu' di 200 giorni senza che si riscontri un apprezzabile aumento delle dimensioni delle particelle. Inoltre, l'impiego di dette emulsioni comprendenti 80-100% in peso di detti fluoroalcani per volume di emulsione, assicura un quantitativo di ossigeno sufficiente derivante dal contatto con l'atmosfera e non richiede ulteriore aggiunta di ossigeno. It has been seen that the above emulsions, which include said fluoroalkanes as main components, are sterilizable at 121 ° C for 15 minutes in the presence of osmotic ingredients without any increase in the size of the particles; furthermore, said emulsions, although comprising up to 100% by weight of said fluoroalkanes by volume of emulsion, are kept at room temperature for long periods, even more than 200 days without an appreciable increase in the size of the particles. Furthermore, the use of said emulsions comprising 80-100% by weight of said fluoroalkanes per emulsion volume, ensures a sufficient quantity of oxygen deriving from contact with the atmosphere and does not require further addition of oxygen.

Ancora, detti fluoroalcani, essendo chimicamente, fisicamente e fisiologicamente inerti, privi di tossicità , nonché* non inquinanti e quindi compatibili con l'ecosistema, ben si prestano ad essere impiegati, secondo il trovato, come trasportatori/mediatori di ossigeno nei sistemi biologici. Furthermore, said fluoroalkanes, being chemically, physically and physiologically inert, free of toxicity, as well as non-polluting and therefore compatible with the ecosystem, are well suited to be used, according to the invention, as transporters / mediators of oxygen in biological systems.

I suddetti composti inoltre, non eliminano acido fluoridrico intramolecolare e di conseguenza non danno luogo a doppi legami fluorurati; sono composti allo stato liquido o solidi incolori, non sono attaccati da soluzioni fortemente acide o fortemente basiche, da nucleofili e da agenti ossidanti. Inoltre non subiscono metabolizzazione ne catabolizzazione. Moreover, the aforesaid compounds do not eliminate intramolecular hydrofluoric acid and consequently do not give rise to fluorinated double bonds; they are compounds in the liquid state or colorless solids, they are not attacked by strongly acidic or strongly basic solutions, by nucleophiles and by oxidizing agents. They also do not undergo metabolism or catabolization.

I fluoroalcani di formula (I) e (II), presentano punti di fusione e di ebollizione che aumentano con l'aumentare del peso molecolare; mostrano inoltre elevata solubilità* per i gas, ad esempio ossigeno ed anidride carbonica (rispettivamente 40-45% e 130-150% volume/volume) , mentre sono scarsamente solubili in acqua. The fluoroalkanes of formula (I) and (II) have melting and boiling points which increase with increasing molecular weight; they also show high solubility * for gases, for example oxygen and carbon dioxide (respectively 40-45% and 130-150% volume / volume), while they are scarcely soluble in water.

Tramite agenti emulsionanti efficaci (ad esempio fosfolipidi quali lecitina da tuorlo d'uovo, lecitina da semi di soia, polimeri a blocchi ossido di etilene/ossido di propilene quali "Pluronic", tensioattivi fluorurati, cioè* composti costituiti da una testa fluorofila ed una coda idrofila connesse da un ponte) e tecniche di emulsione fisica (ultrasuoni, alta pressione, omogeneizzatori "Ultraturrax"), detti fluoroalcani danno luogo ad emulsioni stabili con particelle di dimensioni medie di 80-300 nm. In particolare, i fluoroalcani con punti di ebollizione preferibilmente compresi tra 100 e 300 °C e tensioni di vapore preferibilmente al di sotto di 50 mbar a circa 37 °C, sono particolarmente adatti ad essere impiegati come trasportatori/mediatori di ossigeno nei sistemi biologici, ad esempio come emulsioni in grado di trasportare ossigeno. Through effective emulsifying agents (for example phospholipids such as lecithin from egg yolk, lecithin from soybeans, ethylene oxide / propylene oxide block polymers such as "Pluronic", fluorinated surfactants, ie * compounds consisting of a fluorophilic head and a hydrophilic tail connected by a bridge) and physical emulsion techniques (ultrasound, high pressure, "Ultraturrax" homogenizers), said fluoroalkanes give rise to stable emulsions with particles of average size of 80-300 nm. In particular, fluoroalkanes with boiling points preferably between 100 and 300 ° C and vapor pressures preferably below 50 mbar at about 37 ° C, are particularly suitable for use as oxygen carriers / mediators in biological systems, for example as emulsions capable of carrying oxygen.

Detti fluoroalcani sono solubili a seconda delle dimensioni delle catene, nei periluorocarburi e nei loro derivati (ad esempio eteri periluorurati ad alto peso molecolare come "Hostinert" e "Fomblin") come pure negli idrocarburi e nei loro derivati (ad esempio paraffine liquide, olii siliconici, esteri o acidi grassi) . La solubilità nei periluorocarburi aumenta con l'aumentare della lunghezza del gruppo alchilico fluorurato, mentre la solubilità negli idrocarburi aumenta con l’aumentare della lunghezza del gruppo alchilico non fluorurato. Detti fluoroalcani possono essere pure impiegati come veicoli per farmaci lipofili. Said fluoroalkanes are soluble depending on the size of the chains, in periluorocarbons and their derivatives (for example high molecular weight periluorinated ethers such as "Hostinert" and "Fomblin") as well as in hydrocarbons and their derivatives (for example liquid paraffins, oils silicone, esters or fatty acids). The solubility in periluorocarbons increases with the increase in the length of the fluorinated alkyl group, while the solubility in hydrocarbons increases with the increase in the length of the non-fluorinated alkyl group. Said fluoroalkanes can also be used as vehicles for lipophilic drugs.

Le densità di detti fluoroalcani (D=l,3 - 1,6 g/cm3) sono inferiori di quelle dei perfluorocarburi (D=l,8 -2,0 g/cm^) per la presenza dei gruppi alchilici non fluorurati. Di conseguenza, le emulsioni che comprendono fluoroalcani con solubilità<1 >di ossigeno confrontabili a quelle dei perfluorocarburi con paragonabile capacita<1 >di trasporto di ossigeno, sono caratterizzati da una densità' inferiore dell'intera emulsione e, di conseguenza, quando impiegati come fluidi sostitutivi del sangue, richiedono una inferiore attività' di contrazione del muscolo cardiaco. La tensione superficiale e' pari a 20-30 mN/m a 20 °C ed e' dovuta alla presenza di almeno un gruppo alchilico fluorurato terminale<'>. The densities of said fluoroalkanes (D = 1.3-1.6 g / cm3) are lower than those of the perfluorocarbons (D = 1.8-2.0 g / cm3) due to the presence of the non-fluorinated alkyl groups. Consequently, emulsions comprising fluoroalkanes with <1> oxygen solubility comparable to those of perfluorocarbons with comparable <1> oxygen carrying capacity, are characterized by a lower density of the entire emulsion and, consequently, when used as blood replacement fluids require less heart muscle contraction activity. The surface tension is equal to 20-30 mN / m at 20 ° C and is due to the presence of at least one terminal fluorinated alkyl group <'>.

Prima di essere impiegati nei sistemi biologici secondo il presente trovato, i fluoroalcani di formula (I) e (II) devono essere sottoposti ad un procedimento di elevata purificazione. Before being used in the biological systems according to the present invention, the fluoroalkanes of formula (I) and (II) must be subjected to a highly purified process.

Il suddetto procedimento comprende il trattamento iniziale dei fluoroalcani con una soluzione acida di permanganato di potassio ed il trattamento in autoclave con una miscela di idrato di potassio acquoso 8N, un nucleofilo (ammina secondaria) e CaO o BaO a 180 °C per un periodo di 72 ore. Successivamente il prodotto e' separato dalla fase acquosa e dalla fase amminica, lavato con acido cloridrico acquoso, poi con una soluzione di NaHCO-j ed acqua distillata, anidrif.icato su Na2S04 e CaCl2 e frazionato per mezzo di una colonna efficiente. La purezza e' controllata mediante metodi spettroscopici, quali ad esempio IR, <1>H-NMR, <19>F-NMR e spettroscopia GC/MS. The above process comprises the initial treatment of the fluoroalkanes with an acid solution of potassium permanganate and the autoclaving with a mixture of 8N aqueous potassium hydrate, a nucleophile (secondary amine) and CaO or BaO at 180 ° C for a period of 72 hours. Subsequently the product is separated from the aqueous and amine phases, washed with aqueous hydrochloric acid, then with a solution of NaHCO-j and distilled water, anhydrified on Na2SO4 and CaCl2 and fractionated by means of an efficient column. The purity is controlled by spectroscopic methods, such as for example IR, <1> H-NMR, <19> F-NMR and GC / MS spectroscopy.

Viene pure effettuato un procedimento per la determinazione quantitativa dei componenti che potenzialmente possono dar luogo ad eliminazione intramolecolare di acido fluoridrico. Il procedimento e* basato sulla determinazione del fluoro tramite elettrodi ionosensibili dopo reazione con esametilendiammina in nonano o decano ad una temperatura di 120 °C. Se non vi e' fluoro determinabile (limite di determinazione IO<-5 >mol/1) , i fluoroalcani altamente purificati ottenuti mediante il suddetto procedimento non mostrano limiti di proliferazione per quanto riguarda la sintesi di proteine e di DNA in colture cellulari HELA, M0LT4 o HEP2 , ne* vengono osservati emolisi o segni di infiammazione provocati da reazioni immunologiche, quali ad esempio attivazione complementare e secrezione di interleuchina. I fluoroalcani di formula (I) e (II) pertanto possono essere vantaggiosamente e con tutta sicurezza impiegati in campo medico, biologico e farmaceutico. A process is also carried out for the quantitative determination of the components which can potentially give rise to intramolecular elimination of hydrofluoric acid. The process is based on the determination of fluorine by ion-sensitive electrodes after reaction with hexamethylenediamine in nonane or decane at a temperature of 120 ° C. If there is no determinable fluorine (limit of determination IO <-5> mol / 1), the highly purified fluoroalkanes obtained by the above procedure do not show proliferation limits as regards the synthesis of proteins and DNA in HELA cell cultures, M0LT4 or HEP2, hemolysis or signs of inflammation caused by immunological reactions, such as complementary activation and interleukin secretion, are observed. The fluoroalkanes of formula (I) and (II) can therefore be advantageously and safely used in the medical, biological and pharmaceutical fields.

A solo titolo indicativo e non limitativo del presente trovato, vengono qui di seguito riportati alcuni esempi di pratica realizzazione. By way of indication only and not of limitation of the present invention, some practical embodiment examples are given below.

Allo scopo di assicurare condizioni operative prive di germi e di endotossine, tutti i componenti del sistema di omogeneizzazione sono trattati con etanolo ad elevata purezza, acido fosforico diluito ed infine sottoposti a lavaggio con acqua priva di endotossine. In order to ensure germ-free and endotoxin-free operating conditions, all components of the homogenization system are treated with high purity ethanol, diluted phosphoric acid and finally washed with endotoxin-free water.

ESEMPIO 1 EXAMPLE 1

Preparazione di un'emulsione comprendente: Preparation of an emulsion comprising:

60% (peso/volume) di <F>i3<C>6<-c>8<H>17 60% (weight / volume) of <F> i3 <C> 6 <-c> 8 <H> 17

4,5% (peso/volume) di Lecitina E 100 (Lipoid KG -Germania) in soluzione lattato Ringer. 4.5% (weight / volume) of Lecithin E 100 (Lipoid KG -Germany) in Ringer's lactate solution.

24 g di perfluoroesilottano vengono aggiunti a 22 mi di una soluzione lattato Ringer e 1,8 g di lecitina E 100 (lecitina da tuorlo d'uovo), sotto continua agitazione. L'agitazione viene mantenuta per 1 ora a temperatura ambiente. Successivamente nella miscela viene introdotto un flusso di azoto per un perido di 30 minuti attraverso un filtro sterile (0,2 micron). La miscela viene preemulsionata mediante trattamento con Ultraturrax IKA T 25 a 20.000 giri al minuto per 2 minuti. 24 g of perfluorohexiloctane are added to 22 ml of a Ringer lactate solution and 1.8 g of lecithin E 100 (egg yolk lecithin), under continuous stirring. The stirring is maintained for 1 hour at room temperature. A stream of nitrogen is then introduced into the mixture for a period of 30 minutes through a sterile filter (0.2 micron). The mixture is pre-emulsified by treatment with Ultraturrax IKA T 25 at 20,000 rpm for 2 minutes.

La miscela omogenea e' omogeneizzata per mezzo di un omogeneizzatore Micron Lab 40 (APV Gaulin, Lubeck -Germania) a 500 bar. The homogeneous mixture is homogenized by means of a Micron Lab 40 homogenizer (APV Gaulin, Lubeck -Germany) at 500 bar.

L'emulsione ottenuta e' un liquido lattiginoso, incoiore. The emulsion obtained is a milky, incoior liquid.

La dimensione media delle particelle, determinata mediante analizzatore della distribuzione di particelle inferiori al micron Autosizer II c (Malvern Instruments Ltd.) e' di 166 nm. The average particle size, determined by Autosizer II c sub micron particle distribution analyzer (Malvern Instruments Ltd.) is 166 nm.

ESEMPIO 2. EXAMPLE 2.

Preparazione di un'emulsione sterile comprendente: Preparation of a sterile emulsion comprising:

19,5% (peso/volume) di <F>i3<c>6<-c>8<F>i7 19.5% (weight / volume) of <F> i3 <c> 6 <-c> 8 <F> i7

0,5% (peso/volume) di perfluorodecalina 0.5% (weight / volume) of perfluorodecaline

1,2% (peso/volume) di Lecitina E 100 1.2% (weight / volume) of Lecithin E 100

Soluzione lattato Ringer. Ringer's lactate solution.

7,8 g di perfluoroesilottano sono aggiunti sotto agitazione continua a 0,2 g di perfluorodecalina, 0,48 g di lecitina E 100 e 24 mi di soluzione lattato Ringer. L'agitazione viene mantenuta per 1 ora a temperatura ambiente. Successivamente, nella miscela viene introdotto azoto gassoso attraverso un filtro sterile (0,2 micron) , la miscela viene quindi preemulsionata per mezzo dello strumento Ultraturrax IKA T 25 (2 minuti, 20.000 giri al minuto) e omogeneizzata mediante un omogeneizzatore Micron Lab 40 (10 volte, 500 bar). 7.8 g of perfluorohexiloctane are added under continuous stirring to 0.2 g of perfluorodecaline, 0.48 g of lecithin E 100 and 24 ml of lactate Ringer's solution. The stirring is maintained for 1 hour at room temperature. Subsequently, nitrogen gas is introduced into the mixture through a sterile filter (0.2 micron), the mixture is then pre-emulsified by means of the Ultraturrax IKA T 25 instrument (2 minutes, 20,000 revolutions per minute) and homogenized by means of a Micron Lab 40 homogenizer ( 10 times, 500 bar).

L'emulsione cosi' preparata e' depositata in provette di vetro, che vengono chiuse mediante setti di teflon/silicone, sigillate e successivamente sterilizzate in corrente di vapore ( 121 °C, 1 bar, 20 minuti). The emulsion thus prepared is deposited in glass test tubes, which are closed by means of Teflon / silicone septa, sealed and subsequently sterilized in a stream of steam (121 ° C, 1 bar, 20 minutes).

La dimensione media delle particelle, determinata come nell'esempio 1, e' pari a 170 nm il giorno della preparazione, dopo 100 giorni di stoccaggio a temperatura ambiente e' pari a 190 nm e dopo 140 giorni e' pari a 230 nm. The average particle size, determined as in example 1, is equal to 170 nm on the day of preparation, after 100 days of storage at room temperature it is equal to 190 nm and after 140 days it is equal to 230 nm.

ESEMPIO 3 EXAMPLE 3

58,5% (peso/volume) di <F>i3<C>6<-c>a<H>l758.5% (weight / volume) of <F> i3 <C> 6 <-c> a <H> l7

1,5% (peso/volume) di periluorofluorene 1.5% (weight / volume) of peryluorofluorene

4,5% (peso/volume) di lecitina E 100 4.5% (weight / volume) of lecithin E 100

Soluzione lattato Ringer. Ringer's lactate solution.

23,4 g di perfluoroesilottano vengono aggiunti, sotto continua agitazione, a 0,6 g di perfluorofluorene, 22,15 mi di soluzione lattato Ringer e 1,8 g di lecitina E 100. L'agitazione viene mantenuta per 1 ora a temperatura ambiente. Successivamente nella miscela viene introdotto azoto gassoso tramite un filtro sterile (0,2 micron), la miscela viene preemulsionata tramite lo strumento Ultraturrax IKA T 25 (2min, 20.000 giri al minuto) e omogeneizzata mediante omogeneizzatore Micron Lab 40 (10 volte, 500 bar). La dimensione media delle particelle, determinata come nell'Esempio 1, e' pari a 110 nm determinata nel giono di preparazione. ESEMPIO 4 23.4 g of perfluorohexiloctane are added, under continuous stirring, to 0.6 g of perfluorofluorene, 22.15 ml of Ringer's lactate solution and 1.8 g of lecithin E 100. The stirring is maintained for 1 hour at room temperature . Subsequently, nitrogen gas is introduced into the mixture through a sterile filter (0.2 micron), the mixture is pre-emulsified using the Ultraturrax IKA T 25 instrument (2min, 20,000 rpm) and homogenized by means of a Micron Lab 40 homogenizer (10 times, 500 bar ). The average particle size, determined as in Example 1, is equal to 110 nm determined in the preparation day. EXAMPLE 4

78.0% (peso/volume) di <F>i3<c>6<-c>3<H>i778.0% (weight / volume) of <F> i3 <c> 6 <-c> 3 <H> i7

2,0% (peso/volume) di periluorodecalina 2.0% (weight / volume) of periluorodecaline

6,0% (peso/volume) di lecitina E 100 6.0% (weight / volume) of lecithin E 100

Soluzione lattato Ringer. Ringer's lactate solution.

31,2 g di periluoroesilottano sono aggiunti, sotto agitazione continua, a 0,8 g di perfluorodecalina, 2,4 g di lecitina E 100 e 16 mi di soluzione lattato Ringer. L'agitazione e' mantenuta per 1 ora a temperatura ambiente. Nella miscela viene introdotto azoto gassoso attraverso un filtro sterile (0,2 micron), la miscela viene preemulsionata tramite Ultraturrax IKA T 25 (2 min, 20.000 giri al minuto) ed omogeneizzata mediante omogeneizzatore Micron Lab 40 (10 volte, 500 bar). La dimensione media delle particelle, determinata come nell'esempio 1, e' di 100 nm determinata il giorno della preparazione. 31.2 g of periluorohexiloctane are added, under continuous stirring, to 0.8 g of perfluorodecaline, 2.4 g of lecithin E 100 and 16 ml of Ringer's lactate solution. The stirring is maintained for 1 hour at room temperature. Gaseous nitrogen is introduced into the mixture through a sterile filter (0.2 micron), the mixture is pre-emulsified with Ultraturrax IKA T 25 (2 min, 20,000 rpm) and homogenized with a Micron Lab 40 homogenizer (10 times, 500 bar). The average particle size, determined as in Example 1, is 100 nm determined on the day of preparation.

Claims

CLAIMS 1. Use of fluoroalkanes having the following formulas: F (CF2) n- (CHR) (CHR1) q- (CHR2) pH (I) F (CF2) n- (CHR) m- (CHRl) q- (CHR2) p- (CF2) nF (II) where R, R ^ and R2 are the same or different from each other and selected from hydrogen and C-j ^ - C8 alkyl, n is between 1 and 20 and m, q and p are between 1 and 8, as transporters / mediators of oxygen in biological systems .

2. Use of a fluoroalkane having the following formula: <F> 13 <C> 6 <C> 8 <H> 17 (III) as an oxygen transporter / mediator in biological systems.

3. Use of fluoroalkanes (I) and (II) according to claim 1, said alkanes being in admixture with each other.

4. Use of fluoroalkanes (I) and (II) according to claim 1, said biological systems being blood substitute fluids.

5. Use of fluoroalkanes (I) and (II) according to claim 1, said biological systems being "resuscitative" fluids.

6. Process for the purification of fluoroalkanes having the following formula: <F> (CF2) n- (<CHR>) m <"> i <CHR> 1) q <"> (<CHR> 2> p <H> (I) <F> (<CF> 2) n - <<CHR>> m- (<CHR> l) q- (<CHR> 2) p- (<CF> 2) n <F> (II) where R, Rj ^ and R2 are the same or different from each other and selected from hydrogen and alkyl - C8, n is comprised between 1 and 20 and m, q and p are comprised between 1 and 8, characterized in that it comprises the following steps: - treatment of said fluoroalkanes with an oxidizing acid solution; - subsequent autoclave treatment in a basic environment, in the presence of a nucleophilic agent and an oxide of an alkaline earth metal, to obtain a partially purified product; - separation of said partially purified product, subsequent washing and drying of said product; - fractionation of said product on a column, obtaining a highly purified product.

7. Process according to claim 6, characterized in that said acid solution is a potassium permanganate solution, said autoclave treatment is carried out at 180 ° C for 72 hours in an 8N potassium hydrate solution, in the presence of a secondary amine as the nucleophile and CaO or BaO as oxides.

8. A process for checking the stability * of fluoroalkanes which includes the following steps: - reaction of a fluoroalkane (I) and (II) according to claim 1 with hexamethylenediamine; - determination of fluorine by ion-sensitive electrodes.

9. Process according to claim 8, characterized in that said reaction with hexamethylenediamine is carried out in nonane or decane at a temperature of 120 ° C.

10. Emulsion comprising fluoroalkanes of formula (I) and (II) having the following general formula: F (CF2) n- (CHR) (CHR1) q- (CHR2) pH (I) <F> (CF2) n- (CHR) m- (CHR1) q- (CHR2) p- (CF2) nF (II) where R, R- ^ and R2 are the same or different from each other and selected from hydrogen and alkyl C. ^ - Cg, n and * between 1 and 20 and m, q and p are between 1 and 8, in addition to the usual additives and / or various components.

11. Emulsion according to claim 10, characterized in that it comprises said fluoroalkanes (I) and (II) in admixture with each other.

12. Emulsion according to claim 10, characterized in that it comprises up to 100% by weight of said fluoroalkanes with respect to the emulsion volume.

13. Emulsion according to claim 10, characterized in that it comprises emulsifying agents.

14. Emulsion according to claim 13, characterized in that it comprises from 20 to 100% by weight of said fluoroalkanes and from 0.5 to 8.0% by weight of said emulsifying agents, by volume of emulsion.

15. Emulsion according to claim 13, characterized in that said emulsifying agents are selected from phospholipids, lecithins, fluorinated surfactants, ethylene oxide / propylene oxide block polymers.

16. Emulsion according to claim 13, characterized in that said emulsifying agents consist of a mixture of a biocompatible ethylene oxide - propylene oxide block polymer and of an additional stabilizer such as a phospholipid or a triglyceride.

17. Emulsion according to claim 16, characterized in that said additional stabilizer is a non-ionic fluorinated surfactant consisting of a hydrophilic head and a fluorophilic tail connected by a bridge.

18. Emulsion according to claim 15, characterized in that it comprises from 80 to 100% by weight of said fluoroalkanes and from 6 to 8% by weight of said phospholipids with respect to the emulsion volume.

19. Emulsion according to claim 10, characterized in that it comprises perfluorocarbons.

20. Emulsion according to claim 19, characterized in that said perfluorocarbons are selected from perfluorodecaline, perfluorofluorene.

21. Emulsion according to claim 10, characterized in that said fluoroalkanes have boiling points between 100 and 300 ° C and vapor pressures lower than 50 mbar at 37 ° C.

22. Emulsion according to claim 10, characterized in that it comprises <F> 13 <C> 6-CgHi? as fluoroalkane of formula (I).

23. Use of an emulsion according to claim 10 as a substitute for biological fluids.

24. Use of an emulsion according to claim 23, said biological fluids being blood replacement fluids.

25. Use of an emulsion according to claim 23, said biological fluids being "resuscitative" fluids.

26. Use of an emulsion according to claim 25, said "resuscitative" fluids being used in organ banks and organ transplantation.

27. Use of an emulsion according to claim 23, as a diagnostic means for tomography <19> F-NMR, sonography, X-ray computed tomography.

28. Use of an emulsion according to claim 23 as a cardioplegic agent for oxygenation of ischemic tissues, as a perfusion medium in open heart surgery, and in cancer therapy as an oxygen carrier in radio and chemo therapy.

29. Use of an emulsion according to claim 23 as a transfusion medium.

30. Use of an emulsion according to claim 23 in case of insufficient blood flow to the heart.

31. Use of an emulsion according to claim 23 as an oxygenating emulsion.

32. Use of an emulsion according to claim 23 as an oxygenating medium for cell and tissue cultures.

33. Biological fluid replacement fluids comprising fluoroalkanes having the following formula <F> (<CF> 2) n <"> (<CHR>> m <"> <<CHR> l) q <"> (<CHR> 2> p <H> (<1>) <F> (<CF> 2> n <"> <<CHR>) m <“> (<CHR> 1> q- (<CHR> 2) p <“> (<1 CF> 2> n <F > (: <11>> where R, R1 and R2 are the same or different from each other and selected from hydrogen and alkyl - Cg, n is between 1 and 20 and m, q and p are between 1 and 8.

34. Blood replacement fluids comprising fluoroalkanes having the following formula F (CF2) n "(CHR) m“ (CHRi) q “(CHR2) pH (1) F (cF2) n- (CHR) m- (CHRl) q- (CHR2) p“ iCF2) nF UD where R , R1 and R2 are the same or different from each other and selected from hydrogen and C1 - Cg alkyl, n is between 1 and 20 and m, g and p are between 1 and 8.

35. "Resuscitative" fluids comprising fluoroalkanes having the following formula F (CF2) n- (CHR) m- (CHR!) Q- (CHR2) pH (I) F (CF2) n "(CHR) m" (CHRi) q- (CHR2) p- (CF2) nF (II) where R, R! and R2 are the same or different from each other and selected from hydrogen and alkyl C ^ - Cg, n is comprised between 1 and 20 and m, q and p are comprised between 1 and 8.

36. Use of fluoroalkanes (I) and (II) of claim 1 as vehicles for lipophilic drugs.