A Hemostatic Tissue Adhesive Composition Comprising An NCO-Terminated Urethane Preprolymer

Abstract

The invention relates to a NCO-terminated urethane preprolymer based on at least a polyol oligomer and a diisocyanate, wherein the polyol oligomer is polyhydroxyalkanoate-diol oligomers (oligoPHA-diol) having a hydroxyl value of more than or equal to 149 mgKOH/g, and the diisocyanate is selected from the list consisting of 1,4-butane diisocyanate (BDI), hexamethylene diisocyanate (6-HDI), dimeryl diisocyanate (DDI), pentamethylene diisocyanate (PDI), L-lysine diisocyanate (LDI), 1,7-heptamethylene diisocyanate (7-HDI), and mixtures thereof. The invention also relates to a method for preparing such a NCO-terminated urethane prepolymer and for a method for preparing polyhydroxyalkanoate-diol oligomers (oligoPHA-diol). The invention also relates to a hemostatic tissue adhesive composition and a hemostatic tissue adhesive. The invention also relates to a kit for the preparation of a hemostatic tissue adhesive.

Claims

1. A NCO-terminated urethane preprolymer based on at least a polyol oligomer and a diisocyanate, wherein the polyol oligomer is polyhydroxyalkanoate-diol oligomer (oligoPHA-diol) having a hydroxyl value of more than or equal to 149 mgKOH/g, and the diisocyanate is selected from the list consisting of 1,4-butane diisocyanate (BDI), hexamethylene diisocyanate (6-HDI), dimeryl diisocyanate (DDI), pentamethylene diisocyanate (PDI), L-lysine diisocyanate (LDI), 1,7-heptamethylene diisocyanate (7-HDI), and mixtures thereof.

2. The NCO-terminated urethane preprolymer of claim 1, wherein the oligoPHA-diol are obtained by transesterification of a polyhydroxyalkanoate (PHA) with a reactive solvent selected in the group consisting of 1,2-ethylene glycol, 1,3-propanediol (PDO), 1,4-butanediol (BDO), or mixture thereof.

3. The NCO-terminated urethane preprolymer of claim 1 or 2, wherein the oligoPHA-diol have a hydroxyl value of between 149 mgKOH/g and 560 mgKOH/g, advantageously of between 149 mgKOH/g and 375 mgKOH/g, more advantageously of between 160 mgKOH/g and 375 mgKOH/g, in particular of between 224 mgKOH/g and 375 mgKOH/g.

4. The NCO-terminated urethane preprolymer of any one of claims 1 to 3, wherein the PHA is selected from the group consisting of poly-3-hydroxybutyrate (P3HB), poly-4-hydroxybutyrate (P4HB), poly-3-hydroxyvalerate (P3HV), poly-3-hydroxypropionate (P3HP), poly-4-hydroxyvalerate (P4HV), poly-5-hydroxyvalerate (P5HV), and mixtures thereof, advantageously poly-3-hydroxybutyrate (P3HB), poly-4-hydroxybutyrate (P4HB) or mixtures thereof, more advantageously poly-3-hydroxybutyrate (P3HB).

5. The NCO-terminated urethane preprolymer of any one of claims 1 to 4, wherein the diisocyanate is a mixture of dimeryl diisocyanate (DDI) with a second diisocyanate selected in the group consisting of 1,4-butanediisocyanate (BDI), pentamethylene diisocyanate (PDI), L-lysine diisocyanate (LDI), hexamethylene diisocyanate (6-HDI), 1,7-heptamethylene diisocyanate (7-HDI), and mixtures thereof, in a content ratio (BDI, PDI, LDI, 6-HDI, 7-HDI or mixtures thereof)/(DDI) of between 0,1/99,9 and 99,9/0,1, advantageously between 20/80 and 80/20 or advantageously between 25/75 and 0,1/99,9.

6. The NCO-terminated urethane preprolymer of claim 5, wherein the second diisocyanate is pentamethylene diisocyanate (PDI), hexamethylene diisocyanate (6-HDI), or mixtures thereof, advantageously hexamethylene diisocyanate (6-HDI).

7. The NCO-terminated urethane preprolymer of any one of claims 1 to 6, wherein the NCO-terminated urethane prepolymer has a NCO:OH molar ratio of between 1 and 3, advantageously 2.

8. The NCO-terminated urethane preprolymer of any one of claims 1 to 7, wherein the NCO-terminated urethane preprolymer has a free NCO content of between 4 and 15% by weight, in relation to the total weight of the prepolymer.

9. The NCO-terminated urethane preprolymer of any one of claims 1 to 8, wherein the NCO-terminated urethane preprolymer has a viscosity of between 5 and 120 Pa.Math.s, advantageously between 5 and 60 or advantageously between 70 and 100 Pa.Math.s, the viscosity being measured at temperature at a temperature of 25? C. by shear viscosity between parallel plates.

10. A method for preparing polyhydroxyalkanoate-diol oligomers (oligoPHA-diol) having a hydroxyl value of more than or equal to 149 mgKOH/g, comprising the following steps: a) Heating a reactive solvent at a temperature of more than or equal to the melting point of the PHA and below the reactive solvent boiling point, in particular of between 170? C. and 190? C., advantageously of 180? C., under inert gaz flow, the reactive solvent being selected in the group consisting of 1,2 ethylene glycol, 1,3-propanediol, 1,4-butanediol, and mixtures thereof, preferably 1,4-butanediol; b) Adding dried PHA and stirring; c) Starting the reaction by adding a catalyst; d) Allowing to react for a period of between 15 and 240 minutes, advantageously between 100 and 215 minutes, typically between 205 and 215 minutes; e) Precipitating the mixture obtained after step d) and washing; and f) Recovering the oligoPHA-diol by distillation at a temperature of between 120? C. and 180? C., advantageously between 150? C. and 180? C., typically between 140? C. and 160? C., under reduced pressure.

11. The method of claim 10, wherein the molar ratio (reactive solvent)/(PHA) is of between 2000 and 12000, advantageously between 2000 and 7000 and more advantageously between 5000 and 7000.

12. The method of claim 10 or 11, wherein the PHA is selected from the group consisting of poly-3-hydroxybutyrate (P3HB), poly-4-hydroxybutyrate (P4HB), poly-3-hydroxyvalerate (P3HV), poly-3-hydroxypropionate (P3HP), poly-4-hydroxyvalerate (P4HV), poly-5-hydroxyvalerate (P5HV), and mixtures thereof, advantageously poly-3-hydroxybutyrate (P3HB), poly-4-hydroxybutyrate (P4HB) or mixtures thereof, more advantageously poly-3-hydroxybutyrate (P3HB).

13. A method for preparing NCO-terminated urethane prepolymer as defined in any one of claims 1 to 9, comprising the following steps: a) Mixing together the diisocyanate with the polyhydroxyalkanoate-diol oligomer (oligoPHA-diol) at a temperature of between 20? C. and 110? C., under stirring, the diisocyanate being selected from the list consisting of 1,4-butanediisocyanate (BDI), hexamethylene diisocyanate (6-HDI), dimeryl diisocyanate (DDI), pentamethylene diisocyanate (PDI), L-lysine diisocyanate (LDI), 1,7-heptamethylene diisocyanate (7-HDI), and mixtures thereof, and the oligoPHA-diol having a hydroxyl value of more than or equal to 149 mgKOH/g and/or being obtained according to the method of any one of claims 10 to 12; b) Optionally, when a mixture of DDI with a second diisocyanate is used, adding in step a) the DDI and in step b) adding dropwise the second diisocyanate being selected from the list consisting of 1,4-butanediisocyanate (BDI), pentamethylene diisocyanate (PDI), L-lysine diisocyanate (LDI), hexamethylene diisocyanate (6-HDI), 1,7-heptamethylene diisocyanate (7-HDI), and mixtures thereof; c) Allowing to react until obtaining a free NCO content of between 4 and 18% by weight, in relation to the total weight of the prepolymer, measured by determining the free NCO content by an indirect-titration method. d) Recovering the NCO-terminated urethane prepolymer.

14. A hemostatic tissue adhesive composition comprising A) an NCO-terminated urethane preprolymer as defined in any one of claims 1 to 9 or as obtained according to the method of claim 13, and B) a chain extender.

15. The hemostatic tissue adhesive composition of claim 14, wherein the chain extender B) is selected from the group consisting of N-ethyldiethanolamine, N-butyldiethanolamine, 1,4-butanediol, polyethylene glycol, polypropylene glycol, 1-amino-4-butanol and mixtures thereof, advantageously 1,4-butanediol.

16. The hemostatic tissue adhesive composition of claims 14 or 15, wherein the NCO:OH molar ratio in the composition is of between 1 et 2, advantageously between 1 and 1,5, preferably between 1 and 1,2.

17. A hemostatic tissue adhesive obtained by reacting A) the NCO-terminated urethane prepolymer as defined in any one of claims 1 to 9 or as obtained by the method of claim 13; and B) a chain extender advantageously selected from the group consisting of N-ethyldiethanolamine, N-butyldiethanolamine, 1,4-butanediol, polyethylene glycol, polypropylene glycol, 1-amino-4-butanol, and mixtures thereof, more advantageously 1,4-butanediol.

18. A kit for the preparation of a hemostatic tissue adhesive comprising a composition A comprising an NCO-terminated urethane preprolymer as defined in any one of claims 1 to 9 or as obtained according to the method of claim 13, and a composition B comprising a chain extender advantageously selected from the group consisting of ethyldiethanolamine, N-butyldiethanolamine, 1,4-butanediol, polyethylene glycol, polypropylene glycol, 1-amino-4-butanol, and mixtures thereof, more advantageously 1,4-butanediol, the compositions A and B being packaged separately and being parenterally administrable simultaneously, sequentially or separately.

19. Use of a hemostatic tissue adhesive composition as defined in any one of claims 14 to 16 for the preparation of a hemostatic tissue adhesive.

Description

DESCRIPTION OF FIGURES

[0105] FIG. 1. represents the OligoPHB-diols M.sub.n variation in relation to the reaction time using multiple parameters: a) DBTL molar equivalents (0.7 (rond), 1.5 (triangle), 2.2 (losange), 2.9 (carr?)), at T=180? C. and 6000 molar equivalent BDO (FIG. 1A); b) BDO molar equivalents (3000 (rond), 6000 (triangle), 12000 (losange), 18000 (carre)), at T=180? C. and 2.2 molar equivalents DBTL (FIG. 1B); c) temperature (150? C. (rond), 160? C. (triangle), 180? C. (losange)), with 2.2 molar equivalent DBTL and 6000 molar equivalents BDO (FIG. 1C); d) short diol length (ETG (rond), PDO (triangle), BDO (losange)), T=180? C., 2.2 molar equivalent DBTL and 6000 molar equivalents BDO (FIG. 1D).

[0106] FIG. 2. represents a).sup.1H, b).sup.13C and c).sup.31P NMR spectra of 450 g.Math.mol.sup.?1 oligoPHB-diol in CDCl.sub.3.

[0107] FIG. 3. represents results of FTIR-ATR analysis of OligoPHB-diols with various Mn. From top to bottom, PHB with decreasing Mn: 260 000 g.Math.mol.sup.?1, 1000 g.Math.mol.sup.?1, 450 g.Math.mol.sup.?1 and 3000 g.Math.mol.sup.?1.

[0108] FIG. 4. represents .sup.1H-NMR spectra of a) neat OligoPHB-diol and b) DDI-0% to f) DDI-100% prepolymers, respectively.

[0109] FIG. 5. represents the results of FTIR-ATR analysis from top to bottom: neat OligoPHB-diol, DDI-0% to DDI-100% prepolymers, respectively.

[0110] FIG. 6. represents the evolution of the free NCO groups content (in %) during a) the reaction between prepolymers with DDI-0%, DDI-25%, DDI-50%, DDI-75% or DDI-100% and BDO and b) DDI-50% prepolymer with different chain extenders (BDA, ABO, BDO, PDO and EtG).

[0111] FIG. 7. represents adhesion of the two-component tissue adhesives on muscle tissues for a) all prepolymers (DDI-0%, DDI-25%, DDI-50%, DDI-75% or DDI-100%) with BDO, b) DDI-50% with different chain extenders (BDA, ABO, BDO, PDO and EtG) and c) DDI-50% with BDO on different substrates (Fresh bovine muscle, liver tissue, and porcine skin tissues).

[0112] FIG. 8. represents model TPUs tensile properties of a) all prepolymers (DDI-0%, DDI-25%, DDI-50%, DDI-75% or DDI-100%) with BDO and b) DDI-50% with different chain extenders (BDA, ABO, BDO, PDO and EtG).

EXAMPLES

Materials.

[0113] PHB L88 (Mn=180 000 g/mol, D=2.3 (by SEC)) was kindly supplied by Biocycle, Brazil. IFABOND? was kindly offered by Peters Surgical. 1,4 Butanediol (BDO, 99%) was purchased from Alfa Aesar. Dibutyltin dilaurate (DBTL, 95%), Hexamethylene diisocyanate (HDI, >99%), dibutylamine (>99.5%), bromophenol blue were purchased from Sigma Aldrich. The PHB and the BDO were dried in an oven under vacuum at 40? C. overnight prior to use. Dimeryl diisocyanate (DDI) was kindly supplied by Cognis. Petroleum ether was purchased from VWR.

General Methods and Analysis.

[0114] .sup.1H-NMR spectra in CDCl.sub.3 were implemented with a Bruker 400 MHz spectrophotometer. .sup.1H-NMR calibration was based on the CDCl.sub.3 chemical shift (?H=7.26 ppm).

[0115] Average molar mass (Mn), average mass molar mass (M.sub.n) and the polydispersity (D) were measured by Size Exclusion Chromatography (SEC) using an Acquity APC apparatus from Waters in THE (0.6 mL/min) at 40? C. Three columns (Acquity APC XT 450 ? 2.5 lm 4.69150 mm, 200 and 45) were connected. The calibration was performed using PS standards.

[0116] The hydroxyl index (IOH) was obtained following DIN 53240-2: 2007-11 (2007), or ASTM E1899-02 (2002).

[0117] The free NCO content was obtained by an indirect titration method using DIN 53185, 16945 (1994) or ASTM D1638 (1985)

[0118] Shear viscosity of the oligoPHA-diol and of the prepolymers was measured at 25? C. using a TA Instrument Discovery Hybrid Rheometer HR-3 equipped with 20 mm parallel plates. The frequency range was from 1?10.sup.5 to 100s.sup.?1 and the gap was 500 ?m.

[0119] FTIR-ATR was performed on a Nicolet 380 spectrometer equipped with an ATR diamond module. The spectra were collected with 32 scans.

[0120] Curing times were determined using FTIR-ATR and performed on a Nicolet 380 spectrometer with an ATR diamond module. Both components (prepolymer-BDO) were stabilized one night at 25? C. and then mixed together for 15 s. A drop of mixture was then placed on the ATR diamond module. Spectra were acquired after 30 s, 2, 4, 6, 8, 10, 15, 20, 30 min and 1, 2, 4 and 24 h. The NCO conversion (curing time) was determined following the NCO end-groups peak disappearance at 2255 cm.sup.?1. The remaining free NCO content was then calculated using the NCO peak area at (A(t)) relative to the NCO peak area at t=0 (A(0)), according to Equation 2:

% NCO(t)=A(t)/A(0)?100(2)

[0121] Adhesion strength in contact with fresh bovine muscle was evaluated by lap shear test, in analogy to the revised ASTM F2255-05 (2005). Strips of tissues of approximately 20?10?3 mm were prepared. Approximatively, 0.2 g of adhesive mixture (prepolymer mixed with BDO) were spread on around 100 mm.sup.2 tissue surfaces with a spatula, and two pieces of treated strips were then put in contact over the 100 mm.sup.2, manually pressed together to assure a good contact for few seconds without specific high pressure, to assure inter-adhesion. These specimens were kept at room temperature in an oven with 100% of relative humidity (RH) for 24 h in order to allow curing of the adhesive mixture to the tissue. The evaluations of the adhesion were conducted on a TA Instrument Discovery Hybrid Rheometer HR-3 equipped with the film tension accessory in tensile mode. The crosshead speed was of 5 mm/min. IFABOND? was used as a reference. For the preparation of IFABOND-based specimens, few drops of glues were directly applied on tissue surfaces and two treated strips were pressed together in analogy with the two-component adhesive mixture. Reference tests were performed within the hour after the sample preparation. For each tested adhesive mixture and IFABOND?, sets of minimum five experiments were conducted. The adhesive bond strength (Rmax, in Pa) was obtained by dividing the maximum shear force before failure (in Newtons) by the adhered area (in m.sup.2). Results are presented as the mean value with standard deviation.

[0122] Uniaxial tensile tests were carried out on the TPU materials using an Instron 5567H (USA) machine equipped with a 10 kN load cell. Experiments were measured at room temperature with a constant crosshead speed of 20 mm.Math.min.sup.?1. Sets of five dumbbell-shaped samples with dimensions of approximately 45?5?1 mm.sup.3 were tested. Average Young's modulus, tensile strength at break (?.sub.max) and elongation at break (?.sub.max) were finally determined.

Example 1: PHB-Diol Oligomers (oligoPHB-Diol) Synthesis According to the Invention and Characterization

[0123] Short oligoPHB-diol were synthesized at 180? C. by transesterification reaction of high molar mass PHB using biobased reactive solvent, mostly BDO, with DBTL as catalyst. Reduced catalyst concentrations were used compared to previous studies in order to work in a greener way. Transesterification reaction is described in Scheme 1 and parameters with graphs. Obtained chemical structures were checked and confirmed by .sup.1H NMR and FTIR. The molar masses, D were determined by SEC in THF.

Example of Synthesis

[0124] 1,4 Butanediol (1,4-BDO) (6000 molar equivalents) was heated to 180? C. under argon flow and magnetically stirred. Then, dried PHB (1 molar equivalent) was added and the mixture was stirred until complete PHB complete dissolution. The reaction was started by adding the catalyst, DBTL (2.2 molar equivalent) at 180? C. and the reaction was stopped after 3 h30 by cooling. Precipitation with petroleum ether and several washings with large volumes of petroleum ether were carrying out to remove the catalyst. The trapped petroleum ether was then separated from the mixture using a centrifuge 5804 from Eppendorf, France. Finally, PHB-diol oligomers of 300 g.Math.mol-1 were recovered by 1,4 BDO distillation at 160? C. under reduced pressure. PHB-diol oligomers were dried under vacuum in an oven at 40? C. for at least 12 h before use.

[0125] For other M.sub.n using multiple parameters, reaction times are referred in the graphic from FIG. 1. As it can be seen, oligoPHB-diols M.sub.n decrease over time with a) DBTL molar equivalents (T=180? C. and 6000 molar equivalent BDO) (FIG. 1A), b) BDO molar equivalents (T=180? C. and 2.2 molar equivalents DBTL) (FIG. 1B), c) temperature (with 2.2 molar equivalent DBTL and 6000 molar equivalents BDO) (FIG. 1C), d) short diol length (T=180? C., 2.2 molar equivalent DBTL and 6000 molar equivalents BDO) (FIG. 1D).

##STR00001##

Chemical and Physico-Chemical Properties Analysis

.SUP.1.H NMR Analysis

[0126] Characteristic .sup.1H-NMR signals for hydroxybutyrate (HB) units were observed at ?=5.29, 2.44-2.65, 1.29 ppm for CH(CH.sub.3)CH.sub.2CO, CH(CH.sub.3)CH.sub.2CO and CH(CH.sub.3)CH.sub.2CO protons respectively. Peaks that correspond to oligoPHB-diol primary hydroxyl end-groups were ascribed at ?=3.67 for HOCH.sub.2CH.sub.2-protons and secondary hydroxyl end-groups at ?=4.19 ppm for HOCH(CH.sub.3)CH.sub.2 protons. Small peaks at chemical shifts ?=5.81 and 6.96 were assigned to the presence of vinyl end-groups due to PHB thermal degradation and formation of crotonyl end-groups and can be seen zoomed in the box in FIG. 2.

FTIR-ATR Analysis

[0127] FIG. 3 represents the OligoPHB-diols with various M.sub.n: from top to bottom, PHB with decreasing M.sub.n: 260 000 g.Math.mol.sup.?1, 1000 g.Math.mol.sup.?1, 450 g.Math.mol.sup.?1 and 3000 g.Math.mol.sup.?1. Characteristic peaks from oligoPHB-diol structure were ascribed at 3400, 1720, 1450, 1376, 1172 and 1054 cm.sup.?1 corresponding to OH intermolecular stretching vibration, C?O from ester groups stretching vibration, methyl CH bending vibration, OH bending vibration, CO from ester groups stretching and terminal hydroxyl CO stretching vibration superimposed with COC stretching from ester, respectively. Small peak intensities of cis-disubstituted C?C characteristic stretching vibration and its corresponding CH bending vibration were also found at 1655 cm.sup.?1 and 735 cm.sup.?1, respectively.

Molar Masses, IOH and Viscosity Analyses

[0128]

TABLE-US-00001 TABLE 1 Results of molar masses, IOH and viscosity analyses PHB diol M.sub.n Viscosity (25? C.) (by SEC THF) [Pa .Math. s] I.sub.OH [mgKOH/g] ~260000 g .Math. mol.sup.?1 1000 g .Math. mol.sup.?1 110 700 g .Math. mol.sup.?1 .sup.30640 ? 1261 160 450 g .Math. mol.sup.?1 1.62 ? 0 365 300 g .Math. mol.sup.?1 1.60 ? 0 420

[0129] From 1000 g.Math.mol.sup.?1 to high M.sub.n, the obtained oligoPHB-diols are in form of a solid and thus cannot be processed into a prepolymer without solvent in the second step of reaction (see Debuissy, Pollet, and Averous, 2017).

Example 2: Prepolymers Synthesis According to the Invention (oligoPHB-Diol300+DDI/HDI) and Characterization

[0130] ##STR00002##

Synthesis

[0131] A series of prepolymers was prepared in a one-step bulk process without catalyst in a green approach. For that, in a previously flame dried three-necked bottom flask equipped with argon flow, a precise amount of DDI was added (see table 2). The reaction was set to 75? C. with an oil bath under mechanical stirring. Previously dried oligoPHB-diol (PHB300 as prepared in example 1) was then added, followed by HDI, dropwise. In the case of neat HDI-based prepolymer, HDI was first added in the three-necked bottom flask, followed by the oligoPHB-diol. The reaction proceeded at 75? C., and the extend was monitored by free NCO content (% NCO) for 2-3 h to yield prepolymers with an NCO/OH molar ration of 2. Five different formulations were prepared and stored at room temperature under argon. They were named DDI-X % according to the mol % of DDI in the diisocyanate content, hence with X varying from 0 to 100 (Table 2).

TABLE-US-00002 TABLE 2 Prepolymers formulation (mol % are given in brackets) (Wt % in relation to the total weight of the prepolymer) PHB300 content HDI content DDI content Prepolymer [wt %] [wt %] [wt %] DDI-0% 44 56 (100) 0 (0) DDI-25% 30 28 (75) 42 (25) DDI-50% 23 14 (50) 63 (50) DDI-75% 18 6 (25) 76 (75) DDI-100% 15 0 (0) 85 (100)

Chemical and Physico-Chemical Properties Analysis

NCO Content Determination

[0132] The free NCO content was obtained by an indirect-titration method. For that, DBA (50 mL of a 0.2M solution in dry THF) was added to a known mass of prepolymer (1-2 g) and reacted for 2 min. The resulting amine excess was then back titrated using a standard aqueous HCl 0.5M solution and bromophenol blue as indicator. The NCO content, given in weight percent, was calculated as follows:

[00001]$\%\mathrm{NCO}=\frac{\left(\mathrm{Vb}-V_S\right)?4202?0.5}{M?1000}$

[0133] Where Vb (mL) is the HCl solution volume necessary for the blank titration, Vs (mL) the HCl solution volume required for the sample titration, and M (g) the prepolymer weight.

1H NMR

[0134] FIG. 4. represents .sup.1H-NMR spectra of a) neat OligoPHB-diol and b) DDI-0% to f) DDI-100% prepolymers, respectively. Signals at chemical shifts ?=3.67 and 4.19 ppm from oligoPHB-diol protons next to primary and secondary OH respectively disappeared due to the reaction between OH and NCO groups to form urethane bonds. Chemical shift at ?=3.14 ppm was assigned to CH.sub.2NHCOO protons from HDI and/or DDI and ?=3.29 ppm to protons adjacent to the terminal NCO groups. Characteristic signals of PHB repetitive units at ?=5.29 and 2.44-2.65 ppm were attributed to CH(CH.sub.3)CH.sub.2CO and CH(CH.sub.3)CH.sub.2CO protons, respectively, while peaks in the region ?=1.57-1.77 ppm corresponded to the superimposition of CH.sub.2 protons from all components, thus from BDO, oligoPHB-diol, HDI and DDI chains.

FTIR-ATR Analysis

[0135] FIG. 5. shows the results of FTIR-ATR analysis from top to bottom: neat OligoPHB-diol, DDI-0% to DDI-100% prepolymers, respectively.

[0136] FTIR-ATR analysis was used to confirm the successful synthesis of NCO terminated prepolymers from the oligoPHB-diol. FTIR spectra of oligoPHB-diol and the five prepolymers are depicted in FIG. 5. Polyaddition between oligoPHB-diol and HDI/DDI was demonstrated by the disappearance of the broad OH stretching vibration from oligoPHB-diol at 3400 cm.sup.?1, along with NH stretching band appearance at 3329 cm.sup.?1 from urethane bonds. Other characteristic bands from NCO-terminated polyurethane prepolymer architecture were assigned at 2260, 1700 and 1515 cm.sup.?1. They were attributed to N?C?O stretching vibration from prepolymer terminal NCO functions, C?O stretching band from urethane and NH bending vibration from urethane, respectively.

[0137] As DDI content increased in the prepolymer formulation (from DDI-0% to DDI-100%), CH.sub.3 and CH.sub.2 asymmetric stretching vibrations at 2920 and 2851 cm.sup.?1, as well as CH.sub.z bending vibrations at 1460 cm.sup.?1, became stronger due to numerous CH.sub.2 from DDI long aliphatic chains. This observation was in accordance with the M.sub.n increase exhibited in Table 3. Moreover, with increasing DDI content there was a gradual decline of the terminal NCO band at 2260 cm.sup.?1, when compared to the neat HDI-based prepolymer (DDI-0%). This result is in accordance with the decreasing free % NCO shown in Table 3. Globally, all characteristic urethane peak areas intensities diminished with the addition of DDI in the prepolymer. They decreased to the point where C?O stretching bands from oligoPHB-diol and urethane at 1725 cm.sup.?1 and 1700 cm.sup.?1, respectively, were clearly distinguishable. On the contrary, for DDI-25% and DDI-0%, these bands were superimposed as a result of higher urethane bond content, in the chains.

Viscosity Properties

[0138]

TABLE-US-00003 TABLE 3 Prepolymers main physico-chemical properties Viscosity at 25? C. was M.sub.n M.sub.w Viscosity Free % NCO carried out for the [g .Math. mol.sup.?1] [g .Math. mol.sup.?1] ? [Pa .Math. s] [%] DDI-0% 1577 2517 1.60 17 ? 2 14.0 DDI-25% 2543 5924 2.33 47 ? 1 9.5 DDI-50% 3615 10548 2.92 69 ? 6 7.1 DDI-75% 4102 7976 1.94 83 ? 2 5.7 DDI-100% 3959 7179 1.81 101 ? 0 4.8

Example 3: Hemostatic Tissue Adhesive Synthesis According to the Invention (PHB-Diol Oligomers 300+DDI/HDI)

[0139] The tissue adhesives are composed of two-component systems comprising a prepolymer and BDO. This latter acts as a reactive solvent to maintain a low viscosity for the deliverance of the mix, and at the end to increase the final PU molar mass as chain extender, as for a conventional TPU synthesis (=Hemostatic tissue adhesive).

Characterization

Curing Time

[0140] In order to test the reactivity of the NCO-terminated prepolymers, the curing times of the two-component systems were evaluated, since a fast-curing time is essential for surgical adhesives. The corresponding kinetic curves are displayed in FIG. 6a). A significantly higher kinetic, more specifically in the first 40 min, was associated to an increasing HDI content in the prepolymer. This observation is due to multiple phenomena: (i) an increasing oligoPHB-diol content from DDI-100% to DDI-0%, and hence of residual catalyst content. (ii) The initial prepolymers % NCO: It was indeed previously demonstrated that an increased initial % NCO reduces the prepolymers curing time(Gogoi, Alam, and Khandal 2014). Hence, increasing the HDI content results in higher kinetics due to higher initial % NCO (Table 3). Finally, the increase in reactivity with the HDI content can be explained by (iii) the DDI structure influence on the prepolymers: DDI presents a high hydrophobicity due to the long and flexible aliphatic grafted chains (Scheme 2), and its increasing content results in higher prepolymers M.sub.n and viscosities (Table 3). In this case, without an initial high mixing, hydrophilic BDO and high content DDI prepolymers can undergo phase segregation due to the difference of hydrophobic/hydrophilic balance, associated with the gap of viscosity. With different chain extenders, due to the higher reactivity of NH.sub.2 compared to OH functions and water, in the first 6 min of reaction for ABO and 13 min for BDA, the decrease in free NCO was more drastic than for the OH bifunctional chain extenders (FIG. 6 b)) in the enlarged square). However, after these times the tendency reversed and the remaining % of free NCO functions with BDO, PDO and Etg was lower than ABO and BDA for a same reaction time. At 2 h for instance, the free % NCO was around 43% for ABO and BDA, and around 25% for BDO, PDO and Etg.

Adhesion Strength on Muscle Tissue

[0141] In connection with the targeted biomedical application, adhesives properties of the different systems were further evaluated. The adhesion strength was obtained using a lap-shear test in analogy with ASTM F2255-03 standard. Fresh bovine muscle and liver tissue, as well as porcine skin tissues were used as substrates to test all formulations. Results are depicted in FIG. 7 and display the adhesion strength values (R.sub.max in Pa) for all formulations compared to a well-known commercial reference, IFABOND. Compared to some results obtained in previous studies (McDermott et al. 2004) and to IFABOND, values for the different prepolymers with BDO are rather low, ranging from 2 to 11 kPa. In our two-component systems, increasing HDI content resulted in an improved adhesion strength, most probably due to a higher urethane bonds density and thus physical interactions with the tissues after curing. Moreover, lower initial % NCO in DDI-based systems (Table 3) drops the global adhesion strength capacity of the adhesive. Varying the chain extender using high reactivity functional groups such as NH.sub.2 significantly improved the adhesion on muscle tissues: the average adhesion strength gradually increased from 4590 to 17770 Pa for BDO and BDA, respectively. Reducing the chain extender chain length also increased adhesion but to a much lower extend (up to 5600 Pa for Etg).

TPUs Model Polymers Analysis

Synthesis

[0142] A series of five TPUs (hemostatic tissue adhesive) was prepared from PHB-diol oligomers with HDI and/or DDI as diisocyanates and BDO as chain extender. The general procedure for the TPU preparation was based on a classic two-step method. First, NCO-terminated prepolymers were prepared by adding oligoPHB-diol to different proportions of HDI and DDI with a NCO:OH molar ratio of 2, as described in Example 2. After determination of the NCO content, an exact amount of chain extender was added (See Table 4). The reaction was then vigorously stirred for one minute and subsequently poured in a Teflon mold. Preparations were kept overnight in an oven at 80? C. to ensure completion of the reaction. Finally, TPUs were compression-molded in a hot press at 120? C. with 200 MPa pressure for 5 min, followed by 10 min quenching between two steel-plates to obtain 1 mm thickness films.

TABLE-US-00004 TABLE 4 TPUs-based Hemostatic adhesives compositions Prepolymer OligoPHB- DDI Chain extender diol content HDI content content Chain content TPU [wt %] [wt %] [wt %] extender [wt %] DDI-0%-BDO 39 (1) 49 (2) 0 (0) BDO 12 (1) DDI-25%-BDO 27 (1) .sup.26 (1.5) .sup.38 (0.5) BDO 9 (1) DDI-50%-BDO 21 (1) 13 (1) 59 (1) BDO 7 (1) DDI-75%-BDO 17 (1) 5 (0.5) .sup.71 (1.5) BDO 7 (1) DDI-100%-BDO 14 (1) 0 (0) 81 (2) BDO 5 (1) DDI-50%-BDA 21 (1) 13 (1) 59 (1) BDA 7 (1) DDI-50%-ABO 21 (1) 13 (1) 59 (1) ABO 7 (1) DDI-50%-PDO 21 (1) 14 (1) 61 (1) PDO 4 (1) DDI-50%-Etg 21 (1) 14 (1) 61 (1) Etg 4 (1)

Mechanical Properties

[0143] Uniaxial Stress-strain curves at room temperature are shown in FIG. 8 in order to evaluate the mechanical behavior of the TPU and to mimic a potential stress solicitation of the final materials in situ. The corresponding values are shown in Table 5. Increasing the HDI content resulted to a significant increase of the elastic modulus and tensile strength, up to a Young's modulus of 174 MPa for DDI-0%. DDI-0%-BDO exhibited a typical thermoplastic polymer behavior with plastic deformation region after the yield point, followed by an increasing stress until rupture. The other materials containing increasing DDI contents had higher elongations and lower tensile strengths at break, which are conventional behaviors of thermoplastic elastomers (TPE), as it is usually the case for most TPUs. Elongation at break increased with the DDI fraction, which bring mobility and softness with decreasing Young's modulus, as expected for this type of material. TPU-urea PHB300-ABO and PHB300-BDA are hard and brittle polymers with high Young's Modulus and short elongations at break, compared to PHB300-BDO. It is common for PU-urea to achieve high tensile strength due to greater physical crosslink from NH bonds in urea, which correlates to high Tg(HS) described by DSC earlier. With regard to the chain extender length, harder polymers were obtained by increasing the number of carbons in the chain. In fact, average Young's moduli from 0.6 to 24.4 MPa were obtained for PHB300-Etg and PHB300-BDO, respectively. Usually, increasing the chain length leads to more flexible polymers, but this tendency is reversed for very short chains. Here, it was clear that the incorporation of shorter diols (2 and 3 carbons) resulted in more flexible chains with a gradual increase of the elongation at break (up to 517% with Etg).

TABLE-US-00005 TABLE 5 Mechanical properties of the model TPUs Elastic modulus Tensile strength at Elongation at TPU [MPa] break [MPa] break [%] DDI-0%-BDO 174.0 ? 1.3 11.2 ? 0.5 194 ? 14 DDI-25%-BDO 39.0 ? 1.0 4.4 ? 0.1 97 ? 6 DDI-50%-BDO 24.4 ? 0.3 2.1 ? 0.0 96 ? 1 DDI-75%-BDO 5.5 ? 0.1 0.9 ? 0.0 147 ? 5 DDI-100%-BDO 1.6 ? 0.1 0.5 ? 0.0 185 ? 35 DDI-50%-BDA 27.3 ? 1.5 2.8 ? 0.1 13 ? 1 DDI-50%-ABO 17.7 ? 0.4 3.6 ? 0.1 72 ? 11 DDI-50%-PDO 1.1 ? 0.0 0.7 ? 0.1 336 ? 20 DDI-50%-Etg 0.6 ? 0.2 0.4 ? 0.0 517 ? 12