Branched hydroxyproline polymer, physical hydrogel, and methods for preparing and modifying branched hydroxyproline polymer

Abstract

Disclosed is a branched hydroxyproline polymer having a weight average molecular weight in the range of 1 to 10,000 kDa and a branching degree in the range of 0.01 to 0.5. Further disclosed are an at least partially modified branched hydroxyproline polymer, a method for preparing the branched hydroxyproline polymer, a method for modifying the branched hydroxyproline polymer, a physical hydrogel, and a method for preparing a branched amino acid polymer.

Description

Branched hydroxyproline polymer, physical hydrogel and method for preparing and modifying branched hydroxyproline polymer

[0001] This application claims priority to Chinese Patent Application No. 202311436210.0 filed on October 31, 2023, and the contents of the above-mentioned Chinese patent application disclosure are hereby incorporated by reference in their entirety as a part of this application. Technical Field

[0002] Embodiments of the present disclosure relate to the field of polymers, and in particular to a branched hydroxyproline polymer, an at least partially modified branched hydroxyproline polymer, a method for preparing a branched hydroxyproline polymer, a method for modifying a branched hydroxyproline polymer, a physical hydrogel, and a method for preparing a branched polyamino acid. Background Art

[0003] [Corrected 11.11.2024 according to Rule 26] Polyamino acids (also known as polypolypeptide polymers, synthetic polypolypeptides, etc.) are important bio-based polymers that can be used as drugs, pharmaceutical preparations or carriers. They have important uses in many high-value-added fields such as high-end biomaterials, cosmetics, asymmetric catalysis, and have huge market potential. For example, polylysine can be used in cell culture, antibacterial materials, gene transfection materials and other fields; polyglutamic acid and aspartic acid can be used as pharmaceutical preparations and carriers as well as medical materials. Several nanomedicines based on polyglutamic acid and polyaspartic acid have entered the clinical trial stage in the United States, Japan and China; polyvaline and polyleucine are often used as catalysts for the industrial asymmetric Julia-Colonna epoxidation reaction; polysarcosine is a good pharmaceutical preparation and carrier; glatiramer acetate (i.e., L-alanine-L-glutamic acid-L-lysine-L-tyrosine polypeptide polymer acetate (prepared by polymerization of four amino acid N-carboxyl cyclic anhydrides (hereinafter referred to as NCA)) was approved by the US FDA in 1996 for the treatment of multiple sclerosis. Its efficacy and tolerability have won it a good reputation in Western countries with a large number of multiple sclerosis patients. Its sales in 2012 reached US$4 billion, and it has been among the top 20 best-selling drugs in the world for many years. At present, the synthesis of various polyamino acids has gradually become a research direction that the industry focuses on. For example, polyamino acids can be prepared by ring-opening polymerization of amino acids NCA, but amino acids with active functional groups such as hydroxyl side chains need to have their active functional groups protected before the corresponding amino acid NCA can be synthesized, and then deprotected after the ring-opening polymerization is completed.

[0004] Polyhydroxyproline (HP) is an important model molecule for applications in a wide range of fields, including targeted drug delivery and biomimetic catalysis. The development of functionalization methods and materials for its functionalization is of significant research interest. HP has a unique PPII helical secondary structure. The trans-hydroxyl groups on its side chains give it a more stable PPII helical structure than polyproline, resulting in improved hydrophilicity. Because HP is an endogenous amino acid with modifiable hydroxyl groups, several publications have reported the design of HP derivatives based on HP via solid-phase peptide synthesis. However, the synthesis of HP is extremely difficult: solid-phase peptide synthesis methods can only yield very short oligomers (with a degree of polymerization <50). Traditional NCA ring-opening polymerization requires protecting the hydroxyl groups of HP to produce acetyl-HP (NCA), ring-opening polymerization of acetyl-HP, and subsequent hydrolytic removal of the acetyl protecting group. However, the polymerization rate of acetyl-HP is reportedly slow (polymerization in pyridine requires two days), and the subsequent hydrolytic removal of the acetyl protecting group can easily lead to racemization. The traditional NCA ring-opening polymerization method for synthesizing polyhydroxyproline via a protection-deprotection process is time-consuming and costly. The resulting polyhydroxyproline has a small and uncontrollable molecular weight, which is inconsistent with the concept of green chemistry. Furthermore, the difficulty in synthesizing polyhydroxyproline has led to a lack of research related to its post-modification. Currently, there are no post-modification strategies or applications for polyhydroxyproline.

[0005] Summary of the Invention

[0006] To address the above and other problems, the present disclosure discloses a branched polyamino acid and a novel method for preparing such a branched polyamino acid by ring-opening polymerization of amino acids (NCA) with active functional groups such as hydroxyl side chains. This method can eliminate the protection and deprotection process of the active functional groups and synthesize ultra-high molecular weight polyamino acids in a one-pot process.

[0007] At least one embodiment of the present disclosure discloses a branched hydroxyproline polymer having a weight average molecular weight ranging from 1 kDa to 10,000 kDa and a degree of branching ranging from 0.01 to 0.5.

[0008] In some examples, the branched hydroxyproline polymer has a weight average molecular weight ranging from 1 kDa to 1000 kDa, and a degree of branching ranging from 0.01 to 0.36.

[0009] In some examples, the branched hydroxyproline polymer has a structure shown in formula (I):

[0010] R1-(L1) x -(L2) y -(L3) z -R2

[0011] Formula (I)

[0012] in,

[0013] L1 is

[0014] L2 is

[0015] L3 is

[0016] R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2;

[0017] R1 is a terminal group;

[0018] R2 is independently H or a terminal group at each occurrence;

[0019] Each occurrence of x and y is independently a fraction greater than 0 and less than 1, and each occurrence of z is independently a fraction greater than or equal to 0 and less than 1, provided that the sum of all x, y, and z is 1.

[0020] In some examples, z is 0, and the branched polyhydroxyproline polymer has a structure shown in the following formula (II):

[0021] R1-(L1) x -(L2) y -R2

[0022] Formula (II)

[0023] in,

[0024] L1 is

[0025] L2 is

[0026] R a Each occurrence is independently selected from -(L1) x -(L2) y -R2;

[0027] R1 is a terminal group;

[0028] R2 is independently H or a terminal group at each occurrence;

[0029] Each occurrence of x and y is independently a fraction greater than 0 and less than 1, provided that the sum of all x and y is 1.

[0030] In some examples, z is not 0, and the branched hydroxyproline polymer has a structure shown in formula (I):

[0031] R1-(L1) x -(L2) y -(L3) z -R2

[0032] Formula (I)

[0033] in,

[0034] L1 is

[0035] L2 is

[0036] L3 is

[0037] R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2;

[0038] R1 is a terminal group;

[0039] R2 is independently H or a terminal group at each occurrence;

[0040] Each occurrence of x, y, and z is independently a fraction greater than 0 and less than 1, provided that the sum of all x, y, and z is 1.

[0041] At least one embodiment of the present disclosure further provides an at least partially modified branched hydroxyproline polymer having a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a degree of branching in the range of 0.01 to 0.5. The at least partially modified branched hydroxyproline polymer has a structure represented by formula (III):

[0042] R1'-(L1') x’ -(L2') y’ -(L3') z’ -R2'

[0043] Formula (III)

[0044] in,

[0045] L1' is

[0046] L2' is

[0047] L3' is

[0048] R a 'Each occurrence is independently selected from -(L1') x’ -(L2') y’-(L3') z’ -R2';

[0049] R1' is a terminal group;

[0050] R2' is independently H or a terminal group at each occurrence;

[0051] Each occurrence of R3 is independently selected from H, C 1-20 Alkyl, C 1-20 Alkenyl, C 1-20 Alkynyl, C 1-20 Alkoxy, C 1-20 Alkenyloxy, C 1-20 Alkynyloxy, C 1-20 Alkoxyalkyl, C 1-20 substituted with a 5-6 membered heteroaryl substituent, wherein the heterocyclic ring and heteroaryl optionally contain one to three heteroatoms independently selected from S, O or N,

[0052] Each occurrence of x' and y' is independently a fraction greater than 0 and less than 1, and each occurrence of z' is independently a fraction greater than or equal to 0 and less than 1, provided that the sum of all x', y' and z' is 1.

[0053] In some examples, the at least partially modified branched hydroxyproline polymer is

[0054] R2' and R3 are each independently selected from H, C 1-20 Alkyl, C 1-20 Alkyl acyl, C 1-20 Alkoxyacyl, C 1-20 Alkylsulfonyl and di(C 1-20 The alkyl group in the above group is optionally unsubstituted or substituted by 1 to 3 substituents selected from saturated or unsaturated 5-6 membered heterocyclic groups.

[0055] In some examples, in the at least partially modified branched hydroxyproline polymer, R2' and R3, at each occurrence, are independently selected from H, acetyl, and lipoyl.

[0056] In some examples, the modification degree of the at least partially modified branched hydroxyproline polymer is between 0.1% and 100%.

[0057] At least one embodiment of the present disclosure further provides a method for preparing a branched hydroxyproline polymer, comprising: treating a compound represented by the following formula:

[0058] The branched hydroxyproline polymer is polymerized in an organic solvent in the presence of a base to obtain the branched hydroxyproline polymer, wherein the branched hydroxyproline polymer has a weight average molecular weight ranging from 1 kDa to 10,000 kDa and a branching degree ranging from 0.01 to 0.5.

[0059] In some examples, the base is selected from potassium hydride, sodium hydride, potassium methoxide, sodium methoxide, potassium tert-butoxide, sodium tert-butoxide, triazabicyclo[4.4.0]dec-5-ene (TBD), 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), N,N-diisopropylethylamine (DIPEA) or any mixture thereof.

[0060] In some examples, the organic solvent is selected from dimethyl sulfoxide (DMSO), dimethylformamide (DMF), acetonitrile, dichloromethane (DCM), ethyl acetate, tetrahydrofuran (THF), acetone, and mixtures thereof.

[0061] At least one embodiment of the present disclosure further provides a method for at least partially modifying a branched hydroxyproline polymer, wherein the branched hydroxyproline polymer has a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a degree of branching in the range of 0.01 to 0.5, the method comprising: reacting the branched hydroxyproline polymer with a modifying agent in the presence of a base in an organic solvent to at least partially modify the branched hydroxyproline polymer, wherein the branched hydroxyproline polymer has a structure represented by the following formula (I):

[0062] R1-(L1) x -(L2) y -(L3) z -R2

[0063] Formula (I)

[0064] in,

[0065] L1 is

[0066] L2 is

[0067] L3 is

[0068] R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2;

[0069] R1 is a terminal group;

[0070] R2 is independently H or a terminal group at each occurrence;

[0071] Each occurrence of x and y is independently a fraction greater than 0 and less than 1, and each occurrence of z is independently a fraction greater than or equal to 0 and less than 1, provided that the sum of all x, y, and z is 1,

[0072] And wherein, the modifying agent is selected from the group consisting of:

[0073] Wherein, R is selected from C 1-20 alkyl.

[0074] In some instances, R is methyl.

[0075] In some examples, the base is selected from trimethylamine, triethylamine, triethanolamine, and mixtures thereof.

[0076] In some examples, the organic solvent is selected from DMSO, DMF, acetonitrile, DCM, ethyl acetate, THF, acetone, and mixtures thereof.

[0077] At least one embodiment of the present disclosure further provides a physical hydrogel comprising any one of the at least partially modified branched hydroxyproline polymers described above.

[0078] In some examples, the at least partially modified branched hydroxyproline polymer is an at least partially acylated branched hydroxyproline polymer.

[0079] In some examples, the at least partially modified branched hydroxyproline polymer is an at least partially acetylated branched hydroxyproline polymer or an at least partially lipoylated branched hydroxyproline polymer.

[0080] At least one embodiment of the present disclosure further provides a method for preparing a branched polyamino acid, comprising: reacting a compound of formula (IV):

[0081] in:

[0082] R A and R B One of them is C 1-6 Alkyl, another C substituted with hydroxy 1-6 alkyl,

[0083] Or, R A and R B Together with the ring atoms to which they are attached, they form a five- or six-membered ring substituted with hydroxy,

[0084] The branched polyamino acid is polymerized in an organic solvent in the presence of a base to obtain the branched polyamino acid, which has a weight average molecular weight ranging from 1 kDa to 10,000 kDa and a branching degree ranging from 0.01 to 0.5.

[0085] In some examples, the compound of formula (IV) has a structure selected from the group consisting of:

[0086] Among them, R 1 -(CH2) 1-2 -,

[0087] R 2 and R 3 Each occurrence is optionally independently selected from H, CH3- and CH3CH2-,

[0088] The condition is R 1 、R 2 and R 3 The sum of the number of carbon atoms is not greater than 5;

[0089] R4 is C 1-6 alkyl.

[0090] In some examples, the base is selected from potassium hydride, sodium hydride, potassium methoxide, sodium methoxide, potassium tert-butoxide, sodium tert-butoxide, TBD, DBU, DIPEA, or any mixture thereof.

[0091] In some examples, the organic solvent is selected from DMSO, DMF, acetonitrile, DCM, ethyl acetate, THF, acetone, and mixtures thereof. BRIEF DESCRIPTION OF THE DRAWINGS

[0092] In order to more clearly illustrate the technical solutions of the embodiments of the present disclosure, the drawings of the embodiments will be briefly introduced below. Obviously, the drawings described below only relate to some embodiments of the present disclosure, rather than limiting the present disclosure.

[0093] In order to more clearly illustrate the technical solutions of the embodiments of the present disclosure, the drawings of the embodiments will be briefly introduced below. Obviously, the drawings in the following description only relate to some embodiments of the present disclosure, rather than limiting the present disclosure.

[0094] FIG1 shows a comparison between a conventional polyhydroxyproline synthesis method and the method disclosed herein;

[0095] FIG2 shows the SEC curves of products prepared using 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) and different [NCA] / [base] ratios;

[0096] FIG3 shows size exclusion chromatography (SEC) curves of products prepared using N,N-diisopropylethylamine (DIPEA) and different [NCA] / [base] ratios;

[0097] FIG4 shows SEC curves of products prepared using DIPEA and different [NCA] / [base] ratios;

[0098] FIG5 shows a schematic diagram of the branched structure of a branched hydroxyproline polymer (B-PHyp);

[0099] FIG6 shows the infrared absorption spectra of branched hydroxyproline polymer (B-PHyp) and polyacetylhydroxyproline (PAcHyp);

[0100] Figure 7 shows the relationship between B-PHyp and linear polyhydroxyproline 1 H-NMR (D2O);

[0101] Figure 8 shows the SEC curve changes of B-PHyp (black) and the aqueous phase after adding NaOH (red);

[0102] FIG9 shows the reaction formula for quantitative analysis of secondary amine end groups using diazonium salts and a standard curve prepared using a proline standard solution;

[0103] Figure 10 shows the relationship between B-PHyp and acetyl-branched hydroxyproline polymer (Ac-B-PHyp). 1 H-NMR (D2O) comparison;

[0104] FIG11 shows rheological data of 2 wt % Ac-B-PHyp hydrogel: A) oscillation frequency sweep curve; B) amplitude sweep curve; C) amplitude step sweep curve; D) injectable schematic diagram;

[0105] Figure 12 shows the branched hydroxyproline-proline copolymer 1 H-NMR (D2O);

[0106] FIG13 shows the size exclusion chromatography (SEC) curve of the branched hydroxyproline-proline copolymer. DETAILED DESCRIPTION

[0107] For the purpose of the following detailed description, it should be understood that, unless otherwise specified to the contrary, the present disclosure may adopt various alternative variations and step sequences. In addition, except in any operating examples, or when otherwise indicated, all numerals representing the amount of the components used in the specification and claims should be understood to be modified by the term "about" in all cases. Therefore, unless otherwise specified to the contrary, the numerical parameters set forth in the following specification and the appended claims are approximate values ​​that vary according to the desired performance to be obtained by the present disclosure. At least, it is not intended to limit the application of the doctrine of equivalents to the scope of the claims, and each numerical parameter should at least be interpreted according to the number of reported significant figures and by applying ordinary rounding techniques.

[0108] Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the disclosure are approximations, the numerical values ​​set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0109] Furthermore, it should be understood that any numerical range recited herein is intended to include all subranges subsumed therein. For example, a range of "1 to 10" is intended to include all subranges between (and including) the recited minimum value of 1 and the recited maximum value of 10, i.e., having a minimum value equal to or greater than 1 and a maximum value equal to or less than 10.

[0110] In this disclosure, unless otherwise expressly stated, the use of the singular includes the plural and the plural includes the singular. Furthermore, in this disclosure, unless otherwise expressly stated, the use of "or" means "and / or," even though "and / or" could be explicitly used in certain circumstances. Furthermore, in this disclosure, unless otherwise expressly stated, the use of "a" or "an" means "at least one." For example, "a" polymer, "a" composition, etc., refers to one or more of any of these items.

[0111] Branched hydroxyproline polymers

[0112] The present disclosure provides a branched hydroxyproline polymer. As described herein, the term "hydroxyproline" refers to 4-hydroxypyrrolidine-2-carboxylic acid; the term "hydroxyproline polymer" refers to a polymeric substance formed by polymerization or copolymerization of hydroxyproline and, optionally, one or more other monomers, including but not limited to hydroxyproline homopolymers or copolymers of hydroxyproline and other monomers. For example, the hydroxyproline polymer described in the present disclosure may be a hydroxyproline homopolymer formed by polymerization of only hydroxyproline as a monomer; or, it may be a copolymer formed by hydroxyproline and a monomer such as, but not limited to, proline (2-pyrrolidinecarboxylic acid) (e.g., a hydroxyproline-proline copolymer). As used herein, the term "branched hydroxyproline polymer" refers to a hydroxyproline polymer having branches of varying lengths on a linear main chain; in some cases, one, some, or all of these branches may further have more branches of varying lengths, and these further branches may further have more branches of varying lengths, and so on, thereby forming a branch-shaped macromolecule, i.e., a dendritic polymer. The branched hydroxyproline polymer disclosed in the present invention is a type of highly branched three-dimensional macromolecule with numerous branching points. The molecular chains are not easily entangled, the viscosity does not change with the increase of molecular weight, and it has abundant terminal functional groups, which makes it easy to modify and adapt it, thereby facilitating the synthesis of various functional materials.

[0113] In some aspects, the weight average molecular weight of the branched hydroxyproline polymers described herein can be in the range of 1 kDa to 10,000 kDa, and the degree of branching can be in the range of 0.01 to 0.5. As used herein, the term "weight average molecular weight" is a statistical average molecular weight based on the weight average of molecules of different molecular weights in a polymer and is one of the most basic parameters of a polymer material. As used herein, the term "degree of branching" refers to the ratio of the sum of the number of branching units and the number of terminal units in a branched polymer to the total number of monomer units in the polymer. For example, the weight average molecular weight of the branched hydroxyproline polymers described in the present disclosure can be in the range of 1 kDa to 9,000 kDa, for example, 1 kDa to 8,500 kDa, 1 kDa to 8,000 kDa, 1 kDa to 7,500 kDa, 1 kDa to 7,000 kDa, 1 kDa to 6,500 kDa, 1 kDa to 6,000 kDa, 1 kDa to 5,500 kDa, 1 kDa to 5,000 kDa, 1 kDa to 4,500 kDa, 1 kDa to 4,500 kDa, kDa, 0.02 to 0.48, 0.02 to 0.45, 0.02 to 0.42, 0.02 to 0.40, 0.02 to 0.38, 0.02 to 0.36, or a range between any two of the foregoing endpoints, etc. The degree of branching can be in the range of 0.02 to 0.5, for example, 0.02 to 0.48, 0.02 to 0.45, 0.02 to 0.42, 0.02 to 0.40, 0.02 to 0.38, 0.02 to 0.36, or a range between any two of the foregoing endpoints, etc. The branched hydroxyproline polymer within the above range can be effectively used in macromolecular initiators, post-modification platforms, drug carriers, etc.

[0114] In some aspects, the branched hydroxyproline polymers described herein may have a structure represented by the following formula (I):

[0115] R1-(L1) x -(L2) y -(L3) z -R2

[0116] Formula (I)

[0117] in,

[0118] L1 is

[0119] L2 is

[0120] L3 is

[0121] R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2;

[0122] R1 is a terminal group;

[0123] R2 is independently H or a terminal group at each occurrence;

[0124] Each occurrence of x and y is independently a fraction greater than 0 and less than 1, and each occurrence of z is independently a fraction greater than or equal to 0 and less than 1, provided that the sum of all x, y, and z is 1.

[0125] As shown in the above formula (I), the main chain structure of the branched hydroxyproline polymer described in the present disclosure may include a non-branched hydroxyproline repeating unit L1: and branched hydroxyproline repeating unit L2: And may optionally contain no or a proline repeating unit L3: The repeating units L1, L2 and L3 can be randomly arranged in any order, wherein x, y and z represent the percentage of the corresponding repeating unit in all the repeating units of the branched hydroxyproline polymer. Furthermore, the side chain of the branched repeating unit L2 can also have further branching. That is, -OR in the branched repeating unit L2 a The substituent R a Each occurrence can be independently selected from -(L1) x -(L2) y -(L3) z -R2, thereby forming an "iterative" branched structure. In this way, the linear main chain of the branched hydroxyproline polymer described in the present disclosure carries branches of varying lengths, and one, some, or all of these branches further carry more branches of varying lengths, and these further branches can further carry more branches of varying lengths, and so on, thereby forming a branch-shaped macromolecule, that is, a highly branched dendrimer.

[0126] In some embodiments, the branched hydroxyproline polymer described in the present disclosure may be a branched polyhydroxyproline homopolymer, that is, z may be 0. For example, the branched polyhydroxyproline polymer may have a structure shown in the following formula (II):

[0127] R1-(L1) x -(L2) y -R2

[0128] Formula (II)

[0129] in,

[0130] L1 is

[0131] L2 is

[0132] R a Each occurrence is independently selected from -(L1) x -(L2) y -R2;

[0133] R1 is a terminal group;

[0134] R2 is independently H or a terminal group at each occurrence;

[0135] Each occurrence of x and y is independently a fraction greater than 0 and less than 1, provided that the sum of all x and y is 1.

[0136] As shown in the above formula (II), the main chain structure of the branched hydroxyproline polymer described in the present disclosure may include a non-branched hydroxyproline repeating unit L1: and branched hydroxyproline repeating unit L2: But does not contain the proline repeat unit L3: The repeating units L1 and L2 can be randomly arranged in any order, wherein x and y represent the percentage of the corresponding repeating unit in all the repeating units of the branched hydroxyproline polymer. Furthermore, the side chain of the branched repeating unit L2 can also have further branching. That is, -OR in the branched repeating unit L2 a The substituent R a Each occurrence can be independently selected from -(L1) x -(L2) y -(L3) z -R2, thereby forming an "iterative" branched structure. In this way, the linear main chain of the branched hydroxyproline polymer described in the present disclosure carries branches of varying lengths, and one, some, or all of these branches further carry more branches of varying lengths, and these further branches can further carry more branches of varying lengths, and so on, thereby forming a branch-shaped macromolecule, that is, a highly branched dendrimer.

[0137] In some examples, the branched hydroxyproline polymer described in the present disclosure may have a structure such as shown in the following formula:

[0138] Wherein, R is an end group; x and n represent the percentage of branched repeating units and unbranched repeating units, respectively, of the total repeating units of the branched hydroxyproline polymer described herein, 0≤x<1, 0≤n<1, provided that x+n=1. In some examples, x can be, for example, 1% to 50%, preferably 1% to 28%; n can be, for example, 50% to 99%, preferably 72% to 99%.

[0139] In other embodiments, the branched hydroxyproline polymer described in the present disclosure may be a branched hydroxyproline-proline copolymer, that is, z is not 0. For example, the branched hydroxyproline-proline copolymer may have a structure shown in the following formula (I):

[0140] R1-(L1) x -(L2) y -(L3) z -R2

[0141] Formula (I)

[0142] in,

[0143] L1 is

[0144] L2 is

[0145] L3 is

[0146] R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2;

[0147] R1 is a terminal group;

[0148] R2 is independently H or a terminal group at each occurrence;

[0149] Each occurrence of x, y, and z is independently a fraction greater than 0 and less than 1, provided that the sum of all x, y, and z is 1.

[0150] As used herein, the term "end group" refers to a non-hydrogen group at the end of a polymer molecule chain, i.e., a terminal group / capping group at the polymer chain. The end group described in the present disclosure can be any terminal group commonly used in the art, including but not limited to hydroxyl, carboxyl, amino, amide, ester, etc., and the present disclosure does not impose specific limitations on this.

[0151] At least partially modified branched hydroxyproline polymers

[0152] The present disclosure also provides an at least partially modified branched hydroxyproline polymer having a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a degree of branching in the range of 0.01 to 0.5. As used herein, the term "modification" refers to the phenomenon that the chemical structure of a substance is changed by the introduction or removal of chemical groups. Common modifications include, but are not limited to, acylation and deacylation, sulfonylation and desulfonylation, phosphorylation and dephosphorylation, alkylation and dealkylation, etc. The at least partially modified branched hydroxyproline polymer described in the present disclosure may include, but is not limited to, modification at at least a portion of the free hydroxyl sites of the branched hydroxyproline polymer, for example, acylation, sulfonylation, alkylation, etc.

[0153] In some examples, the at least partially modified branched hydroxyproline polymer described in the present disclosure can have a structure represented by Formula (III):

[0154] R1'-(L1') x’ -(L2') y’ -(L3') z’ -R2'

[0155] Formula (III)

[0156] in,

[0157] L1' is

[0158] L2' is

[0159] L3' is

[0160] R a 'Each occurrence is independently selected from -(L1') x’ -(L2') y’ -(L3') z’ -R2';

[0161] R1' is a terminal group;

[0162] R2' is independently H or a terminal group at each occurrence;

[0163] Each occurrence of R3 is independently selected from H, C 1-20 Alkyl, C 1-20 Alkenyl, C 1-20 Alkynyl, C 1-20 Alkoxy, C 1-20 Alkenyloxy, C 1-20 Alkynyloxy, C 1-20 Alkoxyalkyl, C 1-20substituted with a 5-6 membered heteroaryl substituent, wherein the heterocyclic ring and heteroaryl optionally contain one to three heteroatoms independently selected from S, O or N,

[0164] Each occurrence of x' and y' is independently a fraction greater than 0 and less than 1, and each occurrence of z' is independently a fraction greater than or equal to 0 and less than 1, provided that the sum of all x', y' and z' is 1.

[0165] As shown in the above formula (III), the main chain structure of the at least partially modified branched hydroxyproline polymer described in the present disclosure may include a non-branched hydroxyproline repeating unit L1': and a branched hydroxyproline repeat unit L2': And it may optionally contain no or contain proline repeating units, wherein the repeating units L1', L2' and L3' may be randomly arranged in any order, wherein x', y' and z' respectively represent the percentage of the corresponding repeating unit in all repeating units of the at least partially modified branched hydroxyproline polymer. Furthermore, the side chain of the branched repeating unit L2' may also have further branching. That is, -OR in the above branched repeating unit L2' a Substituent R on ' a 'Each occurrence can be independently selected from -(L1') x’ -(L2') y’ -(L3') z’ R2', thereby forming an "iterative" branched structure. In this way, the linear backbone of the at least partially modified branched hydroxyproline polymer described in the present disclosure carries branches of varying lengths, and one, some, or all of these branches further carry more branches of varying lengths, and these further branches can further carry more branches of varying lengths, and so on, thereby forming a branch-shaped macromolecule, that is, a highly branched dendrimer.

[0166] In some examples, the weight average molecular weight of the at least partially modified branched hydroxyproline polymers described in the present disclosure can be in the range of 1 kDa to 9,000 kDa, for example, 1 kDa to 8,500 kDa, 1 kDa to 8,000 kDa, 1 kDa to 7,500 kDa, 1 kDa to 7,000 kDa, 1 kDa to 6,500 kDa, 1 kDa to 6,000 kDa, 1 kDa to 5,500 kDa, 1 kDa to 5,000 kDa, 1 kDa to 4,500 kDa, 1 kDa to 5 ...7,000 kDa, 1 kDa to 7,500 kDa, 1 kDa to 7,000 kDa, 1 kDa to 7,500 kDa, 1 kDa to 7,000 kDa, 1 kDa to 7,500 kDa, 1 kDa to 7,000 kDa, kDa to 4,000 kDa, 1 kDa to 3,500 kDa, 1 kDa to 3,000 kDa, 1 kDa to 2,500 kDa, 1 kDa to 2,000 kDa, 1 kDa to 1,800 kDa, 1 kDa to 1,500 kDa, 1 kDa to 1,400 kDa, 1 kDa to 1,300 kDa, 1 kDa to 1,250 kDa, 1 kDa to 1,200 kDa, 10 kDa to 1,200 kDa, or a range between any two of the foregoing endpoints, etc. The degree of branching of the at least partially modified branched hydroxyproline polymers described in the present disclosure can be in the range of 0.02 to 0.5, for example, 0.02 to 0.48, 0.02 to 0.45, 0.02 to 0.42, 0.02 to 0.40, 0.02 to 0.38, 0.02 to 0.36, or a range between any two of the above endpoints, etc.

[0167] In other examples, the branched hydroxyproline polymers described in the present disclosure can be modified at 0.1% to 100% of the free amino and hydroxyl groups of the branched hydroxyproline polymer (i.e., the degree of modification is 0.1% to 100%), for example, 10-60% post-acylation modification, for example, 10%, 15%, 20%, 25%, 30% to 35%, 40%, 45%, 50%, 55% or 60% post-acetylation or lipoylation modification, for example, 40% post-acetylation or lipoylation modification.

[0168] In some examples, at least some of the R2' and at least some of the R3 in the at least partially modified branched hydroxyproline polymer of formula (III) above may be the same and selected from C 1-20 Alkyl, C 1-20 Alkyl acyl, C 1-20 Alkoxyacyl, C 1-20 Alkylsulfonyl or di(C 1-20alkyl)aminooxy, the alkyl group in the above group is optionally unsubstituted or substituted with 1-3 substituents selected from saturated or unsaturated 5-6 membered heterocyclic groups; and the remaining R2' and R3 may be H. In a further example, in the at least partially modified branched hydroxyproline polymer represented by formula (II), at least some R2' and at least some R3 may be acetyl groups or thioctanol groups, and the remaining R2' and R3 may be H, for example, at least 10%, 15%, 20%, 25%, 30% to 35%, 40%, 45%, 50%, 55% or 60% of all R2' and R3 may be acetyl groups or thioctanol groups, and the remaining R2' and R3 may be H.

[0169] Method for preparing branched hydroxyproline polymers

[0170] The present disclosure also discloses a method for preparing a branched hydroxyproline polymer, which comprises: reacting a compound represented by the following formula, i.e., hydroxyproline N-carboxyl intracyclic anhydride (hypNCA) or proline N-carboxyl intracyclic anhydride (pNCA):

[0171] The polymer is polymerized in an organic solvent in the presence of a base to obtain a branched hydroxyproline polymer having a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a branching degree in the range of 0.01 to 0.5.

[0172] Generally speaking, polyamino acids are primarily prepared through the ring-opening polymerization of amino acids (NCA). Amino acids with active functional groups, such as hydroxyl side chains, require protection of these active functional groups before synthesizing NCA, followed by deprotection after the ring-opening polymerization is complete. For example, the traditional ring-opening polymerization of hydroxyproline (NCA) often requires first protecting the hydroxyl group to produce acetylhydroxyproline (NCA), then subjecting the acetylhydroxyproline (NCA) to polymerization, and finally hydrolysis and deacetylation. This traditional synthetic route often requires several days for the polymerization step alone, and the protection-deprotection process often causes racemization of the product. This synthetic route for preparing polyhydroxyproline is time-consuming and costly, resulting in a low and uncontrollable molecular weight, which is inconsistent with the principles of green chemistry.

[0173] After extensive work and in-depth research, the inventors have developed a method for the rapid polymerization of hypNCA and / or pNCA in an organic phase. By utilizing the nucleophilicity of the hydroxyl group and adding an organic base to convert HypNCA into an initiator-type monomer, they achieve a one-pot method for the preparation of ultra-high molecular weight hydroxyproline polymers with ester bonds as branching points, with molecular weights reaching 1 kDa and even exceeding 10,000 kDa. The reaction scheme described in this disclosure is illustrated below using hypNCA as an example:

[0174] Scheme 1: Organic polymerization of HypNCA to generate ultrahigh molecular weight branched hydroxyproline polymers; in situ acylation and subsequent modification

[0175] In some aspects, the base suitable for use in the method for preparing a branched hydroxyproline polymer described in the present disclosure is preferably an organic base. Generally, the weaker the alkalinity of the organic base used, the lower the equivalent weight, the greater the molecular weight obtained by polymerization. Some examples of bases suitable for use may include, but are not limited to, a base selected from the group consisting of potassium hydride, sodium hydride, potassium methoxide, sodium methoxide, potassium tert-butoxide, sodium tert-butoxide, triazabicyclo[4.4.0]dec-5-ene (TBD), DBU, DIPEA, or any mixture thereof. For example, when DIPEA is used as a base, a molecular weight exceeding 1,000 kDa can be obtained.

[0176] In other aspects, some examples of organic solvents suitable for use in the method for preparing branched hydroxyproline polymers described herein include, but are not limited to, solvents selected from the group consisting of dimethyl sulfoxide (DMSO), dimethylformamide (DMF), acetonitrile, dichloromethane (DCM), ethyl acetate, tetrahydrofuran (THF), acetone, and mixtures thereof. Organic solvents can facilitate rapid and controllable synthesis of branched hydroxyproline polymers having high molecular weight or even ultrahigh molecular weight.

[0177] In other aspects, the preparation can be carried out in an anhydrous environment. Optionally, an amine initiator such as benzylamine can be added to the reaction to promote or accelerate the reaction process.

[0178] Method for modifying branched hydroxyproline polymers

[0179] The present disclosure also discloses a method for at least partially modifying a branched hydroxyproline polymer, wherein the branched hydroxyproline polymer has a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a degree of branching in the range of 0.01 to 0.5, the method comprising: reacting a branched hydroxyproline polymer of formula (I) with a modifying agent in the presence of a base in an organic solvent to at least partially modify the branched hydroxyproline polymer, wherein the branched hydroxyproline polymer of formula (I) may have a structure shown in formula (I):

[0180] R1-(L1) x -(L2) y -(L3) z -R2

[0181] Formula (I)

[0182] in,

[0183] L1 is

[0184] L2 is

[0185] L3 is

[0186] R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2;

[0187] R1 is a terminal group;

[0188] R2 is independently H or a terminal group at each occurrence;

[0189] Each occurrence of x and y is independently a fraction greater than 0 and less than 1, and each occurrence of z is independently a fraction greater than or equal to 0 and less than 1, provided that the sum of all x, y, and z is 1,

[0190] Wherein, the modifying agent can be selected from the following group:

[0191] Wherein, R is selected from C 1-20 alkyl.

[0192] In some aspects, R can be methyl.

[0193] In other aspects, the base used can be selected from trimethylamine, triethylamine, triethanolamine, and mixtures thereof.

[0194] In yet other aspects, the organic solvent used can be selected from DMSO, DMF, acetonitrile, DCM, ethyl acetate, THF, acetone, and mixtures thereof.

[0195] Physical hydrogel

[0196] The present disclosure also discloses a physical hydrogel comprising any one of the at least partially modified branched hydroxyproline polymers described above.

[0197] As used herein, the term "hydrogel" refers to a class of highly hydrophilic, three-dimensional network-structured gels that rapidly swell in water and, in this swollen state, can retain large volumes of water without dissolving. Water absorption is closely related to the degree of crosslinking; the higher the degree of crosslinking, the lower the water absorption, a characteristic very similar to soft tissue. The water content in a hydrogel can range from as low as a few thousandths to as high as 99% by weight. The aggregated state of the gel is neither completely solid nor completely liquid; it can maintain a certain shape and volume under certain conditions, and solutes therein can diffuse or permeate through the hydrogel. Depending on the bonding of the hydrogel network, hydrogels can be divided into physical hydrogels and chemical hydrogels. As used herein, the term "physical hydrogel" is formed by physical forces such as electrostatic interactions, hydrogen bonds, and chain entanglement. This type of gel is non-permanent and can be converted into a solution by heating, so it is also called a pseudogel or thermoreversible gel. In some aspects, the physical hydrogels described herein can be injectable physical hydrogels. For example, the solid content of such a physical hydrogel can reach 1 wt% or 2 wt%, and can be used to replace the hyaluronic acid-based intra-articular injections currently used in literature and clinics.

[0198] In some examples, the at least partially modified branched hydroxyproline polymer can be any of the at least partially modified branched hydroxyproline polymers described herein. For example, the at least partially modified branched hydroxyproline polymer can be an at least partially acylated branched hydroxyproline polymer, for example, an at least partially acetylated branched hydroxyproline polymer or an at least partially lipoylated branched hydroxyproline polymer.

[0199] Method for preparing branched polyamino acid

[0200] The present disclosure also discloses a method for preparing a branched polyamino acid, which comprises: reacting a compound of formula (IV):

[0201] in:

[0202] R A and R B One of them is C 1-6 Alkyl, another C substituted with hydroxy 1-6 alkyl,

[0203] Or, R A and R B Together with the ring atoms to which they are attached, they form a five- or six-membered ring substituted with hydroxy,

[0204] The branched polyamino acid is polymerized in an organic solvent in the presence of a base to obtain the branched polyamino acid, which has a weight average molecular weight ranging from 1 kDa to 10,000 kDa and a branching degree ranging from 0.01 to 0.5.

[0205] In some examples, the compound of formula (IV) can have a structure selected from the group consisting of:

[0206] Among them, R 1 -(CH2) 1-2 -,

[0207] R 2 and R 3 Each occurrence is optionally independently selected from H, CH3- and CH3CH2-,

[0208] The condition is R 1 、R 2 and R 3 The sum of the number of carbon atoms is not greater than 5;

[0209] R4 is C 1-6 alkyl.

[0210] In some examples, the base used can be selected from potassium hydride, sodium hydride, potassium methoxide, sodium methoxide, potassium tert-butoxide, sodium tert-butoxide, TBD, DBU, DIPEA, or any mixture thereof.

[0211] In other examples, the organic solvent used can be selected from DMSO, DMF, acetonitrile, DCM, ethyl acetate, THF, acetone, and mixtures thereof.

[0212] Example

[0213] The embodiments of the present disclosure will be described in detail below with reference to the examples, but it will be understood by those skilled in the art that the following examples are only used to illustrate the present disclosure and should not be considered as limiting the scope of the present disclosure. Where specific conditions are not specified in the examples, they are carried out according to conventional conditions or the conditions recommended by the manufacturer. Where the reagents or instruments used are not specified by the manufacturer, they are all conventional products that can be obtained commercially. Unless otherwise specified, all amounts listed are described in parts by weight based on the gross weight. The present disclosure should not be construed as being limited to the specific examples described.

[0214] Example 1. Preparation of branched hydroxyproline polymers according to the present invention

[0215] In a glove box, L-hydroxyproline-N-carboxylic anhydride (157 mg, 1.00 mmol, 2000 equiv) was added to a 5 mL glass vial and dissolved in 500 μL of anhydrous DMSO. N,N-diethylisopropylamine (DIPEA) (0.5 M x 1 μL, 0.0005 mmol, 1 equiv) was then added and allowed to react for 30 min. The solution was diluted to 2.5 mL with water, passed through a PD10 desalting column, and lyophilized to obtain the final branched hydroxyproline polymer.

[0216] Branched hydroxyproline polymers were prepared using DBU and DIPEA as bases, anhydrous DMSO as solvent, and different equivalents of L-hydroxyproline-N-carboxylic anhydride. The parameters of the products obtained using different raw materials and feed ratios are listed in Table 1 below.

[0217] Table 1. Molecular weight, dispersion, amino content and branching degree of B-PHyp obtained with different base types and feed ratios

[0218] in,

[0219] Molecular weight and dispersity data were determined by aqueous SEC coupled with a multi-angle light scattering / differential detector in 1×PBS (pH=7.4). The dn / dc (658 nm) was determined using linear hydroxyproline to be 0.159 ml / g.

[0220] The molecular weights of DIPEA-500 and above exceed the separation limit of the column, making it impossible to accurately determine molecular weight and dispersion. We also roughly determined the weight-average molecular weight of DIPEA-4000 using static light scattering to be 1,200 kg / mol. (The dn / dc (658 nm) measured using linear hydroxyproline in PBS solution was 0.159 ml / g.)

[0221] Figure 5 shows a schematic diagram of the hyperbranched structure of B-BHyp. Wherein, the number of branching units is recorded as D, the number of terminal units is recorded as T, and the number of linear units is recorded as L. The degree of branching (DB) is defined as (D+T) / (D+T+L). When the molecular weight of the polymer is large, the number of D and T is substantially equivalent, so it can be simplified to 2T / (2T+L). As can be seen from the structural schematic diagram shown in Figure 5, the end group T contains a secondary amine structure, and L contains an amide structure. Therefore, the degree of branching can be calculated by the ratio of the number of secondary amine and amide groups. Therefore, the inventors used proline calibration and the reaction of diazonium salt with secondary amine. The product content was determined by absorption at 340nm, and the number ratio of secondary amine and amide was further characterized (see Figure 9). The results are converted into the degree of branching and are shown in Table 1 below.

[0222] Figures 2-4 show the SEC curves of products obtained using DBU and DIPEA as bases and at different [NCA] / [base] ratios, respectively. Earlier peaks indicate higher molecular weights. These curves were obtained using a Cytiva Superose 6 column.

[0223] From the results shown in Table 1 and Figures 2-4, it can be seen that when different types of organic bases and feed ratios are used, the results show that: when the alkalinity is weaker and the [NCA] / [base] ratio is lower, the molecular weight of the polymer obtained by the reaction is larger.

[0224] Figure 6 shows the infrared absorption spectra comparison of polyacetylhydroxyproline and B-Hyp, which proves that the 1735 cm -1 The peak represents the ester bond absorption peak. Furthermore, the present inventors further demonstrated the branching of the resulting product using H NMR spectroscopy and hydrolysis. Figures 7 and 8 show the H-NMR (DO) and B-PHyp (black) spectra of polyhydroxyproline (PHyp) and linear polyhydroxyproline, respectively, as well as the aqueous phase SEC curve changes after the addition of NaOH (red).

[0225] Example 2. Preparation of physical hydrogel

[0226] The product obtained in Example 1 was taken out of the glove box and placed at room temperature. 500 μL DMSO was added to the system and diluted to 1 ml. Acetic anhydride (47 μL, 0.5 mmol, 0.5 equivalent) was added under rapid stirring, followed by triethylamine (28 μL, 0.2 mmol, 0.2 equivalent). After about 1 minute of reaction, a gel was formed. After continuing the reaction for 2 hours, 15 ml of water was added to dilute, dialyze, and freeze-dry. Take 20 mg of acetylated branched hydroxyproline polymer, add 1000 μL of pure water, heat until dissolved, and return to room temperature to form a hydrogel. Use 1 H-NMR characterized the degree of acetylation, which was approximately 40% (see Figure 10).

[0227] FIG11 shows rheological data of 2 wt % acetyl-branched hydroxyproline polymer hydrogel: A) oscillation frequency sweep curve; B) amplitude sweep curve; C) amplitude step sweep curve; D) injectable schematic diagram.

[0228] From the above results, it can be seen that the physical hydrogel of the present invention has a low gelling solid content (2 wt %) and good deformation recovery ability.

[0229] Example 3. Preparation of branched hydroxyproline-proline copolymers according to the present invention

[0230] In a glove box, L-hydroxyproline-N-carboxylic anhydride (78.5 mg, 0.500 mmol, 100 equiv) and proline-N-carboxylic anhydride (70.5 mg, 0.500 mmol, 100 equiv) were added to a 5 mL glass vial and dissolved in 500 μL of anhydrous DMSO. N,N-diethylisopropylamine (DIPEA) (0.5 M x 10 μL, 0.005 mmol, 1 equiv) was then added and allowed to react for 30 min. The solution was diluted to 2.5 mL with water, passed through a PD10 desalting column, and lyophilized to obtain the final branched hydroxyproline polymer.

[0231] Figure 12 shows the branched hydroxyproline-proline copolymer 1H-NMR (DO) analysis indicated that the ratio of proline to hydroxyproline in the product was consistent with that in the feed. Figure 13 shows the size exclusion chromatography (SEC) curve of the branched hydroxyproline-proline copolymer. The copolymer and B-PHyp (DIPEA-200) exhibited similar elution volumes, indicating that the two copolymers possessed similar molecular weights.

[0232] The foregoing description is merely an exemplary embodiment of the present disclosure and is not intended to limit the scope of protection of the present disclosure. The scope of protection of the present disclosure is determined by the appended claims.

Claims

1. A branched hydroxyproline polymer having a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a degree of branching in the range of 0.01 to 0.

5.

2. The branched hydroxyproline polymer according to claim 1, wherein the weight average molecular weight is in the range of 1 kDa to 1000 kDa and the degree of branching is in the range of 0.01 to 0.

36.

3. The branched hydroxyproline polymer according to claim 1 or 2, having a structure as shown in formula (I): R1-(L1) x -(L2) y -(L3) z -R2 Formula (I) in, L1 is L2 is L3 R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2; R1 is a terminal group; R2 is independently H or a terminal group at each occurrence; Each occurrence of x and y is independently a fraction greater than 0 and less than 1, and each occurrence of z is independently a fraction greater than or equal to 0 and less than 1, provided that the sum of all x, y, and z is 1.

4. The branched hydroxyproline polymer according to any one of claims 1 to 3, wherein z is 0, and the branched polyhydroxyproline polymer has a structure shown in the following formula (II): R1-(L1) x -(L2) y -R2 Formula (II) in, L1 is L2 is R a Each occurrence is independently selected from -(L1) x -(L2) y -R2; R1 is a terminal group; R2 is independently H or a terminal group at each occurrence; Each occurrence of x and y is independently a fraction greater than 0 and less than 1, provided that the sum of all x and y is 1.

5. The branched hydroxyproline polymer according to any one of claims 1 to 3, wherein z is not 0, and the branched hydroxyproline polymer has a structure shown in formula (I): R1-(L1) x -(L2) y -(L3) z -R2 Formula (I) in, L1 is L2 is L3 R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2; R1 is a terminal group; R2 is independently H or a terminal group at each occurrence; Each occurrence of x, y, and z is independently a fraction greater than 0 and less than 1, provided that the sum of all x, y, and z is 1.

6. An at least partially modified branched hydroxyproline polymer having a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a degree of branching in the range of 0.01 to 0.5, wherein the at least partially modified branched hydroxyproline polymer has a structure represented by formula (III): R1'-(L1') x’ -(L2') y’ -(L3') z’ -R2' Formula (III) in, L1' is L2' is L3' R a 'Each occurrence is independently selected from -(L1') x’ -(L2') y’ -(L3') z’ -R2'; R1' is a terminal group; R2' is independently H or a terminal group at each occurrence; Each occurrence of R3 is independently selected from H, C 1-20 Alkyl, C 1-20 Alkenyl, C 1-20 Alkynyl, C 1-20 Alkoxy, C 1-20 Alkenyloxy, C 1-20 Alkynyloxy, C 1-20 Alkoxyalkyl, C 1-20 substituted with a 5-6 membered heteroaryl substituent, wherein the heterocyclic ring and heteroaryl optionally contain one to three heteroatoms independently selected from S, O or N, Each occurrence of x' and y' is independently a fraction greater than 0 and less than 1, and each occurrence of z' is independently a fraction greater than or equal to 0 and less than 1, provided that the sum of all x', y' and z' is 1.

7. The at least partially modified branched hydroxyproline polymer of claim 6, wherein R2' and R3 are independently selected from H, C 1-20 Alkyl, C 1-20 Alkyl acyl, C 1-20 Alkoxy acyl, C 1-20 Alkylsulfonyl and di(C 1-20 The alkyl group in the above group is optionally unsubstituted or substituted by 1 to 3 substituents selected from saturated or unsaturated 5-6 membered heterocyclic groups.

8. The at least partially modified branched hydroxyproline polymer according to claim 6 or 7, wherein R2' and R3 at each occurrence are independently selected from H, acetyl and lipoyl.

9. An at least partially modified branched hydroxyproline polymer according to any one of claims 6 to 8, wherein the degree of modification is between 0.1% and 100%.

10. A method for preparing a branched hydroxyproline polymer, comprising: The compound shown in the following formula: The branched hydroxyproline polymer is polymerized in an organic solvent in the presence of a base to obtain the branched hydroxyproline polymer, wherein the branched hydroxyproline polymer has a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a branching degree in the range of 0.01 to 0.

5.

11. The method of claim 10, wherein: The base is selected from potassium hydride, sodium hydride, potassium methoxide, sodium methoxide, potassium tert-butoxide, sodium tert-butoxide, triazabicyclo[4.4.0]dec-5-ene, 1,8-diazabicyclo[5.4.0]undec-7-ene, N,N-diisopropylethylamine or any mixture thereof.

12. The method according to claim 10 or 11, wherein: The organic solvent is selected from dimethyl sulfoxide, dimethylformamide, acetonitrile, dichloromethane, ethyl acetate, tetrahydrofuran, acetone and a mixture thereof.

13. A method for at least partially modifying a branched hydroxyproline polymer, wherein the branched hydroxyproline polymer has a weight average molecular weight in the range of 1 kDa to 10,000 kDa and a degree of branching in the range of 0.01 to 0.5, the method comprising: The branched hydroxyproline polymer is reacted with a modifying agent in an organic solvent in the presence of a base to at least partially modify the branched hydroxyproline polymer, wherein the branched hydroxyproline polymer has a structure represented by the following formula (I): R1-(L1) x -(L2) y -(L3) z -R2 Formula (I) in, L1 is L2 is L3 R a Each occurrence is independently selected from -(L1) x -(L2) y -(L3) z -R2; R1 is a terminal group; R2 is independently H or a terminal group at each occurrence; x and y are independently fractions greater than 0 and less than 1 at each occurrence, and z is independently fractions greater than or equal to 0 and less than 1 at each occurrence, provided that the sum of all x, y, and z is 1, And wherein the modifying agent is selected from the group consisting of: Where R is selected from C 1-20 alkyl.

14. The method of claim 13, wherein: R is methyl.

15. The method according to claim 13 or 14, wherein: The base is selected from trimethylamine, triethylamine, triethanolamine and mixtures thereof.

16. The method according to any one of claims 13 to 15, wherein: The organic solvent is selected from dimethyl sulfoxide, dimethylformamide, acetonitrile, dichloromethane, ethyl acetate, tetrahydrofuran, acetone and a mixture thereof.

17. A physical hydrogel comprising an at least partially modified branched hydroxyproline polymer according to any one of claims 6 to 9.

18. The physical hydrogel according to claim 17, wherein The at least partially modified branched hydroxyproline polymer is an at least partially acylated branched hydroxyproline polymer.

19. The physical hydrogel according to claim 17 or 18, wherein The at least partially modified branched hydroxyproline polymer is an at least partially acetylated branched hydroxyproline polymer or an at least partially lipoylated branched hydroxyproline polymer.

20. A method for preparing a branched polyamino acid, comprising: The compound of formula (IV): in: R A and R B One of them is C 1-6 Alkyl, another C substituted with hydroxy 1-6 alkyl, Or, R A and R B Together with the ring atoms to which they are connected, they form a five- or six-membered ring substituted with a hydroxyl group, and are polymerized in an organic solvent in the presence of a base to obtain the branched polyamino acid, wherein the weight average molecular weight of the branched polyamino acid is in the range of 1 kDa to 10,000 kDa, and the degree of branching is in the range of 0.01 to 0.

5.

21. The method of claim 20, wherein: The compound of formula (IV) has a structure selected from the group consisting of: Among them, R 1 -(CH2) 1-2 -, R 2 and R 3 Each occurrence is optionally independently selected from H, CH3- and CH3CH2-, The condition is R 1 , R 2 and R 3 The sum of the number of carbon atoms is not greater than 5; R4 is C 1-6 alkyl.

22. The method of claim 20 or 21, wherein: The base is selected from potassium hydride, sodium hydride, potassium methoxide, sodium methoxide, potassium tert-butoxide, sodium tert-butoxide, triazabicyclo[4.4.0]dec-5-ene, 1,8-diazabicyclo[5.4.0]undec-7-ene, N,N-diisopropylethylamine or any mixture thereof.

23. The method of any one of claims 20 to 22, wherein: The organic solvent is selected from dimethyl sulfoxide, dimethylformamide, acetonitrile, dichloromethane, ethyl acetate, tetrahydrofuran, acetone and a mixture thereof.

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