Full-automatic solid-phase polypeptide synthesis system and synthesis method based on flow chemistry

Abstract

The invention discloses a full-automatic solid-phase polypeptide synthesis system and a full-automatic solid-phase polypeptide synthesis method based on flow chemistry, and belongs to the technical field of chemical synthesis automation equipment. Wherein the liquid storage module is used for respectively storing different liquid reaction reagents, including a deprotection reagent, a main solvent, an activating agent, alkali and amino acid, required by solid-phase polypeptide synthesis; the transmission module is connected between the liquid storage module and the mixer and is used for selecting and accurately extracting the corresponding required liquid reaction reagent from the liquid storage module to the mixer according to different stages of solid-phase polypeptide synthesis, and the liquid reaction reagent is rapidly and fully mixed when flowing through the mixer; the preheating module is connected between the mixer and the reactor, and is used for preheating the mixed liquid reaction reagent from the mixer, so that the preheated liquid reaction reagent completes the reaction of the corresponding stage of solid-phase polypeptide synthesis when flowing through the reactor. The method is high in polypeptide synthesis speed, high in synthesized polypeptide purity, simple in instrument structure and low in cost.

Description

Technical Field

[0001] This invention relates to the field of automated equipment technology for chemical synthesis, and in particular to a fully automated solid-phase peptide synthesis system and method based on flow chemistry. Background Technology

[0002] Solid-phase peptide synthesis systems, based on the principle of solid-phase peptide synthesis, have become key instruments in peptide synthesis, playing an irreplaceable role in basic research and drug development. However, the development of peptide synthesis systems in China is currently slow, facing problems such as slow synthesis speed and low purity of synthesized peptides. Existing imported instruments are expensive and complex in structure, resulting in high operating and maintenance costs for domestic universities, research institutions, and drug development companies, making it difficult to meet the growing research needs in the field of peptides. Summary of the Invention

[0003] This invention aims to at least partially solve one of the technical problems in related technologies. Therefore, one objective of this invention is to propose a fully automated solid-phase peptide synthesis system and method based on flow chemistry, which offers fast synthesis speed, high peptide purity, simple structure, and low cost.

[0004] The fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention includes a liquid storage module, a transport module, a mixer, a preheating module, a reactor, and a control module; The liquid storage module is used to store different liquid reaction reagents required for solid-phase peptide synthesis, including deprotection reagents, main solvents, activators, bases, and amino acids. The transmission module is connected between the liquid storage module and the mixer, and is used to select and precisely extract the corresponding liquid reaction reagents from the liquid storage module to the mixer according to different stages of solid-phase peptide synthesis, so that the corresponding liquid reaction reagents are quickly and thoroughly mixed when flowing through the mixer. The preheating module is connected between the mixer and the reactor. The preheating module preheats the mixed liquid reaction reagent from the mixer so that the preheated liquid reaction reagent completes the corresponding stage of solid-phase peptide synthesis when it flows through the reactor. The control module controls the operation of the fully automated solid-phase peptide synthesis system based on flow chemistry.

[0005] The working principle of the fully automated solid-phase peptide synthesis system based on flow chemistry in this invention embodiment is as follows: according to the target peptide sequence of solid-phase peptide synthesis, the corresponding amino acids are coupled sequentially on the solid-phase resin in the reactor. The coupling of each type of amino acid is carried out in a cycle of "deprotection reaction - deprotection flow path cleaning - condensation reaction - condensation reaction flow path cleaning". One cycle completes the coupling of one type of amino acid. The cycle of "deprotection reaction - deprotection flow path cleaning - condensation reaction - condensation reaction flow path cleaning" is repeated until all amino acids are coupled.

[0006] One cycle completes the coupling process of one amino acid, which is divided into four stages, as follows: Deprotection reaction: The deprotecting agent in the storage module is selected and precisely extracted through the transfer module, and the extracted deprotecting agent flows sequentially through the transfer module, the mixer, the preheating module and the reactor. The deprotecting agent deprotects the Fmoc groups on the solid resin in the reactor, exposing the amine reactive groups.

[0007] Deprotection flow path cleaning: The main solvent in the storage module is selected and precisely extracted through the transfer module to flush the deprotection reaction flow path from the previous step.

[0008] Condensation reaction: The activator, alkali, and corresponding amino acids in the storage module are simultaneously selected and precisely extracted through the transmission module, so that the activator, alkali, and corresponding amino acids flow through the mixer and are quickly and thoroughly mixed. When the mixed liquid reaction reagent flows through the preheating module, it is preheated and activated to form a preheated liquid reaction reagent. When the preheated liquid reaction reagent flows through the reactor, a condensation reaction occurs. The condensation reaction is a condensation reaction between the activated amino acids and the amine reactive groups on the resin in the reactor.

[0009] Condensation reaction flow path cleaning: The main solvent in the storage module is precisely extracted through the transfer module to flush the condensation reaction flow path from the previous step.

[0010] It should be noted that the flow path downstream of the outlet of the transport module in the fully automated solid-phase peptide synthesis system based on flow chemistry of this invention is a common flow path for the above four stages. Therefore, after the deprotection reaction is completed, the deprotection reaction flow path needs to be cleaned to avoid the deprotection reagent remaining in the flow path. After the condensation reaction is completed, the condensation reaction flow path needs to be cleaned to avoid the mixed reaction solution of activator, base and amino acid reagent remaining in the flow path, thereby improving the purity of the synthesized peptide.

[0011] The fully automated solid-phase peptide synthesis system based on flow chemistry of this invention has the following technical advantages: First, by setting up the transfer module, the required liquid reaction reagents can be precisely extracted from the storage module according to different stages of solid-phase peptide synthesis. The flow rate of the liquid reaction reagents can be precisely controlled by the transfer module, such as the flow rate and time of a volumetric pump, with a large variable flow rate range, which is beneficial for achieving rapid and efficient solid-phase peptide synthesis. Second, by setting up the mixer, different liquid reaction reagents can be quickly and thoroughly mixed when flowing through the mixer simultaneously, which is beneficial for achieving rapid and efficient solid-phase peptide synthesis. Third, by setting up the preheating module, the reaction liquid reagents are preheated to a predetermined temperature before synthesis. Pre-activation allows for precise control of amino acid activation temperature, facilitating rapid and efficient solid-phase peptide synthesis. Fourth, the reactor is a unidirectional flow reactor, offering advantages such as faster synthesis speed, higher efficiency, easier scale-up, and higher safety compared to traditional batch reactors. Fifth, all reactions are completed within a single reactor, eliminating the need for resin transfer, significantly simplifying the operation process. The coordinated control of selector valves and volumetric pumps enables liquid reagent delivery, significantly reducing equipment costs and space requirements. Sixth, the synthesis method can be customized through programming, and commands can be executed automatically, precisely controlling the transfer of amino acids to the mixer for pre-activation. Precise programming enables automatic deprotection reactions and solid-phase condensation, making operation flexible and convenient.

[0012] The fully automated solid-phase peptide synthesis system based on flow chemistry of this invention is achieved by sequentially connecting the transport module, the mixer, the preheating module, and the reactor. A control module controls the collaborative operation of these modules, precisely controlling the addition, activation, and reaction of liquid reagents. This enables continuous flow chemistry-based solid-phase peptide synthesis, resulting in fast synthesis speed, high efficiency, high peptide purity, and easy scale-up. It allows for the synthesis of high-molecular-weight peptides according to actual needs, greatly expanding the diversity of peptide synthesis. The system is structurally sound, low-cost, safe and flexible to use, and widely applicable, meeting the needs of peptide synthesis from laboratory to pilot-scale operations.

[0013] In some embodiments, the transmission module includes a first transmission component, a second transmission component, and a third transmission component; The first transmission component is connected between the liquid storage module and the inlet of the mixer, and is used to accurately extract alkali from the liquid storage module to the mixer; The second transmission component is connected between the reservoir module and the inlet of the mixer, and is used to select and precisely extract the deprotection reagent, main solvent and activator from the reservoir module to the mixer; The third transmission component is connected between the liquid storage module and the inlet of the mixer, and is used to select and precisely extract amino acids and main solvent from the liquid storage module to the mixer.

[0014] In some embodiments, the liquid storage module includes reagent bottles for storing different types of liquid reaction reagents, wherein at least 20 of the reagent bottles are used to store at least 20 kinds of amino acids respectively; The first transfer component includes a first volumetric pump, the inlet of which is connected to the reagent bottle storing the alkali, and the outlet of which is connected to the inlet of the mixer. The second transfer component includes a first selection valve and a second volumetric pump. The inlet of the second volumetric pump is connected to the reagent bottle storing the deprotection reagent, the reagent bottle storing the main solvent, and the reagent bottle storing the activator through the first selection valve. The outlet of the second volumetric pump is connected to the inlet of the mixer. The third transfer component includes a second selection valve and a third volumetric pump. There is at least one second selection valve. The inlet of the third volumetric pump is connected to the reagent bottle storing the main solvent and the reagent bottle storing amino acids respectively through at least one of the second selection valves. The outlet of the third volumetric pump is connected to the inlet of the mixer.

[0015] In some embodiments, the plurality of inlet ends of the first selection valve are respectively connected to the plurality of reagent bottles storing a plurality of activators, the reagent bottle storing a deprotection reagent, and the reagent bottle storing the main solvent; the outlet end of the first selection valve is connected to the inlet end of the second volumetric pump; and the outlet end of the second volumetric pump is connected to the inlet end of the mixer. There are multiple second selection valves. The multiple inlet ends of one second selection valve are respectively connected to the outlet ends of the remaining second selection valves. The outlet end of one second selection valve is connected to the inlet end of the third volumetric pump. The outlet end of the third volumetric pump is connected to the inlet end of the mixer. The multiple outlet ends of the remaining second selection valves are respectively connected to the reagent bottle storing different types of amino acids and the reagent bottle storing the main solvent.

[0016] In some embodiments, the mixer is a spiral column structure consisting of a liquid conduit wound around a cylindrical rod.

[0017] In some embodiments, the inner diameter of the liquid conduit of the mixer is 0.75 to 1 mm, the outer diameter of the liquid conduit of the mixer is 1.6 to 2 mm, the diameter of the cylindrical rod is 6 to 10 mm, and the number of spiral turns of the mixer is at least 20.

[0018] In some embodiments, the preheating module includes a converter and a plurality of heating devices connected in parallel to the converter, the converter being connected between the mixer and the reactor, and the plurality of heating devices having different preheating temperatures.

[0019] In some embodiments, the heating device includes: A heating element, wherein a first temperature control component is provided inside the heating element, and the first temperature control component is adapted to heat the heating element; A heat-insulating outer tube is sleeved around the outer periphery of the heating core; wherein... At least one of the heating core and the heat-insulating outer tube has a spiral groove; after the heating core is sleeved with the heat-insulating outer tube, a heating cavity is formed at the spiral groove; the heating cavity is suitable for accommodating liquid pipelines.

[0020] In some embodiments, the reactor includes: A base on which a support assembly is provided; A first fixing seat is disposed on the bracket assembly, and a first fluid channel is formed inside the first fixing seat; A second fixing seat is disposed on the first fixing seat and is detachably connected to the first fixing seat; a reaction chamber and a second fluid channel are formed inside the second fixing seat, the reaction chamber is open to the upward, and the second fluid channel communicates with the reaction chamber and the first fluid channel. A reaction tube is disposed within the reaction chamber and is adapted to load a solid support; the reaction tube has a fluid inlet and a fluid outlet, and the fluid outlet is connected to the second fluid channel. A cover body is disposed on the second fixed base and is detachably connected to the second fixed base; and a third fluid channel is formed in the cover body, the third fluid channel being connected to the fluid inlet. A second temperature control component is disposed on the first fixed base and / or the second fixed base to regulate the temperature inside the reaction chamber.

[0021] In some embodiments, a gas path module is also included; the gas path module is connected to the liquid storage module and is used to input protective gas into the liquid storage module to protect each liquid reaction reagent.

[0022] This invention also proposes a fully automated solid-phase peptide synthesis method based on flow chemistry.

[0023] According to the fully automated solid-phase peptide synthesis method based on flow chemistry of the present invention, solid-phase peptide synthesis is performed using the fully automated solid-phase peptide synthesis system based on flow chemistry of the present invention described above, and includes the following steps. S1: Deprotection reaction: The deprotection agent in the storage module is selected and precisely extracted through the transfer module, and the extracted deprotection agent flows sequentially through the transfer module, the mixer, the preheating module and the reactor. The deprotection agent deprotects the Fmoc groups on the solid resin in the reactor, exposing the amine reactive groups. S2: Deprotection flow path cleaning: The main solvent in the storage module is selected and precisely extracted through the transfer module to rinse the deprotection reaction flow path in step S1; S3: Condensation reaction: The activator, alkali, and corresponding amino acids in the storage module are simultaneously and precisely selected and extracted through the transmission module, so that the activator, alkali, and corresponding amino acids flow through the mixer in sequence and are quickly and thoroughly mixed. When the mixed liquid reaction reagent flows through the preheating module, it is preheated and activated to form a preheated liquid reaction reagent. When the preheated liquid reaction reagent flows through the reactor, a condensation reaction occurs. The condensation reaction is a condensation reaction between the activated amino acids and the amine reactive groups on the resin in the reactor. S4: Condensation reaction flow path cleaning: The main solvent in the storage module is selected and precisely extracted through the transfer module to flush the condensation reaction flow path in step S3. S5: Repeat steps S1 to S4 in a loop until all amino acid coupling is completed.

[0024] In other words, solid-phase peptide synthesis involves sequentially coupling the corresponding types of amino acids onto the solid-phase resin in the reactor according to the target peptide sequence. The coupling of each type of amino acid is carried out in a cycle of "deprotection reaction - deprotection flow path cleaning - condensation reaction - condensation reaction flow path cleaning". One cycle completes the coupling of one type of amino acid. The cycle of "deprotection reaction - deprotection flow path cleaning - condensation reaction - condensation reaction flow path cleaning" is repeated until all amino acids are coupled.

[0025] Since the fully automated solid-phase peptide synthesis method based on flow chemistry in this embodiment of the invention uses the fully automated solid-phase peptide synthesis system based on flow chemistry in the aforementioned embodiment of the invention, the fully automated solid-phase peptide synthesis method based on flow chemistry in this embodiment of the invention has essentially the same technical effects as the fully automated solid-phase peptide synthesis system based on flow chemistry in the aforementioned embodiment of the invention, and will not be repeated here.

[0026] Additional aspects and advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. Attached Figure Description

[0027] Figure 1This is a schematic diagram of the principle of a fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention; Figure 2 This is a schematic diagram of the deprotection reaction flow path (dashed line) of the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention; Figure 3 This is a schematic diagram of the deprotection flow path cleaning (dashed line) of the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention; Figure 4 This is a schematic diagram (dashed line) of a condensation reaction flow path of a fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention; Figure 5 This is a schematic diagram of the cleaning (dashed line) of a condensation reaction flow path in a fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention. Figure 6 This is a schematic diagram of the gas path module of the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention; Figure 7 This is a schematic diagram of one end face of the first selection valve of the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention, showing that the first selection valve has multiple inlet ends; Figure 8 This is a schematic diagram of another end face of the first selection valve of the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention, showing that the first selection valve has an outlet end; Figure 9 This is a schematic diagram of the mixer in a fully automated solid-phase peptide synthesis system based on flow chemistry, according to an embodiment of the present invention. Figure 10 This is a schematic diagram of the heating device of the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention. Figure 11 This is a schematic diagram of the heating core of the heating device in the fully automated solid-phase peptide synthesis system based on flow chemistry, according to an embodiment of the present invention. Figure 12 This is a schematic diagram of the first cross-sectional structure of the reactor in the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention. Figure 13 This is a schematic diagram of the second cross-sectional structure of the reactor in the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention. Figure 14 This is a schematic cross-sectional view of the second station of the reactor in the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention. Figure 15This is a schematic diagram illustrating the principle of deprotection and condensation reactions in the reactor of the fully automated solid-phase peptide synthesis system based on flow chemistry, according to an embodiment of the present invention. Figure 16 To synthesize ACP 65-74 Schematic diagram of high performance liquid chromatography analysis of crude peptides; Figure 17 To synthesize ACP 65-74 Schematic diagram of mass spectrometry analysis; Figure 18 This is a schematic diagram of the high-performance liquid chromatography analysis of the synthesized crude ABC-20 peptide; Figure 19 This is a schematic diagram of the mass spectrometry analysis of the synthesized ABC-20. Figure 20 This is a schematic diagram showing the real-time monitoring results of the ultraviolet monitoring module during the synthesis of peptide ABC-20 using the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention. Figure 21 This is a schematic diagram of the high-performance liquid chromatography analysis of the synthesized crude HD-6 peptide; Figure 22 This is a schematic diagram of the mass spectrometry analysis of synthesized HD-6; Figure 23 This is a list of the meanings of English words or English abbreviations appearing in this invention.

[0028] Figure Labels Liquid storage module 1; Reagent bottle 101; Transmission module 2; First transmission component 201; First volumetric pump 2011; Second transmission assembly 202; first selection valve 2021; second volumetric pump 2022; Third transmission component 203; Second selection valve 2031; Third volumetric pump 2032; Four-way valve 3; Mixer 4; Liquid tube 401; cylindrical rod 402; Preheating module 5; Converter 501; Heating device 502; Heating core 5021; Protruding edge 50211; Notch 50212; Mounting cavity 50213; Detection hole 50214; Insulated outer tube 5022; Spiral groove 5023; Plug 5024; Through hole 50241; limit pin 5025; pin hole 5026; Reactor 6; Base 610; support rod 620; first fixed seat 630; first fluid channel 631; First mounting channel 632; temperature sensor mounting cavity 633; ​​first fastening hole 634; Second fixed seat 640; reaction chamber 641; second fluid channel 642; heating chamber 643; Limiting groove 644; First sealing ring mounting groove 645; Second sealing ring mounting groove 646; Second fastening hole 647; cover 650; third fluid channel 651; guide section 652; Third sealing ring mounting groove 653; First sieve plate 661; Second sieve plate 662; Heating rod fixing assembly 670; Air circuit module 7; Air source 701; pressure regulator 702; multi-port air valve 703; rotor flow meter 704; Ultraviolet monitoring module 8; Back pressure valve 9; 10 waste liquid containers. Detailed Implementation

[0029] Embodiments of the present invention are described in detail below, examples of which are shown in the accompanying drawings, wherein the same or similar reference numerals denote the same or similar elements or have the same or similar functions throughout. The following is combined Figures 1 to 18 This invention describes a fully automated solid-phase peptide synthesis system and method based on flow chemistry, as described in embodiments of the present invention.

[0030] like Figures 1 to 14 As shown, the fully automated solid-phase peptide synthesis system based on flow chemistry according to an embodiment of the present invention includes a liquid storage module 1, a transport module 2, a mixer 4, a preheating module 5, a reactor 6, an ultraviolet monitoring module 8, and a control module.

[0031] The storage module 1 is used to store different liquid reaction reagents required for solid-phase peptide synthesis. These liquid reaction reagents include deprotection reagents, main solvents, activators, bases, and amino acids. The deprotection reagent can be piperidine or morpholine, which are commonly used. The main solvent can be DMF, which serves as both a solvent for dissolving amino acids and activators, and is also used to clean the corresponding flow paths. Activators can be PyAOP, PyBOP, HATU, HBTU, TBTU, etc., and can be flexibly selected according to requirements. Bases can be DIEA, DBU, etc., and amino acids can be a variety of amino acids, such as the existing 20 natural amino acids, or the existing 20 natural amino acids and a variety of non-natural amino acids. Each of the above-mentioned different liquid reaction reagents is stored independently in the storage module. For example, the storage module 1 uses multiple reagent bottles 101, each storing one type of liquid reaction reagent, and the different liquid reaction reagents do not interfere with each other.

[0032] The transfer module 2 is connected between the liquid storage module 1 and the mixer 4. It is used to select and precisely extract the corresponding liquid reaction reagents from the liquid storage module 1 to the mixer 4 according to different stages of solid-phase peptide synthesis, ensuring rapid and thorough mixing of the required liquid reaction reagents as they flow through the mixer 4. For example, during the deprotection reaction stage of solid-phase peptide synthesis, such as... Figure 2 As shown, the transfer module 2 precisely extracts the deprotection reagent from the storage module 1 and feeds it to the mixer 4; during the cleaning stage of solid-phase peptide synthesis, as... Figure 3 and Figure 5 As shown, the transfer module 2 precisely extracts the main solvent from the storage module 1 and supplies it to the mixer 4; during the condensation reaction stage of solid-phase peptide synthesis, as... Figure 4 As shown, the transfer module 2 simultaneously and precisely extracts the alkali, activator, and one of the amino acids from the storage module 1 and feeds them to the mixer 4. The mixer 4 can rapidly and thoroughly mix the liquid reaction reagents flowing through it. For example, during the condensation reaction stage of solid-phase peptide synthesis, the mixer 4 efficiently and thoroughly mixes the alkali, activator, and one of the amino acids flowing through it simultaneously. The transfer module 2, under the control of the control module, enables the selection of the type of liquid reaction reagent and the precise extraction of the selected reagent.

[0033] A preheating module 5 is connected between the mixer 4 and the reactor 6. The preheating module 5 preheats the mixed liquid reaction reagent from the mixer 4, ensuring that the preheated liquid reaction reagent completes the corresponding stage of solid-phase peptide synthesis when it flows into the reactor 6. By connecting the preheating module 5 between the outlet of the mixer 4 and the inlet of the reactor 6, the mixed liquid reaction reagent flowing from the outlet of the mixer 4 is preheated as it passes through the preheating module 5. The preheated liquid reaction reagent flows out from the outlet of the preheating module 5 and into the reactor 6. After passing through the reactor 6, it becomes waste liquid and is discharged from the outlet of the reactor 6.

[0034] The ultraviolet monitoring module 8 is located at the rear end of the reactor 6 and is used to monitor the ultraviolet absorbance of the liquid flowing out of the reactor 6 in real time, thereby providing online feedback on the efficiency of the amino acid condensation reaction and deprotection reaction, and realizing online evaluation and feedback of the synthesis efficiency.

[0035] The control module manages the operation of the fully automated solid-phase peptide synthesis system based on flow chemistry. By setting up the control module, other related modules such as transport module 2, preheating module 5, reactor 6, and ultraviolet monitoring module 8 can work together to achieve fully automated solid-phase peptide synthesis based on flow chemistry with high synthesis efficiency.

[0036] like Figures 2 to 5 and Figure 15As shown, the working principle of the fully automated solid-phase peptide synthesis system based on flow chemistry in this embodiment of the invention is as follows: according to the target peptide sequence of solid-phase peptide synthesis, the corresponding amino acids are coupled sequentially on the solid-phase resin in reactor 6. The coupling of each type of amino acid is carried out in a cycle of "deprotection reaction - deprotection flow path cleaning - condensation reaction - condensation reaction flow path cleaning". The coupling of one type of amino acid is completed in one cycle. The cycle of "deprotection reaction - deprotection flow path cleaning - condensation reaction - condensation reaction flow path cleaning" is repeated until all amino acids are coupled.

[0037] One cycle completes the coupling process of one amino acid, which is divided into four stages, as follows: Deprotection reaction: such as Figure 2 As shown, the deprotecting agent in the storage module 1 is selected and precisely extracted by the transfer module 2, and the extracted deprotecting agent flows sequentially through the transfer module 2, mixer 4, preheating module 5 and reactor 6. The deprotecting agent deprotects the Fmoc groups on the solid resin in reactor 6, exposing the amine reactive groups.

[0038] Cleaning of the protection flow path: such as Figure 3 As shown, the main solvent in the storage module 1 is selected and precisely extracted by the transmission module 2 to flush the deprotection reaction flow path of the previous step.

[0039] Condensation reaction: such as Figure 4 As shown, the activator, alkali and corresponding amino acids in the storage module 1 are simultaneously selected and precisely extracted by the transmission module 2, so that the activator, alkali and corresponding amino acids flow through the mixer 4 at the same time and are quickly and thoroughly mixed. When the mixed liquid reaction reagent flows through the preheating module 5, it is preheated and activated to form a preheated liquid reaction reagent. When the preheated liquid reaction reagent flows through the reactor 6, it undergoes a condensation reaction. The condensation reaction is the condensation reaction between the activated amino acids and the amine reactive groups on the resin in the reactor 6.

[0040] Condensation reaction flow path cleaning: such as Figure 5 As shown, the main solvent in the storage module 1 is precisely extracted by the transfer module 2 to flush the condensation reaction flow path of the previous step.

[0041] It should be noted that the flow path downstream of the outlet of the transport module 2 in the fully automated solid-phase peptide synthesis system based on flow chemistry of this embodiment is a common flow path for the above four stages. Therefore, after the deprotection reaction is completed, the deprotection reaction flow path needs to be cleaned to avoid the deprotection reagent remaining in the flow path. After the condensation reaction is completed, the condensation reaction flow path needs to be cleaned to avoid the mixed reaction solution of activator, base and amino acid reagent remaining in the flow path, thereby improving the purity of the synthesized peptide.

[0042] The fully automated solid-phase peptide synthesis system based on flow chemistry of this invention has the following technical advantages: First, by setting up a transfer module 2, the required liquid reaction reagents can be precisely extracted from the storage module 1 according to different stages of solid-phase peptide synthesis. The flow rate of the liquid reaction reagents can be precisely controlled by the transfer module, such as the flow rate and time of a volumetric pump, with a large variable flow rate range, which is beneficial for achieving rapid and efficient solid-phase peptide synthesis. Second, by setting up a mixer 4, different liquid reaction reagents can be quickly and thoroughly mixed when flowing through the mixer 4 simultaneously, which is beneficial for achieving rapid and efficient solid-phase peptide synthesis. Third, by setting up a preheating module 5, the reaction liquid reagents are preheated to a predetermined temperature for pre-activation, which can precisely control the amino acid activation temperature, which is beneficial for achieving rapid and efficient solid-phase peptide synthesis. Fourth, the reaction... The reactor 6 is a unidirectional flow reactor, which has advantages over traditional batch reactors, such as faster synthesis speed, higher efficiency, easier scale-up, and higher safety. Fifth, all reactions are completed within a single reactor 6, eliminating the need for resin transfer and significantly simplifying the operation process. Liquid reagent delivery is achieved through the coordinated control of selector valves and volumetric pumps, significantly reducing equipment costs and space requirements. Sixth, it is equipped with an ultraviolet monitoring module 8, which monitors the ultraviolet absorbance of the liquid flowing out of reactor 6 in real time, thereby providing online feedback on the efficiency of amino acid condensation and deprotection reactions, enabling online evaluation and feedback of synthesis efficiency. Seventh, through programmed custom synthesis methods, commands are executed automatically, precisely controlling the transfer of amino acids to mixer 4 for pre-activation. Deprotection reactions and solid-phase condensation are performed automatically through precise programming, making operation flexible and convenient.

[0043] The fully automated solid-phase peptide synthesis system based on flow chemistry in this invention consists of a transport module 2, a mixer 4, a preheating module 5, and a reactor 6 connected sequentially. A control module manages the collaborative operation of these modules, precisely controlling the addition, activation, and reaction of liquid reagents. This enables continuous flow chemistry-based solid-phase peptide synthesis, resulting in rapid, efficient, and highly pure synthesized peptides. The system is easily scaled up and can synthesize high-molecular-weight peptides according to specific needs, greatly expanding the diversity of peptide synthesis. It features a rational structure, low cost, safe and flexible operation, and wide applicability, meeting the needs of peptide synthesis from laboratory to pilot-scale. Furthermore, the ultraviolet monitoring system at the reactor's downstream end provides real-time online feedback on the reaction efficiency of each step in the synthesis process, facilitating the optimization of reaction conditions.

[0044] In some embodiments, the transmission module 2 includes a first transmission component 201, a second transmission component 202, and a third transmission component 203.

[0045] The first transmission component 201 is connected between the inlet end of the liquid storage module 1 and the mixer 4, and is used to accurately extract alkali from the liquid storage module 1 to the mixer 4.

[0046] The second transmission component 202 is connected between the liquid storage module 1 and the inlet of the mixer 4, and is used to select and precisely extract the deprotection reagent, main solvent and activator from the liquid storage module 1 to the mixer 4, selecting one liquid reaction reagent at a time.

[0047] The third transmission component 203 is connected between the inlet end of the liquid storage module 1 and the mixer 4, and is used to select and accurately extract amino acids and main solvent from the liquid storage module 1 to the mixer 4, selecting one liquid reaction reagent at a time.

[0048] In other words, the transmission module 2 is divided into three paths according to the different types of liquid reaction reagents: the first transmission component 201 is a separate path for alkali, the second transmission component 202 is a separate path for activator and deprotection reagent, and the third transmission component 203 is a separate path for amino acid reagent. The second transmission component 202 and the third transmission component 203 are respectively connected to the reagent bottle 101 of the liquid storage module 1 that stores the main solvent.

[0049] During solid-phase peptide synthesis, the transport module 2 operates under the control of the control module. When entering the deprotection reaction stage, the second transport component 202 selects and precisely extracts the deprotection reagent, allowing it to flow sequentially through the second transport component 202, mixer 4, preheating module 5, and reactor 6. When entering the deprotection flow path cleaning stage, the second transport component 202 selects and precisely extracts the main solvent, allowing it to flow sequentially through the second transport component 202, mixer 4, preheating module 5, and reactor 6. When entering the condensation reaction stage, the first transport component 201 precisely extracts... Simultaneously, the second transfer component 202 selects and precisely extracts the activator, and the third transfer component 203 selects and precisely extracts the corresponding type of amino acid. This allows the alkali, activator, and corresponding type of amino acid to simultaneously enter the mixer 4 and flow sequentially through the preheating module 5 and the reactor 6. When entering the condensation reaction flow path cleaning stage, the second transfer component 202 and the third transfer component respectively select and precisely extract the main solvent. The main solvent then flows through the second transfer component 202 and the third transfer component 203 respectively, and then simultaneously flows sequentially through the mixer 4, the preheating module 5, and the reactor 6. Therefore, the transfer module 2, with its simple structure consisting of the first transfer component 201, the second transfer component 202, and the third transfer component 203, also simplifies the logic of the control module, making control convenient and less prone to errors.

[0050] In some embodiments, the liquid storage module 1 includes reagent bottles 101 for storing different types of liquid reaction reagents, wherein at least 20 reagent bottles 101 are used to store at least 20 kinds of amino acids, which is beneficial to expanding the diversity of peptide synthesis and to synthesizing high molecular weight synthetic peptides.

[0051] The first transfer component 201 includes a first volumetric pump 2011. The inlet of the first volumetric pump 2011 is connected to the reagent bottle 101 storing alkali, and the outlet of the first volumetric pump 2011 is connected to the inlet of the mixer 4. Thus, the first volumetric pump 2011 operates under the control of the control module, providing power to draw alkali from the reagent bottle 101. Simultaneously, it can precisely control the flow rate of the alkali, thereby precisely controlling the amount of alkali drawn, achieving accurate alkali extraction. The first volumetric pump 2011 can adjust the flow rate of the alkali, with a wide variable flow rate range.

[0052] The second transfer assembly 202 includes a first selection valve 2021 and a second volumetric pump 2022. The inlet of the second volumetric pump 2022 is connected via the first selection valve 2021 to a reagent bottle 101 storing deprotection reagent, a reagent bottle 101 storing main solvent, and a reagent bottle 101 storing activator, respectively. The outlet of the second volumetric pump 2022 is connected to the inlet of the mixer 4. Figure 1 and Figure 7 and Figure 8 As shown, the first selector valve 2021 has multiple inlet terminals (such as...). Figure 7 The diagram shows that the first selector valve 2021 has ten inlet terminals and one outlet terminal (e.g., ...). Figure 8 As shown, the first selection valve 2021 opens the corresponding inlet end of the first selection valve 2021 under the control of the control module according to different stages of solid-phase peptide synthesis. The second volumetric pump 2022 operates under the control of the control module, which can provide power and precisely control the flow rate, thereby precisely controlling the extraction volume. The second volumetric pump 2022 can adjust the flow rate, and the flow rate has a large variable range. For example, when entering the deprotection stage, the first selection valve 2021 opens the inlet end connected to the reagent bottle 101 storing the deprotection reagent, and at the same time, the second volumetric pump 2022 precisely extracts the deprotection reagent; as another example, when entering the deprotection flow path cleaning or condensation reaction flow path cleaning stage, the first selection valve 2021 opens the inlet end connected to the reagent bottle 101 storing the main solvent, and at the same time, the second volumetric pump 2022 precisely extracts the main solvent; as yet another example, when entering the condensation reaction stage, if there are two reagent bottles 101 storing activators, storing two types of activators such as PyAOP and HATU respectively, then the first selection valve 2021 opens the inlet end connected to one of the reagent bottles 101 storing activators, and at the same time, the second volumetric pump 2022 precisely extracts the corresponding type of activator.

[0053] The third transfer assembly 203 includes a second selection valve 2031 and a third volumetric pump 2032. There is at least one second selection valve 2031. The inlet of the third volumetric pump 2032 is connected to a reagent bottle 101 storing the main solvent and a reagent bottle 101 storing amino acids, respectively, via at least one second selection valve 2031. The outlet of the third volumetric pump 2032 is connected to the inlet of the mixer 4. The structure of the second selection valve 2031 is similar to that of the first selection valve 2021. The configuration of the second selection valve 2031 and the third volumetric pump 2032 are similar to the functions of the first selection valve 2021 and the second volumetric pump 2022 described above, and will not be repeated here.

[0054] It should be noted that the number of reagent bottles 101 storing the same main solvent can be set as needed. For example, setting three bottles can save space and simplify logic control.

[0055] In some embodiments, multiple inlet terminals of the first selection valve 2021 are respectively connected to multiple reagent bottles 101 storing multiple activators, a reagent bottle 101 storing deprotection reagents, and a reagent bottle 101 storing the main solvent. The outlet terminal of the first selection valve 2021 is connected to the inlet terminal of the second volumetric pump 2022, and the outlet terminal of the second volumetric pump 2022 is connected to the inlet terminal of the mixer 4. The multiple reagent bottles 101 storing multiple activators can be understood as multiple activators being stored separately in multiple reagent bottles 101. For example, the two activators PyAOP and HATU are stored separately in two reagent bottles 101.

[0056] There are multiple second selection valves 2031. Multiple inlet terminals of one second selection valve 2031 are connected to the outlet terminals of the remaining second selection valves 2031. The outlet terminal of one second selection valve 2031 is connected to the inlet terminal of a third volumetric pump 2032, and the outlet terminal of the third volumetric pump 2032 is connected to the inlet terminal of the mixer 4. The multiple outlet terminals of the remaining second selection valves 2031 are connected to reagent bottles 101 storing different types of amino acids and reagent bottles 101 storing the main solvent. The number of second selection valves 2031 can be set according to the number of amino acid types. For example, if there are 20 types of amino acids, four second selection valves 2031 can be set.

[0057] Therefore, it is beneficial to achieve peptide synthesis and makes the logic control simpler, more convenient, and less prone to errors.

[0058] In some embodiments, the outlet ends of the first volumetric pump 2011, the second volumetric pump 2022, and the third volumetric pump 2032 are connected to the inlet end of the mixer 4 via a four-way valve 3.

[0059] In some embodiments, a check valve is provided between the first volumetric pump 2011 and the mixer 4, the second volumetric pump 2022 and the mixer 4, and the third volumetric pump 2032 and the four-way valve 3.

[0060] In some embodiments, such as Figure 9 As shown, the mixer 4 is a spiral column structure formed by a liquid pipe 401 coiled around a cylindrical rod 402. This mixer 4 has a simple structure, short mixing time, and high mixing efficiency.

[0061] In some embodiments, the inner diameter of the liquid pipe 401 of the mixer 4 is 0.75~1 mm, the outer diameter of the liquid pipe 401 of the mixer 4 is 1.6~2 mm, the diameter of the cylindrical rod 402 is 6~10 mm, and the number of spiral turns of the mixer 4 is at least 20. The mixer 4 has a simple structure, short mixing time, and high mixing efficiency.

[0062] Mixer 4 is made of heat- and corrosion-resistant materials, such as polytetrafluoroethylene, which is safe and reliable.

[0063] In some embodiments, such as Figure 1 As shown, the preheating module 5 includes a converter 501 and multiple heating devices 502 connected in parallel to the converter 501. The converter 501 is connected between the mixer 4 and the reactor 6, and the preheating temperatures of the multiple heating devices 502 are different. The converter 501 and the heating devices 502 operate under the control of the control module. The converter 501 can switch any one of the multiple heating devices 502 to the position between the mixer 4 and the reactor 6, allowing the liquid reaction reagent to flow from the mixer 4 through the connected heating device 502 and then through the reactor 6. Since the multiple heating devices 502 are pre-set and heated to different preheating temperatures, the converter 501 can switch the corresponding heating device 502 to the position between the mixer 4 and the reactor 6 according to the preheating temperature required by the liquid reaction reagent. This improves the preheating efficiency and meets the requirements of solid-phase peptide synthesis under different preheating temperature conditions, which is beneficial to improving the synthesis efficiency.

[0064] like Figure 10 and Figure 11As shown, the heating device 502 includes a heating core 5021, a heat-insulating outer tube 5022, and a liquid pipeline. A first temperature control component is provided inside the heating core 5021, which is adapted to heat the heating core 5021. The heat-insulating outer tube 5022 is sleeved on the outer periphery of the heating core 5021. At least one of the heating core 5021 and the heat-insulating outer tube 5022 forms a spiral groove 5023. After the heating core 5021 and the heat-insulating outer tube 5022 are sleeved, a heating cavity is formed at the spiral groove 5023. The heating cavity is adapted to accommodate the liquid pipeline, and both ends of the liquid pipeline are connected to a converter. The converter can connect the liquid pipeline of any one of the multiple heating devices 502 between the outlet end of the mixer and the inlet end of the reactor module.

[0065] A spiral heating cavity is formed between the heating core 5021 and the heat-insulating outer tube 5022 by a spiral groove 5023, and the liquid pipeline is coiled inside the heating cavity. The first temperature control component is used to heat the heating core 5021, which can transfer heat energy to the liquid pipeline inside the heating cavity to heat the liquid in the pipeline. The spiral groove 5023 can be located on the outer wall of the heating core 5021, forming a heating cavity between the spiral groove 5023 and the inner wall of the heat-insulating outer tube 5022; alternatively, it can be located on the inner wall of the heat-insulating outer tube 5022, forming a heating cavity between the spiral groove 5023 and the outer wall of the heating core 5021. Further, the outer wall of the heating core 5021 and the inner wall of the heat-insulating outer tube 5022 can each be provided with a spiral groove 5023, with the spiral grooves 5023 facing each other radially and fitting together to form a spiral channel accommodating the liquid pipeline, within which the heating cavity is formed.

[0066] By incorporating a heating core 5021 and an insulated outer tube 5022, a heating chamber isolated from the external environment can be formed, enabling safe heating of the liquid pipeline. The spiral groove 5023, designed to accommodate the liquid pipeline, increases the contact area between the liquid pipeline and the heating core 5021, effectively improving heat transfer efficiency. The first temperature control component enables precise temperature control and automated program control of the heating process, ensuring convenient operation. Furthermore, the spiral groove 5023 is highly adaptable to different specifications of liquid pipelines. This heating device 502 employs a spiral-wound mixing pipeline and a PID-controlled temperature heating core 5021 to enhance activation efficiency. Its overall structure is simple, compact, easy to disassemble, flexible in placement, and highly adaptable, making it suitable for applications such as continuous flow automated synthesis reactions.

[0067] In some embodiments, the heating core 5021 has protruding edges 50211 at both axial ends. The outer peripheral wall of the protruding edge 50211 is adapted to mate with the inner peripheral wall of the heat-insulating outer tube 5022. The protruding edge 50211 also has notches 50212 to allow liquid pipes to pass through, accommodating the two ends of the liquid pipes extending out of the heating cavity. In this embodiment, the protruding edge 50211 enables the mating connection between the heating core 5021 and the heat-insulating outer tube 5022. Simultaneously, the protruding edge 50211 creates a certain gap between the outer peripheral wall of the heating area on the heating core 5021 and the inner wall of the heat-insulating outer tube 5022. This gap, in conjunction with the spiral groove 5023, facilitates the accommodation of the liquid pipe. Furthermore, the notches 50212 on the protruding edge 50211 allow the two ends of the liquid pipe to extend out of the heating cavity for connection with external reaction devices, detection devices, or other systems.

[0068] In some embodiments, the heating device 502 further includes two plugs 5024, which are respectively disposed at both ends of the heat-insulating outer tube 5022 and detachably connected to the heat-insulating outer tube 5022 to form a closed heating space within the heat-insulating outer tube 5022. In this embodiment, by providing the plugs 5024, both ends of the heat-insulating outer tube 5022 can be sealed, and the two plugs 5024 and the heat-insulating outer tube 5022 define a closed heating space. This improves the heat insulation effect of the heating space and enhances the safety of the heating device 502, preventing operators from being burned by the heating core 5021.

[0069] In some embodiments, the plug 5024 has a through hole 50241 that allows a liquid conduit to pass through; both ends of the liquid conduit are adapted to extend out of the heating chamber and through the through hole 50241 to extend outside the heating space. In this embodiment, by providing a through hole 50241 in the plug 5024, a channel for the liquid conduit to enter and exit the heating space can be provided, so that both ends of the liquid conduit can be connected to external reaction devices, detection devices, or other systems.

[0070] In some embodiments, a limiting member is provided at the through hole 50241 of the plug 5024. The limiting member is suitable for limiting the liquid pipeline to improve the structural stability of the heating process. Specifically, the limiting member can be constructed as an elastic pipe clamp, a buckle, a spring, or the like, forming an adjustable clamping space at the through hole 50241 to clamp and limit liquid pipelines of different diameters.

[0071] In some embodiments, the two ends of the heat-insulating outer tube 5022 are formed with first threaded portions, and the inner or outer peripheral wall of the plug 5024 is formed with a second threaded portion that mates with the first threaded portion, so as to facilitate a detachable connection between the plug 5024 and the heat-insulating outer tube 5022 via threads. In this embodiment, the plug 5024 and the heat-insulating outer tube 5022 are detachably connected via threaded engagement, which is a simple, efficient, and easy-to-assemble / disassemble connection method. As shown in the figure, the two ends of the outer peripheral wall of the heat-insulating outer tube 5022 are formed with first threaded portions, and the inner peripheral wall of the plug 5024 is formed with a second threaded portion, with the first threaded portion and the second threaded portion engaging for connection.

[0072] In some embodiments, the heating device 502 further includes a limiting component adapted to selectively connect the heating core 5021 and the heat-insulating outer tube 5022, or selectively connect the plug 5024 and the heating core 5021, to restrict the movement of the heating core 5021 relative to the heat-insulating outer tube 5022. In this embodiment, by providing the limiting component, the relative movement of the heating core 5021 and the heat-insulating outer tube 5022 can be restricted to ensure high structural stability during the heating process.

[0073] In some embodiments, at least one plug 5024 has a first connecting portion, and the heating core 5021 has a second connecting portion. A limiting component can be selectively connected to the first and second connecting portions to restrict movement of the heating core 5021 relative to the plug 5024. In this embodiment, since the plug 5024 and the heat-insulating outer tube 5022 are stably connected, the heating core 5021, the heat-insulating outer tube 5022, and the plug 5024 can maintain structural stability by restricting the relative movement of the heating core 5021 and the plug 5024.

[0074] In some embodiments, the limiting component is configured as a limiting pin 5025, and at least one of the first connecting portion and the second connecting portion is configured as a pin hole 5026. The limiting pin 5025 and the pin hole 5026 cooperate to realize the limiting connection between the heating core 5021 and the plug 5024, which can restrict the relative circumferential rotation of the heating core 5021 within the heat-insulating outer tube 5022. The plug 5024 or the heating core 5021 can be fixedly connected to the limiting pin 5025 as a single unit.

[0075] In some embodiments, the first temperature control component includes at least one heating rod, and at least one mounting cavity 50213 extending axially along the heating core 5021 is formed within the heating core 5021, with the heating rod correspondingly disposed within each mounting cavity 50213. In this embodiment, heating the heating core 5021 by using heating rods results in high heating efficiency and uniform heating; simultaneously, the heating rods have a modular structure and are disposed within the mounting cavities 50213 of the heating core 5021, allowing for convenient installation, removal, and replacement.

[0076] In some embodiments, the first temperature control component further includes at least one temperature sensor. The heating core 5021 has at least one detection hole 50214, and the temperature sensor is correspondingly disposed within each detection hole 50214 to facilitate real-time detection of the temperature of the heating core 5021. In this embodiment, by setting a temperature sensor, the heating temperature of the heating core 5021 can be monitored in real time, allowing for adjustment of the heating rod's operating state based on the detection results, thus achieving precise temperature control. This embodiment establishes a real-time feedback mechanism by setting a temperature sensor, which helps to achieve automated temperature regulation, dynamic adjustment, and on-demand heating; it can also identify temperature anomalies, improving the safety of the device.

[0077] In some embodiments, the detection hole 50214 is opened near the spiral groove 5023 in order to obtain the heating temperature of the liquid pipeline more accurately and realistically.

[0078] In some embodiments, the control module controls the heating rod based on the detection results of the temperature sensor to achieve automated temperature regulation.

[0079] In some embodiments, the inner wall of the heat-insulating outer tube 5022 is clearance-fitted with the outer wall of the heating core 5021. In this embodiment, the clearance fit between the heating core 5021 and the heat-insulating outer tube 5022 facilitates the connection between the heating core 5021 and the heat-insulating outer tube 5022, making it easier to assemble and disassemble the device. Simultaneously, the heating core 5021 and the heat-insulating outer tube 5022 will change size due to thermal expansion at high temperatures; the clearance fit can accommodate this expansion, preventing structural damage caused by temperature changes.

[0080] In some embodiments, the spiral grooves 5023 are configured in multiple ways, with different pitches and / or groove depths. In this embodiment, by setting multiple spiral grooves 5023 with different pitches and / or groove depths, different specifications of liquid pipelines can be more accurately adapted, further improving the adaptability of the device; wherein, different types of liquid pipelines can be flexibly selected to adapt to various application scenarios.

[0081] In some embodiments, the heat-insulating outer tube 5022 may be made of polytetrafluoroethylene, which has good thermal insulation effect.

[0082] In specific use, the heating device 502 is used as follows: according to the actual situation such as the reaction scale, a corresponding liquid pipeline is placed in the spiral groove 5023 on the outer surface of the heating core 5021, the heat-insulating outer tube 5022 is fitted on, the plug 5024 is installed, and the heat-insulating outer tube 5022 is connected to the heating core 5021 for limiting; the first temperature control component is used to control the heating rod to heat to the required temperature, and the liquid reaction reagent is rapidly preheated after passing through the liquid pipeline.

[0083] The preheating module 5 integrates multiple heating devices 502 channels, with a maximum heating temperature of 90°C, which significantly improves the reaction rate and shortens the total coupling time of a single amino acid to 2 minutes.

[0084] In some embodiments, such as Figures 12 to 14 As shown, reactor 6 includes a base 610, a first fixed seat 630, a second fixed seat 640, a reaction tube, a cover 650, and a second temperature control assembly. A support assembly is mounted on the base 610; the first fixed seat 630 is mounted on the support assembly, and a first fluid channel 631 (i.e., the outlet end of reactor 6) is formed within the first fixed seat 630; the second fixed seat 640 is mounted on the first fixed seat 630 and detachably connected to it; a reaction chamber 641 and a second fluid channel 642 are formed within the second fixed seat 640, the reaction chamber 641 opening upwards, and the second fluid channel 642 communicating with both the reaction chamber 641 and the first fluid channel 631; the reaction tube is disposed within the reactor... Inside cavity 641, reaction tube is adapted to load solid carrier; and reaction tube has a fluid inlet and a fluid outlet, the fluid outlet being connected to a second fluid channel 642; cover 650 is disposed on second fixed seat 640 and detachably connected to second fixed seat 640; and cover 650 has a third fluid channel 651, the third fluid channel 651 being connected to fluid inlet (i.e., inlet end of reactor 6); second temperature control component is disposed on second fixed seat 640 and / or first fixed seat 630 to regulate the temperature inside reaction cavity 641.

[0085] During the reaction in reactor 6, a solid carrier is filled into the reaction tube, which is then placed in the reaction chamber 641. The first fluid channel 631, the second fluid channel 642, the fluid outlet of the reaction tube, the fluid inlet of the reaction tube, and the third fluid channel 651 are sequentially connected. Reactants are introduced into the third fluid channel 651, transported to the reaction tube, react with the solid carrier, and then flow out through the second fluid channel 642 and the first fluid channel 631, achieving a fluidized solid-phase synthesis reaction. Flow chemistry synthesis offers advantages such as robust reaction, high synthesis efficiency, high safety, and fast reaction speed. Furthermore, the second temperature control component can establish the required temperature conditions for the reaction. The base 610, the first fixed seat 630, the second fixed seat 640, the reaction tube, and the cover 650 are all detachable, making assembly simple and efficient, and facilitating the replacement of the reaction tube and internal solid carrier at any time. Simultaneously, the overall structure is robust, making it easy to place and use. The second mounting base 640 can be adapted to reaction tubes of different specifications via the reaction chamber 641. The size of the reaction tube can be flexibly adjusted according to requirements to adapt to different synthesis reaction scales.

[0086] Reactor 6 employs a detachable structure to establish a fluidized solid-phase synthesis reaction apparatus, suitable for automated continuous flow synthesis. It boasts high mass transfer efficiency and can improve reaction efficiency through continuous operation. It can accommodate solid-phase carriers of various sizes and types; precise temperature control is possible during the reaction; and its simple, compact structure facilitates easy assembly and disassembly, offering high flexibility and adaptability. Through modular design, layered sealing, and integrated temperature control, it solves the problems of poor flexibility and adaptability, low reaction efficiency, uneven temperature control, and easy leakage inherent in existing reactor 6 technologies. It can be widely applied in the solid-phase synthesis of peptides, oligonucleotides, and other related fields.

[0087] Furthermore, reactor 6 can be flexibly matched with automated systems to suit automated continuous flow synthesis reactions.

[0088] In some embodiments, the support assembly includes a plurality of struts 620, and the base 610 and the first fixed base 630 are connected by the plurality of struts 620.

[0089] In some embodiments, the first fluid channel 631 can be connected to a detection instrument or a waste liquid recovery container via a pipeline to facilitate the collection or detection of the liquid after the reaction. In some embodiments, the third fluid channel 651 can be connected to a pump structure via a pipeline to facilitate the pumping of reaction reagents into the reaction tube, thereby achieving automated flow synthesis reaction.

[0090] In some embodiments, such as Figure 12 , 13 As shown, the cover 650, the second fixed seat 640, the first fixed seat 630, the base 610 and the base 610 are arranged in sequence along the vertical direction. The base 610 and the support rod 620 can play a supporting role, so that the reactor 6 can be stably placed in the experimental space.

[0091] In some embodiments, the second temperature control component includes a heating rod and a temperature sensor. The heating rod and temperature sensor are connected to a control module. At least a portion of the heating rod is disposed within a second mounting base 640 to facilitate heating the reaction chamber 641. The temperature sensor is disposed within either a first mounting base 630 or a second mounting base 640 to detect the reaction temperature. In this embodiment, by using a heating rod, the reaction chamber 641 can be heated to achieve the required temperature conditions for the reaction. By using a temperature sensor, the reaction temperature can be detected to determine whether the temperature within the reaction chamber 641 has reached the required temperature, allowing for timely temperature adjustment via the heating rod to achieve precise temperature control and ensure the normal progress of the reaction. The temperature sensor can be a thermocouple temperature sensor. The control module controls the heating rod based on the data from the temperature sensor, achieving automated regulation of the reaction temperature.

[0092] In some embodiments, the first mounting base 630 has a first mounting channel 632, and the second mounting base 640 has a heating cavity 643, which communicates with the first mounting channel 632. At least a portion of the heating rod extends into the heating cavity 643 through the first mounting channel 632. In this embodiment, as... Figure 13 As shown, the first mounting channel 632 and the heating chamber 643 form a heating rod mounting cavity. By setting the first mounting channel 632 and the heating chamber 643, the heating rod is placed close to the reaction chamber 641, realizing the integrated design of the second temperature control component, the first fixing seat 630, and the second fixing seat 640, making the structure of the reactor 6 more compact.

[0093] In some embodiments, the heating rod is installed in the heating rod mounting cavity by a heating rod fixing assembly 670. The heating rod fixing assembly 670 specifically includes a pressure plate disposed on the bottom surface of the first fixing seat 630 and fastening bolts for fixing the pressure plate, wherein the pressure plate can limit the heating rod.

[0094] In some embodiments, at least two heating chambers 643 are configured and spaced apart around the outer periphery of the reaction chamber 641; multiple heating rods are configured to correspond one-to-one with each heating chamber 643. In this embodiment, as... Figure 13 As shown, the heating chamber 643 and the heating rod are constructed in at least two parts and are spaced apart around the outer periphery of the reaction chamber 641, which can improve the heating effect of the reaction chamber 641, make the heating efficiency higher, and make the heating effect of the reaction chamber 641 more uniform.

[0095] In some embodiments, the first mounting base 630 has a downwardly opening temperature sensor mounting cavity 633, and the temperature sensor is disposed within the temperature sensor mounting cavity 633. Further, the second mounting base 640 also has a temperature sensor detection cavity communicating with the temperature sensor mounting cavity 633, so that the temperature sensor can extend into the second mounting base 640, bringing the temperature sensor probe closer to the reaction cavity 641, thereby improving the reliability and accuracy of temperature detection.

[0096] In some embodiments, the reactor 6 further includes a first sieve plate 661 and a second sieve plate 662. The first sieve plate 661 is disposed on the cover 650 and is adapted to isolate the third fluid channel 651 and the fluid inlet; the second sieve plate 662 is disposed at the bottom end of the reaction tube to isolate the fluid outlet and the second fluid channel 642. In this embodiment, by setting the first sieve plate 661, impurities can be intercepted to prevent them from entering the reaction tube and ensure the smooth progress of the reaction; by setting the second sieve plate 662, the solid carrier can be prevented from flowing out, structural blockage can be avoided, and the liquid after the reaction can be collected more easily.

[0097] In some embodiments, reactor 6 further includes a first sealing ring and a second sealing ring. The first sealing ring is disposed between the first fixed seat 630 and the second fixed seat 640 to seal the connection between the first fluid channel 631 and the second fluid channel 642. The second sealing ring is disposed at the bottom end of the reaction tube to seal the connection between the fluid outlet and the second fluid channel 642. In this embodiment, by providing the first and second sealing rings, the sealing effect of reactor 6 can be improved, preventing reactant leakage.

[0098] Furthermore, a first sealing ring mounting groove 645 is formed on the bottom surface of the second fixed seat 640 that abuts against the first fixed seat 630. The first sealing ring is disposed in the first sealing ring mounting groove 645. When the first fixed seat 630 and the second fixed seat 640 are assembled, the first sealing ring is squeezed to achieve a sealing effect. A second sealing ring mounting groove 646 is formed on the inner wall of the second fluid channel 642 or the bottom peripheral wall of the reaction tube. The second sealing ring is disposed in the second sealing ring mounting groove 646. When the reaction tube is placed into the reaction chamber 641, at least part of the bottom end of the reaction tube extends into the second fluid channel 642. The bottom end of the reaction tube squeezes the second sealing ring to achieve a sealing effect.

[0099] In some embodiments, the cover 650 has a flow guide 652 that extends into the reaction tube; a portion of the third fluid channel 651 is formed in the flow guide 652 to facilitate communication with a fluid inlet. In this embodiment, by providing the flow guide 652 and extending it into the reaction tube after structural assembly, the fluid delivery path can be optimized, ensuring that the reactants are smoothly delivered from the third fluid channel 651 into the reaction tube, while also preventing leakage of reactants in the gap between the cover 650 and the second fixing seat 640.

[0100] In some embodiments, the reactor 6 further includes a third sealing ring disposed on the outer periphery of the flow guide 652 to form a seal between the flow guide 652 and the reaction tube. In this embodiment, the third sealing ring improves the sealing effect between the cover 650 and the reaction tube, preventing reactants from leaking from the gap between the flow guide 652 and the inner wall of the reaction tube. Further, in some embodiments, a third sealing ring mounting groove 653 is formed on the outer periphery of the flow guide 652. The third sealing ring is disposed in the third sealing ring mounting groove 653 and, during assembly of the reaction tube, the second fixing seat 640, and the cover 650, is squeezed by the inner wall of the reaction tube to achieve a sealing effect.

[0101] In the above embodiments, by setting the first sealing ring, the second sealing ring, and the third sealing ring, the cover 650, the second fixing seat 640, and the first fixing seat 630 can form a leak-proof structure.

[0102] In some embodiments, the second fixing seat 640 forms a limiting groove 644, and the reaction tube has a protruding edge that engages with the limiting groove 644 to limit the position of the reaction tube. In this embodiment, by providing a limiting groove 644 in the second fixing seat 640, an engagement with the reaction tube is achieved to limit the position of the reaction tube, thereby ensuring that reaction tubes of different specifications can be stably assembled in the reaction chamber 641.

[0103] In some embodiments, the reactor 6 further includes a connecting assembly adapted to connect the first fixing seat 630, the second fixing seat 640, and the cover 650. In this embodiment, by providing a connecting assembly to connect the first fixing seat 630, the second fixing seat 640, and the cover 650, the stability of the structural assembly can be improved, while also enhancing the sealing effect of the structure. Specifically, in some embodiments, such as Figure 12 As shown, the first fixing seat 630 has at least one first fastening hole 634, and the second fixing seat 640 has at least one second fastening hole 647 on one side. The first fastening hole 634 and the second fastening hole 647 are threaded holes, and are tightly connected by fasteners such as bolts. The other side of the second fixing seat 640 has at least one third fastening hole, and the cover 650 has at least one fourth fastening hole. The third fastening hole and the fourth fastening hole are threaded holes, and are tightly connected by fasteners such as bolts. In this embodiment, the various structures are connected by threaded fasteners, which allows for quick assembly and disassembly.

[0104] In practical applications, reactor 6 involves placing corresponding reaction tubes in reaction chamber 641 based on actual conditions such as reaction scale and liquid flow rate; loading corresponding solid support into the reaction tubes; covering with cover 650; and securing cover 650, second fixing seat 640, and first fixing seat 630 with fasteners; using a second temperature control component to control the heating rod to the required temperature; and allowing the liquid reaction reagent to flow from top to bottom through the reaction tubes, reacting with the solid support within, and then flowing out from the first fluid channel 631 of the first fixing seat 630. This reactor 6 can achieve synthesis scales of 0.001~0.5 mmol, making it more widely applicable.

[0105] In some embodiments, such as Figure 6 As shown, it also includes a gas path module 7; the gas path module 7 is connected to the liquid storage module 1 and is used to input protective gas into the liquid storage module 1 to protect each liquid reaction reagent.

[0106] Specifically, the gas path module 7 mainly consists of a gas source 701, a pressure regulating valve 702, multiple gas pipes, multiple multi-port valves 703, and multiple rotor flow meters 704; the pressure regulating valve 702 and rotor flow meters 704 operate under the control of the control module. The purpose of setting up the gas path module 7 is to fill each reagent bottle 101 of the liquid storage module 1 with inert gas, such as nitrogen or argon, to protect the liquid reaction reagents, such as amino acids, from oxidation or water absorption and precipitation.

[0107] In some embodiments, the system further includes an ultraviolet (UV) monitoring module 8, a back pressure valve 9, and a waste liquid tank 10. The UV monitoring module 8 is connected to the outlet of the reactor 6, and the UV monitoring module 8, back pressure valve 9, and waste liquid tank 10 are connected in series. The UV monitoring module 8 is responsible for real-time online monitoring of peptide synthesis efficiency; the back pressure valve 9 regulates the overall pipeline pressure of the fully automated solid-phase peptide synthesis system based on flow chemistry to ensure pressure stability; and the waste liquid tank 10 collects the waste liquid flowing out from the outlet of the reactor 6.

[0108] Equipped with an ultraviolet monitoring module 8, it can monitor the efficiency of amino acid condensation and deprotection reactions in real time, enabling precise control of the synthesis process.

[0109] The following examples illustrate in detail the types of reagents and synthesis processes of the fully automated solid-phase peptide synthesis system based on flow chemistry of the present invention.

[0110] The types of reaction reagents are as follows: Firstly, regarding amino acids: Currently, there are 20 natural amino acids and various non-natural amino acids, all of which use Fmoc to protect their N-termini. Therefore, in addition to reagent bottles 101 containing the 20 natural amino acids, the liquid storage module 1 also reserves at least 10 reagent bottles 101 for non-natural amino acids.

[0111] Second, deprotection reagents: the currently commonly used reagent is piperidine.

[0112] Thirdly, activators: PyAOP, HATU, etc., can be selected flexibly according to needs.

[0113] Fourthly, alkali: here it is DIEA. It is connected separately to a volumetric pump, namely the first volumetric pump 2011.

[0114] Fifth, the primary solvent: here it is DMF. DMF is both a solvent for dissolving amino acids and a solvent for dissolving activators. As the primary solvent, a DMF line is connected to each selection for cleaning the corresponding pipeline.

[0115] In the liquid pipeline, the reagents are arranged according to the following overall scheme: a separate alkali transport component 201, a second transport component 202 for activators and deprotection reagents, and a third transport component 203 for amino acid reagents.

[0116] The synthesis process is as follows: The entire solid-phase peptide synthesis process is divided into four stages: deprotection reaction, deprotection flow path cleaning, condensation reaction, and condensation flow path cleaning.

[0117] Deprotection reaction: Piperidine is used to deprotect the Fmoc groups on the solid resin, exposing the amine reactive groups. At this time, the second volumetric pump 2022 is turned on, and the first selection valve 2021 is switched to open the piperidine flow path. Piperidine flows from the piperidine reagent bottle 101 to the first selection valve 2021, then to the second volumetric pump 2022, through the four-way valve 3, the mixer 4, and the converter 501. It is heated in the preheating module 5 (usually at 90 degrees Celsius), and then flows to the reactor 6 for the deprotection reaction. Finally, it flows through the flow cell at the ultraviolet monitoring module 8 and the back pressure valve 9 to the waste liquid tank 10.

[0118] Cleaning the protection flow path: The pipeline is cleaned using the main solvent (DMF). At this time, the second volumetric pump 2022 is turned on, and the first selector valve 2021 is switched to open the DMF flow path. The DMF flows through the first selector valve 2021 to the second volumetric pump 2022, through the four-way valve 3, the mixer 4, and through the converter 501. It is heated in the preheating module 5 (usually at 90 degrees Celsius), then flows to the reactor 6, and finally flows through the flow cell at the ultraviolet monitoring module 8, the back pressure valve 9, and to the waste liquid tank 10.

[0119] Condensation reaction: Solid-phase condensation of amino acids takes place. At this time, the first volumetric pump 2011, the second volumetric pump 2022, and the third volumetric pump 2032 are all open. The second volumetric pump 2022 is connected to the first selector valve 2021 to switch to the activator flow path (e.g., HATU or PyAOP). The four second selector valves 2031 connected to the third volumetric pump 2032 switch the flow path according to the corresponding amino acid position. The first volumetric pump 2011 is connected to the alkali DIEA. The three liquids are transported by the first volumetric pump 2011, the second volumetric pump 2022, and the third volumetric pump 2032, simultaneously reaching the four-way valve 3 position. After mixing by the mixer 4, the liquid flows to the converter 501, where it is switched to the corresponding preheating module 5 to heat to the corresponding temperature as needed. Then it flows to the reactor 6, where the activated amino acids undergo a condensation reaction with the amine groups on the resin, thus attaching to the solid-phase resin. The waste liquid flows out from the outlet of the reactor 6 and continues to flow through the flow cell at the ultraviolet monitoring module 8, the back pressure valve 9, and then to the waste liquid tank 10.

[0120] Condensation reaction flow path cleaning: The pipeline is cleaned using the main solvent (DMF). At this time, the second volumetric pump 2022 and the third volumetric pump 2032 are opened, and the first selector valve 2021 switches to the DMF flow path. DMF flows through the first selector valve 2021 to the second volumetric pump 2022, and then to the four-way valve 3. The four second selector valves 2031 connected to the third volumetric pump 2032 switch the flow path to the DMF reagent bottle 101 according to the corresponding position of the previously extracted amino acid. DMF flows through the corresponding second selector valve 2031 to the four-way valve 3. The two DMF streams converge at the four-way valve 3, flow through the mixer 4 and converter 501, are heated in the preheating module 5 (usually to 90 degrees Celsius), then flow to the reactor 6, and finally flow through the flow cell at the ultraviolet monitoring module 8 and the back pressure valve 9 to the waste tank 10.

[0121] Each amino acid thereafter is subjected to a cycle of deprotection reaction, deprotection flow path cleaning, condensation reaction, and condensation reaction cleaning.

[0122] It is particularly important to emphasize that in the fully automated solid-phase peptide synthesis system based on flow chemistry of this invention, reactor 6 can achieve a synthesis scale of 0.001~0.5 mmol, making it more widely applicable. The preheating module 5 integrates multiple heating devices 502 channels, with a maximum heating temperature of 100℃, significantly improving the reaction rate and reducing the total coupling time of a single amino acid to 2 minutes. The solvent system used can adapt to a wider temperature range and is suitable for high-temperature synthesis reactions. Equipped with an ultraviolet monitoring module 8, it can monitor the efficiency of amino acid condensation and deprotection reactions in real time, achieving precise control of the synthesis process. All reactions are completed within a single reactor 6, eliminating the need for resin transfer, greatly simplifying the operation process. Liquid reaction reagent delivery is achieved through the coordinated control of selector valves and volumetric pumps, requiring only 3 volumetric pumps, significantly reducing equipment costs and space requirements. It can be configured with at least 30 reagent bottles 101, comprehensively covering 20 natural amino acids and accommodating no less than 10 non-natural amino acids, greatly expanding the diversity of peptide synthesis. Amino acids and activators are stored separately, allowing for in-situ activation of amino acids during synthesis via preheating module 5. This avoids degradation of liquid reaction reagents and allows for long-term storage. All amino acid reagents are under inert gas protection, which not only improves reagent stability but also avoids the risk of pipeline blockage.

[0123] This invention also proposes a fully automated solid-phase peptide synthesis method based on flow chemistry.

[0124] According to the fully automated solid-phase peptide synthesis method based on flow chemistry of the present invention, solid-phase peptide synthesis is performed using the fully automated solid-phase peptide synthesis system based on flow chemistry of the present invention described above, and includes the following steps (see...). Figures 2 to 5 and Figure 15 (as shown) S1: Deprotection reaction: The deprotection agent in the storage module 1 is selected and precisely extracted through the transfer module 2, and the extracted deprotection agent flows sequentially through the transfer module 2, mixer 4, preheating module 5 and reactor 6. The deprotection agent deprotects the Fmoc groups on the solid resin in reactor 6, exposing the amine reactive groups. S2: Deprotection flow path cleaning: The main solvent in the storage module 1 is selected and precisely extracted through the transfer module 2 to rinse the deprotection reaction flow path in step S1. S3: Condensation reaction: Typically, the transfer module 2 simultaneously selects and precisely extracts the activator, alkali, and corresponding amino acids from the storage module 1, allowing the activator, alkali, and corresponding amino acids to flow sequentially through the mixer 4 and mix rapidly and thoroughly. When the mixed liquid reaction reagent flows through the preheating module 5, it is preheated and activated to form a preheated liquid reaction reagent. When the preheated liquid reaction reagent flows through the reactor 6, a condensation reaction occurs. The condensation reaction is a condensation reaction between the activated amino acids and the amine reactive groups on the resin in the reactor 6. S4: Condensation reaction flow path cleaning: The main solvent in the storage module 1 is selected and precisely extracted through the transfer module 2 to rinse the condensation reaction flow path in step S3. S5: Repeat steps S1 to S4 until all amino acids are spliced ​​together.

[0125] In other words, solid-phase peptide synthesis involves sequentially coupling the corresponding types of amino acids onto the solid-phase resin in reactor 6 according to the target peptide sequence. The coupling of each type of amino acid is carried out in a cycle of "deprotection reaction - deprotection flow path cleaning - condensation reaction - condensation reaction flow path cleaning". One cycle completes the coupling of one type of amino acid. The cycle of "deprotection reaction - deprotection flow path cleaning - condensation reaction - condensation reaction flow path cleaning" is repeated until all amino acids are coupled.

[0126] Since the fully automated solid-phase peptide synthesis method based on flow chemistry in this embodiment of the invention uses the fully automated solid-phase peptide synthesis system based on flow chemistry in the aforementioned embodiment of the invention, the fully automated solid-phase peptide synthesis method based on flow chemistry in this embodiment of the invention has essentially the same technical effects as the fully automated solid-phase peptide synthesis system based on flow chemistry in the aforementioned embodiment of the invention, and will not be repeated here.

[0127] In some embodiments, prior to step S1, the peptide synthesis system needs to be initialized. Initialization includes testing and initialization of the selection valves (i.e., the first selection valve 2021 and the second selection valve 2031), flow rate calibration of the pumps (i.e., the first volumetric pump 2011, the second volumetric pump 2022, and the third volumetric pump 2032), calibration and initialization of the ultraviolet monitoring module 8, temperature calibration and initialization, check valve testing, and mixing test of the mixer 4. The flow rate calibration for the pumps has a relative standard error of 5%; the ultraviolet monitoring absorbance calibration parameters include background, acquisition time, and integrated intensity; the temperature calibration error is ±1℃.

[0128] In some embodiments, the fully automated solid-phase peptide synthesis method based on flow chemistry specifically includes the following steps: (1) Add different liquid reaction reagents to the corresponding reagent bottles 101.

[0129] (2) Turn on the protective gas source 701 and adjust the protective gas pressure to 0.1-0.15 MPa.

[0130] (3) Turn on the power of the polypeptide synthesis system.

[0131] (4) Turn on the computer.

[0132] (5) Turn on the pump switch.

[0133] (6) Turn on the switch of the preheating module 5.

[0134] (7) Open the operating software.

[0135] (8) Initialization of the polypeptide synthesis system.

[0136] (9) Pressure self-test of polypeptide synthesis system.

[0137] (10) Run each volumetric pump to clean the pipeline. (11) Users can customize and save the target polypeptide sequence, synthesis program, execution program, or directly open the pre-defined method program file.

[0138] (12) Open reactor 6 and add solid resin to reactor 6.

[0139] (13) Execute the solid-phase polypeptide synthesis program to start the synthesis.

[0140] (14) After the synthesis is completed, run each volume pump to clean the pipeline.

[0141] (15) Shut down the solid-phase polypeptide synthesis system.

[0142] (16) Open reactor 6 and remove the resin.

[0143] Example The following examples illustrate the synthesis of polypeptides ACP65-74 (10 amino acids), ABC-20 (20 amino acids), and HD6 (32 amino acids).

[0144] First, the fully automated solid-phase peptide synthesis system based on flow chemistry was self-tested to confirm that it could operate normally.

[0145] For peptide ACP65-74, enter the synthetic peptide sequence “VQAAIDYING” in the control module; for peptide ABC-20, enter the synthetic peptide sequence “VYWTSPFMKLIHEQCNRADG” in the control module; for peptide HD-6, enter the synthetic peptide sequence “AFTGHGRRSGYSTEYSYGTGTVMGINHRFGGL” in the control module.

[0146] Flow rate settings: First volumetric pump 2011 is set at 5 mL / min, second volumetric pump 2022 and third volumetric pump 2032 are set at 40 mL / min. Preheating temperature is 90 degrees Celsius, and reactor 6 heating temperature is 90 degrees Celsius.

[0147] The total time for each amino acid synthesis cycle was set to 2 minutes, including 30 seconds for the deprotection reaction, 30 seconds for cleaning the deprotection flow path, 30 seconds for the condensation reaction, and 30 seconds for cleaning the condensation reaction flow path.

[0148] 300 mg of Rink-Amide resin with a substitution degree of 0.5 mmol / g was loaded into reactor 6, and the solid-phase peptide synthesis system was started to run the peptide synthesis step. The deprotection process of the synthesis was monitored in real time using UV monitoring module 8 (311 nm wavelength).

[0149] After completing the peptide synthesis, the resin was removed from reactor 6 and the peptide was cleaved using a trifluoroacetic acid cleavage reagent. The peptide was then precipitated with diethyl ether and dried to obtain crude peptide powder.

[0150] The crude peptide powder was dissolved in a water / acetonitrile mixed solvent and analyzed by reversed-phase high-performance liquid chromatography.

[0151] For ACP65-74, by Figure 16 The liquid chromatography shows a single main peak with a retention time of 17.5 minutes. High-resolution mass spectrometry analysis of the collected eluent from this main peak indicates a molecular weight of 1062 Da, which is consistent with the theoretical molecular weight of ACP65-74 (see [reference]). Figure 17 This proves that the polypeptide was successfully synthesized in high purity.

[0152] For ABC-20, by Figure 18As can be seen, the liquid chromatography output shows a single main peak with a retention time of 25.3 minutes. High-resolution mass spectrometry analysis of the collected eluent from this main peak indicated a molecular weight of 2394 Da, which is consistent with the theoretical molecular weight of ABC-20 (see [reference needed]). Figure 19 This proves that the polypeptide was successfully synthesized with high purity. Figure 20 As can be seen, the height of the UV absorption peak did not change significantly during the synthesis process. Furthermore, integrating the peak areas shows no significant decrease in peak area. This indicates that the synthesis proceeded stably.

[0153] For HD-6, by Figure 21 As can be seen, the liquid chromatography output shows a single main peak with a retention time of 16.5 minutes. High-resolution mass spectrometry analysis of the collected eluent from this main peak indicates a molecular weight of 3436 Da, which is consistent with the theoretical molecular weight of HD-6 (see [reference needed]). Figure 22 This proves that the polypeptide was successfully synthesized in high purity.

[0154] Taking ABC-20 as an example, the entire synthesis of a single amino acid takes only 2 minutes, with a total time of only 40 minutes. In contrast, the Libertyblue 2.0, one of the fastest commercially available peptide synthesizers, takes approximately 120 minutes to synthesize ABC-20. Furthermore, the synthesis of HD-6 using this invention takes only 64 minutes, while the Libertyblue 2.0 takes approximately 180 minutes. Therefore, the fully automated solid-phase peptide synthesis system and method based on flow chemistry of this invention significantly reduces peptide synthesis time while ensuring the accuracy and purity of the synthesized peptides.

[0155] Finally, please note that the meanings of the English words or abbreviations used in this article can be found in [link to article]. Figure 23 The table content.

[0156] Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those skilled in the art can make changes, modifications, substitutions, and variations to the above embodiments within the scope of the present invention.

Claims

1. A fully automated solid-phase peptide synthesis system based on flow chemistry, characterized in that, It includes a liquid storage module, a transfer module, a mixer, a preheating module, a reactor, and a control module; The liquid storage module is used to store different liquid reaction reagents required for solid-phase peptide synthesis, including deprotection reagents, main solvents, activators, bases, and amino acids. The transmission module is connected between the liquid storage module and the mixer, and is used to select and precisely extract the corresponding liquid reaction reagents from the liquid storage module to the mixer according to different stages of solid-phase peptide synthesis, so that the corresponding liquid reaction reagents are quickly and thoroughly mixed when flowing through the mixer. The preheating module is connected between the mixer and the reactor. The preheating module preheats the mixed liquid reaction reagent from the mixer so that the preheated liquid reaction reagent completes the corresponding stage of solid-phase peptide synthesis when it flows through the reactor. The control module controls the operation of the fully automated solid-phase peptide synthesis system based on flow chemistry.

2. The fully automated solid-phase peptide synthesis system based on flow chemistry according to claim 1, characterized in that, The transmission module includes a first transmission component, a second transmission component, and a third transmission component; The first transmission component is connected between the liquid storage module and the inlet of the mixer, and is used to accurately extract alkali from the liquid storage module to the mixer; The second transmission component is connected between the reservoir module and the inlet of the mixer, and is used to select and precisely extract the deprotection reagent, main solvent and activator from the reservoir module to the mixer; The third transmission component is connected between the liquid storage module and the inlet of the mixer, and is used to select and precisely extract amino acids and main solvent from the liquid storage module to the mixer.

3. The fully automated solid-phase peptide synthesis system based on flow chemistry according to claim 2, characterized in that, The liquid storage module includes reagent bottles for storing different types of liquid reaction reagents, wherein at least 20 of the reagent bottles are used to store at least 20 kinds of amino acids; and at least 2 of the reagent bottles are used to store at least 2 kinds of activators. The first transfer component includes a first volumetric pump, the inlet of which is connected to the reagent bottle storing the alkali, and the outlet of which is connected to the inlet of the mixer. The second transfer component includes a first selection valve and a second volumetric pump. The inlet of the second volumetric pump is connected to the reagent bottle storing the deprotection reagent, the reagent bottle storing the main solvent, and the reagent bottle storing the activator through the first selection valve. The outlet of the second volumetric pump is connected to the inlet of the mixer. The third transfer component includes a second selection valve and a third volumetric pump. There is at least one second selection valve. The inlet of the third volumetric pump is connected to the reagent bottle storing the main solvent and the reagent bottle storing amino acids respectively through at least one of the second selection valves. The outlet of the third volumetric pump is connected to the inlet of the mixer.

4. The fully automated solid-phase peptide synthesis system based on flow chemistry according to claim 3, characterized in that, The multiple inlet ends of the first selection valve are respectively connected to the multiple reagent bottles corresponding to storing multiple activators, the reagent bottle storing deprotection reagents, and the reagent bottle storing the main solvent. The outlet end of the first selection valve is connected to the inlet end of the second volumetric pump, and the outlet end of the second volumetric pump is connected to the inlet end of the mixer. There are multiple second selection valves. The multiple inlet ends of one second selection valve are respectively connected to the outlet ends of the remaining second selection valves. The outlet end of one second selection valve is connected to the inlet end of the third volumetric pump. The outlet end of the third volumetric pump is connected to the inlet end of the mixer. The multiple outlet ends of the remaining second selection valves are respectively connected to the reagent bottle storing different types of amino acids and the reagent bottle storing the main solvent.

5. The fully automated solid-phase peptide synthesis system based on flow chemistry according to claim 1, characterized in that, The mixer is a spiral column structure consisting of a liquid tube wound around a cylindrical rod.

6. The fully automated solid-phase peptide synthesis system based on flow chemistry according to claim 1, characterized in that, The preheating module includes a converter and multiple heating devices connected in parallel to the converter. The converter is connected between the mixer and the reactor, and the preheating temperatures of the multiple heating devices are different.

7. The fully automated solid-phase peptide synthesis system based on flow chemistry according to claim 6, characterized in that, The heating device includes: A heating element, wherein a first temperature control component is provided inside the heating element, and the first temperature control component is adapted to heat the heating element; A heat-insulating outer tube is sleeved around the outer periphery of the heating core; wherein... At least one of the heating core and the heat-insulating outer tube has a spiral groove; after the heating core is sleeved with the heat-insulating outer tube, a heating cavity is formed at the spiral groove; the heating cavity is suitable for accommodating liquid pipelines.

8. The fully automated solid-phase peptide synthesis system based on flow chemistry according to claim 1, characterized in that, The reactor includes: A base on which a support assembly is provided; A first fixing seat is disposed on the bracket assembly, and a first fluid channel is formed inside the first fixing seat; A second fixing seat is disposed on the first fixing seat and is detachably connected to the first fixing seat; a reaction chamber and a second fluid channel are formed inside the second fixing seat, the reaction chamber is open to the upward, and the second fluid channel communicates with the reaction chamber and the first fluid channel. A reaction tube is disposed within the reaction chamber and is adapted to load a solid carrier; the reaction tube has a fluid inlet and a fluid outlet, and the fluid outlet is connected to the second fluid channel. A cover body is disposed on the second fixed base and is detachably connected to the second fixed base; and a third fluid channel is formed in the cover body, the third fluid channel being connected to the fluid inlet. A second temperature control component is disposed on the first fixed base and / or the second fixed base to regulate the temperature inside the reaction chamber.

9. The fully automated solid-phase peptide synthesis system based on flow chemistry according to claim 1, characterized in that, It also includes a gas path module; the gas path module is connected to the liquid storage module and is used to input protective gas into the liquid storage module to protect each liquid reaction reagent.

10. A fully automated solid-phase peptide synthesis method based on flow chemistry, characterized in that, Solid-phase peptide synthesis using the fully automated solid-phase peptide synthesis system based on flow chemistry as described in any one of claims 1 to 9 includes the following steps: S1: Deprotection reaction: The deprotection agent in the storage module is selected and precisely extracted through the transfer module, and the extracted deprotection agent flows sequentially through the transfer module, the mixer, the preheating module and the reactor. The deprotection agent deprotects the Fmoc groups on the solid resin in the reactor, exposing the amine reactive groups. S2: Deprotection flow path cleaning: The main solvent in the storage module is selected and precisely extracted through the transfer module to rinse the deprotection reaction flow path in step S1; S3: Condensation reaction: The activator, alkali, and corresponding amino acids in the storage module are simultaneously and precisely selected and extracted through the transmission module, so that the activator, alkali, and corresponding amino acids flow through the mixer in sequence and are quickly and thoroughly mixed. When the mixed liquid reaction reagent flows through the preheating module, it is preheated and activated to form a preheated liquid reaction reagent. When the preheated liquid reaction reagent flows through the reactor, a condensation reaction occurs. The condensation reaction is a condensation reaction between the activated amino acids and the amine reactive groups on the resin in the reactor. S4: Condensation reaction flow path cleaning: The main solvent in the storage module is selected and precisely extracted through the transfer module to flush the condensation reaction flow path in step S3. S5: Repeat steps S1 to S4 in a loop until all amino acid coupling is completed.

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