Preparation method of N-acetyl-5-bromo-chloro-3-indolone, N-acetyl-5-bromo-chloro-3-indolone, 5-bromo-chloro-3-indoloside and chromogenic culture medium

Abstract

This application relates to the field of organic synthesis technology, and discloses a method for preparing N-acetyl-5-bromo-chloro-3-indolone. The method comprises: under alkaline conditions, 5-bromo-chloroindole undergoes an oxidation reaction to introduce an acetoxy group at the 3-position of the indole ring, yielding 5-bromo-chloro-3-acetoxyindole; wherein the chlorine atom is located on the benzene ring of the indole ring; 5-bromo-chloro-3-acetoxyindole undergoes an N-acetylation reaction to introduce an acetyl group at the nitrogen atom at the 1-position of the indole ring, yielding N-acetyl-5-bromo-chloro-3-acetoxyindole; under acidic conditions, N-acetyl-5-bromo-chloro-3-acetoxyindole undergoes a selective hydrolysis reaction to remove the acetyl group from the oxygen atom, yielding N-acetyl-5-bromo-chloro-3-indolone. This improves the safety and efficiency of the preparation process. This application also discloses an N-acetyl-5-bromo-chloro-3-indolone, a 5-bromo-chloro-3-indoloside, and a colorimetric culture medium.

Description

Technical Field

[0001] This application relates to the field of organic synthesis technology, for example to a method for preparing N-acetyl-5-bromo-chloro-3-indolone, N-acetyl-5-bromo-chloro-3-indolone, 5-bromo-chloro-3-indoloside, and a colorimetric culture medium. Background Technology

[0002] Chromogenic media are isolation media that achieve the detection of specific microorganisms through a colorimetric reaction between microbial metabolic enzymes and chromogenic substrates. Currently, 5-bromo-4-chloro-3-indole glycosides and 5-bromo-6-chloro-3-indole glycosides are widely used as chromogenic substrates for the detection of microorganisms such as *Escherichia coli*, coliform bacteria, *Escherichia coli* O157, *Staphylococcus aureus*, and *Cronobacter sakazakii*. However, existing methods for preparing 5-bromo-4-chloro-3-indole glycosides and 5-bromo-6-chloro-3-indole glycosides generally suffer from problems such as long reaction steps, low yields, and cumbersome operations. Some processes also require the use of highly toxic reagents such as allyl alcohol and allyl bromoacetate, posing significant safety risks. Therefore, developing synthetic routes that are short in steps, simple in operation, and highly safe is of great importance. Using N-acetyl-5-bromo-4-chloro-3-indoleone and N-acetyl-5-bromo-6-chloro-3-indoleone as intermediates, 5-bromo-4-chloro-3-indole glycoside and 5-bromo-6-chloro-3-indole glycoside can be efficiently prepared through glycosylation and deprotection reactions.

[0003] The relevant technology uses 2-[(carboxymethyl)amino]-5-bromo-4-chlorobenzoic acid as the starting material, and constructs an indole heterocyclic core through an intramolecular cyclization-diacetization tandem reaction in an acetic anhydride / sodium acetate system to obtain N-acetyl-5-bromo-6-chloro-3-acetoxyindole; then, the intermediate is placed in an aqueous sulfuric acid solution and subjected to acetoxy hydrolysis and enol-ketone tautomerism rearrangement to obtain N-acetyl-5-bromo-6-chloro-3-indoleone.

[0004] In the process of implementing the embodiments of this disclosure, at least the following problems were found in the related art: Among the related technologies, the preparation route of N-acetyl-5-bromo-6-chloro-3-indolone requires a ring-condensation process, resulting in low overall reaction safety.

[0005] It should be noted that the information disclosed in the background section above is only used to enhance the understanding of the background of this application, and therefore may include information that does not constitute prior art known to those skilled in the art. Summary of the Invention

[0006] To provide a basic understanding of some aspects of the disclosed embodiments, a brief summary is given below. This summary is not intended as a general commentary, nor is it intended to identify key / important components or describe the scope of protection of these embodiments, but rather as a prelude to the detailed description that follows.

[0007] This disclosure provides a method for preparing N-acetyl-5-bromo-chloro-3-indolone, N-acetyl-5-bromo-chloro-3-indolone, 5-bromo-chloro-3-indoloside, and a colorimetric culture medium to improve the safety of the preparation process, shorten the preparation steps, and increase the yield and preparation efficiency.

[0008] In some embodiments, the preparation method of N-acetyl-5-bromo-chloro-3-indolone includes: under alkaline conditions, 5-bromo-chloroindole undergoes an oxidation reaction to introduce an acetoxy group at the 3-position of the indole ring to obtain 5-bromo-chloro-3-acetoxyindole; wherein the chlorine atom is located on the benzene ring of the indole ring; 5-bromo-chloro-3-acetoxyindole undergoes an N-acetylation reaction to introduce an acetyl group at the nitrogen atom at the 1-position of the indole ring to obtain N-acetyl-5-bromo-chloro-3-acetoxyindole; under acidic conditions, N-acetyl-5-bromo-chloro-3-acetoxyindole undergoes a selective hydrolysis reaction to remove the acetyl group on the oxygen atom to obtain N-acetyl-5-bromo-chloro-3-indolone.

[0009] In some embodiments, the N-acetyl-5-bromo-chloro-3-indolone is prepared using the preparation method described in any of the foregoing disclosed embodiments.

[0010] In some embodiments, the 5-bromo-chloro-3-indole glycoside is prepared by glycosylation and deprotection reaction using N-acetyl-5-bromo-chloro-3-indole ketone as described in the foregoing disclosed embodiments.

[0011] In some embodiments, the chromogenic culture medium uses 5-bromo-chloro-3-indoleglycoside as the chromogenic substrate as described in the foregoing disclosed embodiments.

[0012] The preparation method of N-acetyl-5-bromo-chloro-3-indolone, N-acetyl-5-bromo-chloro-3-indolone, 5-bromo-chloro-3-indoloside, and chromogenic culture medium provided in this disclosure can achieve the following technical effects: Starting with 5-bromo-chloroindole, the target product is directly obtained through a three-step process of oxidative acetylation, N-acetylation, and selective hydrolysis. Compared with existing technologies, this method avoids the two-step process of first preparing 3-hydroxyindole and then esterifying it, as well as the ring-condensation process involved in the Bischler-Möhlau type intramolecular cyclization-diacetization tandem reaction. The overall reaction steps are significantly shortened, and the operation is simpler. Furthermore, this method avoids the safety hazards associated with the ring-condensation process from the outset and avoids the use of highly toxic reagents such as allyl alcohol and allyl bromoacetate. The reaction conditions are mild, effectively improving production safety. In addition, each step of this method yields a high yield, improving the overall yield and production efficiency, thus enabling the efficient preparation of N-acetyl-5-bromo-chloro-3-indole ketone, providing a high-quality intermediate for the preparation of 5-bromo-chloro-3-indole glycosides and chromogenic culture media.

[0013] 5-Bromo-chloro-3-indole glycoside can be used as a highly sensitive chromogenic substrate, enabling rapid visual identification of bacterial colonies with low background interference, and is suitable for rapid detection of pathogenic microorganisms such as Escherichia coli and Staphylococcus aureus.

[0014] The above general description and the description below are exemplary and illustrative only and are not intended to limit this application. Attached Figure Description

[0015] One or more embodiments are illustrated by way of example with reference to the accompanying drawings. These illustrations and drawings do not constitute a limitation on the embodiments. Elements having the same reference numerals in the drawings are shown as similar elements. The drawings are not to be scaled. And wherein: Figure 1 This is a flowchart illustrating a method for preparing N-acetyl-5-bromo-chloro-3-indolone according to an embodiment of this disclosure; Figure 2 This is a synthetic route diagram of a method for preparing N-acetyl-5-bromo-chloro-3-indolone provided in the embodiments of this disclosure; Figure 3 This is a synthetic route diagram for the preparation of N-acetyl-5-bromo-4-chloro-3-indolone provided in the embodiments of this disclosure; Figure 4 This is a synthetic route diagram for the preparation of N-acetyl-5-bromo-6-chloro-3-indolone provided in the embodiments of this disclosure; Figure 5 The N-acetyl-5-bromo-4-chloro-3-indolone synthesized in Example 10 of this disclosure is 1 H-NMR spectrum; Figure 6The N-acetyl-5-bromo-4-chloro-3-indolone synthesized in Example 10 of this disclosure is 13 C-NMR spectrum; Figure 7 This is the ESI-HRMS spectrum of N-acetyl-5-bromo-4-chloro-3-indolone synthesized in Example 10 of this disclosure; Figure 8 The N-acetyl-5-bromo-6-chloro-3-indolone synthesized in Example 10 of this disclosure is 1 H-NMR spectrum; Figure 9 The N-acetyl-5-bromo-6-chloro-3-indolone synthesized in Example 10 of this disclosure is 13 C-NMR spectrum; Figure 10 This is the ESI-HRMS spectrum of N-acetyl-5-bromo-6-chloro-3-indolone synthesized in Example 10 of this disclosure. Detailed Implementation

[0016] To provide a more detailed understanding of the features and technical content of the embodiments of this disclosure, the implementation of the embodiments of this disclosure will be described in detail below with reference to the accompanying drawings. The accompanying drawings are for illustrative purposes only and are not intended to limit the embodiments of this disclosure. In the following technical description, for ease of explanation, several details are used to provide a full understanding of the disclosed embodiments. However, one or more embodiments may still be implemented without these details. In other cases, well-known structures, steps, and apparatus may be simplified to simplify the illustrations.

[0017] The terms "first," "second," etc., used in the specification, claims, and accompanying drawings of this disclosure are used to distinguish similar objects and are not necessarily used to describe a specific order or sequence. It should be understood that such terms can be used interchangeably where appropriate for the embodiments of this disclosure described herein. Furthermore, the terms "comprising" and "having," and any variations thereof, are intended to cover non-exclusive inclusion.

[0018] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The term "multiple" means two or more. In embodiments of this disclosure, the character " / " indicates an "or" relationship between the preceding and following objects. For example, A / B means: A or B. The term "and / or" describes an association between objects, indicating that three relationships can exist. For example, A and / or B means: A or B, or, A and B.

[0019] It should be noted that, unless otherwise specified, the embodiments and features described in the present disclosure can be combined with each other.

[0020] Those skilled in the art will understand that in the methods described in this application and other parts thereof, for example, in the methods of various embodiments, examples, or claims, the order in which the steps are written does not imply a strict execution order and does not constitute any limitation on the implementation process. The detailed execution order of each step should be determined by its function and possible internal logic. Unless otherwise specified, all steps of this application may be performed sequentially or randomly, preferably sequentially. For example, if the method includes steps A and B, it means that the method may include steps A and B performed sequentially, or it may include steps B and A performed sequentially. For example, if the method may also include step C, it means that step C can be added to the method in any order. For example, the method may include steps A, B, and C, or it may include steps A, C, and B, or it may include steps C, A, and B, etc.

[0021] In this application, open-ended technical features or solutions described using terms such as "containing," "including," or "comprising" do not exclude additional members beyond those listed unless otherwise specified. They can be considered as providing both closed-ended features or solutions comprised of the listed members and open-ended features or solutions that include additional members beyond the listed members. For example, if A includes a1, a2, and a3, it may also include other members or exclude additional members unless otherwise specified. This can be considered as providing both the feature or solution that "A consists of a1, a2, and a3" and the feature or solution that "A includes not only a1, a2, and a3, but also other members."

[0022] The "range" disclosed in this application can be defined in the form of a lower limit and an upper limit. A given range is defined by selecting a lower limit and an upper limit, which define the boundaries of the specific range. Ranges defined in this way can include or exclude endpoints. Any endpoint can be included or excluded independently and can be combined arbitrarily; that is, any lower limit can be combined with any upper limit to form a range. For example, if ranges of 10~80 and 30~40 are listed for a specific parameter, it is expected that ranges of 10~40 and 30~80 will also be included. Furthermore, if minimum range values ​​of 1 and 2 are listed, and if maximum range values ​​of 3, 4, and 5 are also listed, then the following ranges can all be expected: 1~3, 1~4, 1~5, 2~3, 2~4, and 2~5. In this application, unless otherwise stated, the numerical range "a~b" represents a shortened representation of any combination of real numbers between a and b, where a and b are real numbers. For example, the numerical range "0~5" indicates that all real numbers between "0" and "5" have been listed in this article; "0~5" is simply a shortened representation of these numerical combinations. Furthermore, when a parameter is described as an integer ≥2, it is equivalent to listing integers such as 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, etc. For instance, when a parameter is described as an integer selected from "2~10", it is equivalent to listing the integers 2, 3, 4, 5, 6, 7, 8, 9, and 10.

[0023] In this application, unless otherwise specified, A (e.g., B) means that B is a non-limiting example of A, and it is understood that A is not limited to B.

[0024] Combination Figure 1 As shown, this disclosure provides a method for preparing N-acetyl-5-bromo-chloro-3-indolone, comprising: S110. Under alkaline conditions, 5-bromo-chloroindole undergoes an oxidation reaction to introduce an acetoxy group at the 3-position of the indole ring, yielding 5-bromo-chloro-3-acetoxyindole; wherein the chlorine atom is located on the benzene ring of the indole ring.

[0025] S120 and 5-bromo-chloro-3-acetoxyindole undergo an N-acetylation reaction to introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, yielding N-acetyl-5-bromo-chloro-3-acetoxyindole.

[0026] S130. Under acidic conditions, N-acetyl-5-bromo-chloro-3-acetoxyindole undergoes a selective hydrolysis reaction to remove the acetyl group on the oxygen atom, yielding N-acetyl-5-bromo-chloro-3-indoleone.

[0027] Combination Figure 2As shown in the embodiments of this disclosure, the synthetic route for N-acetyl-5-bromo-chloro-3-indolone is as follows:

[0028] The oxidation reaction in S110 can also be called oxidative acetoxylation, which involves simultaneously introducing an acetoxy group at the 3-position of the indole ring under oxidative conditions. In step S110, 5-bromo-chloroindole (Formula II) is used as the starting material, an oxidant is added, and the oxidation reaction is carried out under alkaline conditions to introduce an acetoxy group at the 3-position of the indole ring, yielding 5-bromo-chloro-3-acetoxyindole (Formula III). The oxidant can be a high-valence iodine oxidant, such as iodobenzene diacetate (DIB) or Dess-Martin periodinane (DMP). Direct acetoxylation of the indole ring at the 3-position can be achieved in a single step, avoiding the two-step process of preparing 3-hydroxyindole followed by esterification in traditional methods, thus simplifying the operation and improving reaction efficiency.

[0029] In step S120, the 5-bromo-chloro-3-acetoxyindole of Formula III obtained in step S110 undergoes an N-acetylation reaction to introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, yielding N-acetyl-5-bromo-chloro-3-acetoxyindole of Formula IV. The introduction of this N-acetyl group protects the nitrogen atom of the indole ring, preventing side reactions in subsequent reactions. Furthermore, it provides a stable amide structure for the target product. This amide bond remains stable under the acidic hydrolysis conditions of step S130, thereby achieving selective removal of the acetyl group from the oxygen atom, which is beneficial for the separation and purification of the final product.

[0030] In step S130, under acidic conditions, the N-acetyl-5-bromo-chloro-3-acetoxyindole of formula IV obtained in step S120 is selectively hydrolyzed, removing only the acetyl group on the oxygen atom while leaving the acetyl group on the nitrogen atom unchanged, to obtain the target product N-acetyl-5-bromo-chloro-3-indolone of formula I. This step utilizes the characteristic that the hydrolytic activity of the ester bond (acetoxy group) under acidic conditions is much higher than that of the amide bond (N-acetyl group), removing only the acetyl group on the oxygen atom while retaining the acetyl group on the nitrogen atom, thereby achieving selective hydrolysis.

[0031] In Formulas I to IV above, the chlorine atoms are all located on the benzene ring of the indole ring, for example, at positions 4, 6, or 7. In specific embodiments of this application, based on reaction mechanism analysis, it can be understood that when the chlorine atoms are located at other positions on the benzene ring (e.g., position 7), the three-step transformation can also be achieved by appropriately adjusting the reaction conditions.

[0032] The method for preparing N-acetyl-5-bromo-chloro-3-indolone provided in this disclosure uses 5-bromo-chloroindole as the starting material and directly obtains the target product through a three-step process of "oxidative acetylation, N-acetylation, and selective hydrolysis". Compared with the prior art, this method avoids the two-step process of first preparing 3-hydroxyindole and then esterifying it, and also avoids the ring-condensation process involved in the Bischler-Möhlau type intramolecular cyclization-diacetization tandem reaction, shortening the reaction steps and simplifying the operation. Simultaneously, this preparation method avoids the safety hazards caused by the ring-condensation process from the source and avoids the use of highly toxic reagents such as allyl alcohol and allyl bromoacetate. The reaction conditions are mild, effectively improving the safety of the preparation process and reducing costs. Furthermore, each step of this preparation method achieves a high yield, improving the overall yield and preparation efficiency, thereby enabling the efficient preparation of N-acetyl-5-bromo-chloro-3-indolone, and providing a high-quality intermediate for the preparation of 5-bromo-chloro-3-indole glycosides and chromogenic culture media.

[0033] Optionally, the chlorine atom is located at the 4-position of the indole ring.

[0034] Combination Figure 3 As shown in the figure, this embodiment provides a method for preparing N-acetyl-5-bromo-4-chloro-3-indolone, and its synthetic route is as follows:

[0035] In step S110, 5-bromo-chloroindole undergoes an oxidation reaction under alkaline conditions to introduce an acetoxy group at the 3-position of the indole ring, yielding 5-bromo-chloro-3-acetoxyindole. This includes: using 5-bromo-4-chloroindole of Formula II-1 as the starting material, adding an oxidant, and carrying out an oxidation reaction under alkaline conditions to introduce an acetoxy group at the 3-position of the indole ring, yielding 5-bromo-4-chloro-3-acetoxyindole of Formula III-1.

[0036] In step S120, 5-bromo-chloro-3-acetoxyindole undergoes an N-acetylation reaction to introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, yielding N-acetyl-5-bromo-chloro-3-acetoxyindole, comprising: N-acetylation of 5-bromo-4-chloro-3-acetoxyindole of Formula III-1 to introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, yielding N-acetyl-5-bromo-4-chloro-3-acetoxyindole of Formula IV-1.

[0037] In step S130, under acidic conditions, N-acetyl-5-bromo-chloro-3-acetoxyindole undergoes a selective hydrolysis reaction to remove the acetyl group on the oxygen atom, yielding N-acetyl-5-bromo-chloro-3-indolone. This includes: under acidic conditions, selectively hydrolyzing N-acetyl-5-bromo-4-chloro-3-acetoxyindole as shown in Formula IV-1 to remove only the acetyl group on the oxygen atom, while the acetyl group on the nitrogen atom remains unchanged, yielding the target product N-acetyl-5-bromo-4-chloro-3-indolone as shown in Formula I-1.

[0038] In this embodiment, the chlorine atom is located at position 4 of the indole ring, adjacent to the bromine atom at position 5, while only two adjacent hydrogen atoms remain at positions 6 and 7 on the benzene ring. This substitution mode has been verified by Examples 1-10 and exhibits high reaction yield. The prepared N-acetyl-5-bromo-4-chloro-3-indoleone can serve as a key intermediate for the synthesis of 5-bromo-4-chloro-3-indole glycosides, which can then be used in chromogenic media.

[0039] Optionally, the chlorine atom is located at the 6 position of the indole ring.

[0040] Combination Figure 4 As shown in the figure, this embodiment provides a method for preparing N-acetyl-5-bromo-6-chloro-3-indolone, and its synthetic route is as follows:

[0041] In step S110, 5-bromo-chloroindole undergoes an oxidation reaction under alkaline conditions to introduce an acetoxy group at the 3-position of the indole ring, yielding 5-bromo-chloro-3-acetoxyindole. This includes: using 5-bromo-6-chloroindole of Formula II-2 as the starting material, adding an oxidant, and carrying out an oxidation reaction under alkaline conditions to introduce an acetoxy group at the 3-position of the indole ring, yielding 5-bromo-6-chloro-3-acetoxyindole of Formula III-2.

[0042] In step S120, 5-bromo-chloro-3-acetoxyindole undergoes an N-acetylation reaction to introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, yielding N-acetyl-5-bromo-chloro-3-acetoxyindole, comprising: N-acetylation of the 5-bromo-6-chloro-3-acetoxyindole shown in Formula III-2 to introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, yielding N-acetyl-5-bromo-6-chloro-3-acetoxyindole shown in Formula IV-2.

[0043] In step S130, under acidic conditions, N-acetyl-5-bromo-chloro-3-acetoxyindole undergoes a selective hydrolysis reaction to remove the acetyl group on the oxygen atom, yielding N-acetyl-5-bromo-chloro-3-indolone. This includes: under acidic conditions, selectively hydrolyzing N-acetyl-5-bromo-6-chloro-3-acetoxyindole as shown in Formula IV-2 to remove only the acetyl group on the oxygen atom, while the acetyl group on the nitrogen atom remains unchanged, yielding the target product N-acetyl-5-bromo-6-chloro-3-indolone as shown in Formula I-2.

[0044] In this embodiment, the chlorine atom is located at position 6 of the indole ring, in a meta position with the bromine atom at position 5, while the two hydrogen atoms at positions 4 and 7 of the benzene ring are isolated from each other. This substitution mode was also verified by Examples 1-10, showing excellent reaction yield. The prepared N-acetyl-5-bromo-6-chloro-3-indoleone can be used as a key intermediate for the synthesis of 5-bromo-6-chloro-3-indole glycoside. This glycoside exhibits a purple-red color in microbial colorimetric detection, complementing the blue-green color of the 4-chloro isomer, thus meeting the colorimetric requirements of different detection scenarios or different strains.

[0045] The two schemes described above correspond to isomers of the chlorine atom at the 4- and 6-positions of the indole ring, respectively. Both employ the same three-step synthetic strategy, differing only in the position of the chlorine atom in the starting material. Experimental data show that both schemes achieve high yields and offer advantages such as short steps, high safety, and avoidance of highly toxic reagents. Those skilled in the art can select appropriate starting materials based on the colorimetric requirements of the target glycoside substrate, such as blue-green or purple-red, and use the method of this invention for preparation.

[0046] Optionally, in step S110, the oxidant used in the oxidation reaction is a high-valent iodine oxidant capable of providing acetoxy groups.

[0047] High-valence iodine oxidants that can provide acetoxy groups are organic iodine compounds whose molecular structure contains an acetoxy group (-OAc) and whose iodine atom is in a high valence state (such as +3 or +5). These compounds can oxidize the 3-position CH bond of the indole ring under alkaline conditions, and at the same time, they act as a source of acetoxy groups, introducing the acetoxy group into the 3-position of the indole ring, thereby realizing a one-step oxidative acetoxylation reaction.

[0048] Optionally, in step S110, the oxidant used in the oxidation reaction is one of iodophenylacetic acid or Dess-Martin reagent.

[0049] Iodophenyl diacetic acid (DIB) and Des Martin reagent (DMP) are both high-valent iodine reagents. Under alkaline conditions, they can efficiently achieve acetoxylation at the 3-position of the indole ring. They are characterized by mild performance and good selectivity, and can avoid triggering other side reactions of the indole ring.

[0050] Optionally, in step S110, the oxidant used in the oxidation reaction is iodophenyl diacetic acid.

[0051] DIB has advantages such as low cost, moderate reactivity, and simple post-processing. Furthermore, using DIB as an oxidant can achieve higher reaction yields.

[0052] Optionally, in step S110, the alkaline reagent used in the alkaline conditions of the oxidation reaction is one of potassium hydroxide, sodium hydroxide, 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), 1,5-diazabicyclo[4.3.0]non-5-ene (DBN), or triethylamine.

[0053] In this embodiment, the alkaline reagent is used to promote the deprotonation of the nitrogen atom in the indole ring, forming a nitrogen anion, which then reacts with the oxidant to initiate an oxidative acetoxylation reaction. These alkaline reagents include both inorganic strong bases, such as potassium hydroxide and sodium hydroxide, and organic strong bases, such as DBU, DBN, and triethylamine, all of which can effectively promote the oxidation reaction.

[0054] Optionally, in step S110, the alkaline reagent used in the alkaline conditions of the oxidation reaction is potassium hydroxide.

[0055] Potassium hydroxide is a strong alkaline substance and inexpensive, and it can efficiently promote the deprotonation of nitrogen atoms in the indole ring, thereby obtaining a higher reaction yield.

[0056] Optionally, in step S110, the oxidation reaction is carried out in a first solvent, which is one or more of acetonitrile, dioxane, N,N-dimethylformamide (DMF), and N,N-dimethylacetamide (DMAC).

[0057] All of the above solvents are polar aprotic solvents, capable of effectively dissolving the reaction substrate 5-bromochloroindole, basic reagents, and oxidizing agents, providing a homogeneous reaction environment conducive to the smooth progress of the reaction. Acetonitrile has moderate polarity and a suitable boiling point; dioxane (1,4-dioxane) exhibits good solubility and stability; DMF and DMAC are strongly polar aprotic solvents with excellent solubility for a variety of reactants. Furthermore, two or more of the above solvents can be used as a mixed solvent, and the polarity and solubility of the reaction system can be altered by adjusting the mixing ratio, thereby further optimizing the reaction effect.

[0058] Optionally, in step S110, the oxidation reaction is carried out in a first solvent, which is acetonitrile.

[0059] Acetonitrile exhibits good chemical stability against high-valent iodine oxidants (such as iodophenylacetic acid and Dess-Martin reagent), resulting in higher reaction yields.

[0060] Optionally, in step S110, the reaction temperature of the oxidation reaction is 10~80 °C.

[0061] Reaction temperature is a key factor affecting the reaction rate and selectivity of the oxidation reaction in step S110. When the reaction temperature is in the range of 10~80 °C, the reaction rate is moderate, which can effectively promote the acetoxylation reaction at the 3-position of the indole ring while inhibiting the occurrence of side reactions, thereby ensuring a high reaction yield. The reaction temperature can be any temperature value in the range of 10~80 °C, such as 10 °C, 15 °C, 20 °C, 25 °C, 30 °C, 35 °C, 40 °C, 45 °C, 50 °C, 60 °C, 70 °C, or 80 °C, etc.

[0062] Optionally, in step S110, the reaction temperature of the oxidation reaction is 30~40 °C. Optionally, the reaction temperature of the oxidation reaction is 35 °C.

[0063] Within a reaction temperature range of 30–40 °C, and especially at 35 °C, the oxidation reaction rate and selectivity in step S110 achieve a good balance. On the one hand, the stability of the reaction substrate, basic reagent, and oxidant is maintained, avoiding side reactions caused by excessively high temperatures. On the other hand, a moderate reaction rate avoids slow reactions due to excessively low temperatures, thereby promoting the forward reaction and increasing the yield of the target product.

[0064] Optionally, in step S110, the molar ratio of 5-bromo-chloroindole, basic reagent and oxidant in the oxidation reaction is 1:(1.5~3):(1.2~2).

[0065] Within this ratio range, both the basic reagent and the oxidant are in excess relative to the substrate 5-bromochloroindole, which is beneficial for ensuring complete substrate conversion. An excess of the basic reagent promotes complete deprotonation of the indole ring nitrogen, accelerating the formation of the nitrogen anion. An excess of the oxidant ensures complete acetoxylation at the 3-position of 5-bromochloroindole. This ratio range ensures a high reaction yield while avoiding waste of raw materials and post-processing burdens caused by excessive excess. Specifically, the molar amount of 5-bromochloroindole is 1; the molar amount of the basic reagent can be any value within the range of 1.5 to 3, such as 1.5, 1.6, 1.7, 1.8, 1.9, 2, 2.2, 2.3, 2.5, 2.6, 2.8, or 3; and the molar amount of the oxidant can be any value within the range of 1.2 to 2, such as 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, or 2.

[0066] Optionally, in step S110, the molar ratio of 5-bromo-chloroindole, the basic reagent, and the oxidant in the oxidation reaction is 1:2:1.5. This optimizes the reactant ratio, ensuring sufficient deprotonation and acetylation while avoiding waste from excessive reagents.

[0067] Optionally, in step S110, potassium hydroxide is used as an alkaline reagent and iodophenyl diacetic acid is used as an oxidant in the oxidation reaction, and the molar ratio of 5-bromo-chloroindole, potassium hydroxide and iodophenyl diacetic acid is 1:2:1.5.

[0068] Potassium hydroxide provides a strongly alkaline environment, which efficiently promotes the deprotonation of the nitrogen atom in the indole ring. Iodophenyl diacetic acid (DIB), as a mild and efficient oxidative acetoxylation reagent, can rapidly generate an active intermediate under alkaline conditions, introducing the acetoxy group at the 3-position of the indole ring. This combination of conditions achieves the advantages of short reaction time, high yield, and simple operation.

[0069] Optionally, in step S110, after 5-bromo-chloroindole undergoes an oxidation reaction under alkaline conditions to introduce an acetoxy group at the 3-position of the indole ring, the reaction further includes: adding ethyl acetate to the reaction solution of the oxidation reaction, washing the organic phase with saturated brine, drying it with anhydrous magnesium sulfate and filtering it, concentrating the filtrate under reduced pressure, and separating the residue by column chromatography.

[0070] Following the oxidation reaction, the above post-treatment steps effectively remove unreacted raw materials, basic reagents, oxidants, and byproducts from the reaction system, thereby obtaining the high-purity target intermediate 5-bromo-chloro-3-acetoxyindole. Specifically, adding ethyl acetate to the reaction solution extracts the organic products in the reaction mixture, transferring the target compound from the polar reaction solvent (such as acetonitrile) to the organic phase. Washing the organic phase with saturated brine removes residual water-soluble impurities (such as inorganic salts and excess alkali) and reduces the moisture content, facilitating subsequent drying. Anhydrous magnesium sulfate further removes trace amounts of moisture from the organic phase, preventing adverse effects on subsequent reactions (such as N-acetylation). Reduced-pressure concentration removes low-boiling-point solvents (such as ethyl acetate) at lower temperatures, preventing thermal decomposition of the target product. Column chromatography purification effectively removes unreacted 5-bromo-chloroindole raw materials, residual basic reagents, oxidants, and byproducts, improving the chromatographic purity of the target product and providing a high-quality intermediate for the next reaction, ensuring the stability and reproducibility of the overall yield of the three-step method. After separation by column chromatography, the residue yields 5-bromo-chloro-3-acetoxyindole.

[0071] Optionally, the column chromatography uses a mixed solvent of petroleum ether and ethyl acetate as the eluent, with a volume ratio of petroleum ether to ethyl acetate of 1:1.

[0072] A 1:1 volume ratio of petroleum ether to ethyl acetate was used as the eluent. This ratio allows for effective separation based on the polarity difference between the target product and impurities. Petroleum ether is a nonpolar component, while ethyl acetate is a polar component. The 1:1 volume ratio provides moderate elution strength, allowing the target product to move smoothly from the column while simultaneously eluting the more polar byproducts and the less polar nonpolar impurities, thus yielding high-purity 5-bromochloro-3-acetoxyindole.

[0073] Optionally, in step S120, the acetylation reagent used in the N-acetylation reaction is one of acetic anhydride and acetyl chloride.

[0074] Both acetic anhydride and acetyl chloride can effectively introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, generating N-acetyl-5-bromo-chloro-3-acetoxyindole. Acetic anhydride exhibits a mild reaction, producing acetic acid as a byproduct during acetylation, which is less corrosive to equipment and easily removed through subsequent treatment. Acetyl chloride, on the other hand, is more reactive and can complete acetylation at lower temperatures or in a shorter time.

[0075] Optionally, in step S120, the acetylation reagent used in the N-acetylation reaction is acetic anhydride.

[0076] When acetic anhydride is used as an acetylation reagent, it reacts with indole nitrogen atoms to produce an N-acetyl product and acetic acid. Acetic acid is a weak acidic small molecule that can be removed by simple water washing or distillation, leaving no residue in the reaction system that could affect subsequent steps. The reaction is mild, and the byproducts are easily removed. Moreover, acetic anhydride is not highly corrosive to equipment, the reaction system does not require strictly anhydrous conditions, it is highly safe, inexpensive, suitable for scale-up production, and even more suitable for industrial production.

[0077] Optionally, in step S120, the reaction temperature of the N-acetylation reaction is 10~100 °C.

[0078] Within this temperature range, the reaction system possesses sufficient energy to overcome the activation barrier of the acetylation reaction, promoting the transfer of the acetyl group from the acetylation reagent to the nitrogen atom at the 1-position of the indole ring, while effectively suppressing side reactions. Specifically, when the reaction temperature is greater than or equal to 10 °C, the reaction rate can be increased, the reaction time shortened, and the reaction proceeds fully. When the temperature is less than or equal to 100 °C, the self-decomposition of the acetylation reagent can be avoided or mitigated, while reducing the probability of side reactions occurring at other positions of the indole ring. The reaction temperature can be any value within the range of 10–100 °C, such as 10 °C, 20 °C, 30 °C, 40 °C, 50 °C, 55 °C, 60 °C, 66 °C, 70 °C, 80 °C, 90 °C, 100 °C, etc. Those skilled in the art can adjust the temperature within this range according to the actual reaction progress; for example, a lower temperature can be used in the initial stage of the reaction to control exothermic reactions, and the temperature can be appropriately increased in the later stage to promote complete reaction.

[0079] Optionally, in step S120, the reaction temperature of the N-acetylation reaction is 60~70 °C.

[0080] The N-acetylation reaction, occurring at a temperature of 60–70 °C, precisely matches the kinetic characteristics of the reaction, providing optimal energy support for the directional transfer of the acetyl group from the acetylation reagent to the nitrogen atom at the 1-position of the indole ring. This avoids both the slow reaction rate and raw material residue caused by low temperatures, and the side reactions induced by high temperatures, achieving efficient and directional execution of the main reaction and effectively improving reaction selectivity and product purity. Within this temperature range, the kinetic rate of the acetylation reaction is moderate, ensuring that the reaction is completed within a reasonable time, shortening the production cycle, while effectively avoiding side reactions such as the self-decomposition of the acetylation reagent and non-directional acylation of other active sites on the indole ring caused by high temperatures, thus ensuring the structural integrity of the product. Simultaneously, the target product, N-acetyl-5-bromo-chloro-3-acetoxyindole, exhibits excellent chemical stability within this temperature range, resisting hydrolysis, decomposition, and other degradation reactions, thus ensuring the smooth progress of subsequent separation, purification, post-processing operations, and long-term storage of the product.

[0081] Optionally, in step S120, the N-acetylation reaction is carried out in a second solvent, which is tetrahydrofuran (THF).

[0082] THF is a commonly used polar aprotic solvent that can readily dissolve the substrate 5-bromochloro-3-acetoxyindole, acetylation reagents, and the catalyst DMAP. THF is stable to acids, bases, and acetylation reagents and does not participate in side reactions. When acetylation reagent is acetic anhydride, it exhibits good compatibility with the catalyst DMAP. Under DMAP catalysis, the N-acetylation of 5-bromochloro-3-acetoxyindole can be completed in tetrahydrofuran at 10–100 °C.

[0083] Optionally, in step S120, the reaction temperature of the N-acetylation reaction is the reflux temperature of the second solvent; when the second solvent is tetrahydrofuran, the reaction temperature of the N-acetylation reaction is the reflux temperature of tetrahydrofuran, which is approximately 66°C under standard atmospheric pressure.

[0084] Using the reflux temperature of the second solvent in the reaction maintains a constant and relatively high temperature in the N-acetylation reaction system, providing sufficient activation energy for the N-acetylation reaction, thereby increasing the reaction rate and shortening the reaction time. Taking tetrahydrofuran as an example, its reflux temperature (approximately 66 °C) can both increase the reaction rate and prevent the decomposition of the acetylation reagent or the indole ring side reaction caused by excessively high temperature. In addition, reflux operation can effectively maintain the homogeneity of the reaction system, promote the formation of a highly active acetylopyridinium intermediate between the catalyst and the acetylation reagent, and enable the efficient transfer of the acetyl group to the indole nitrogen atom. Therefore, using the reflux temperature of tetrahydrofuran as the reaction condition can balance reaction rate, selectivity, and process stability. When the reaction temperature is 66 °C, the acetylation reaction has a moderate reaction rate, which can ensure that the reaction is completed within a reasonable time while avoiding side reactions caused by high temperature, thus maximizing the synergistic optimization of reaction rate, product purity, and product stability.

[0085] Optionally, in step S120, the N-acetylation reaction is carried out in the presence of a catalyst, wherein the catalyst is 4-dimethylaminopyridine (DMAP).

[0086] Using DMAP as a catalyst for N-acetylation reactions, its strong nucleophilicity rapidly activates the acetylation reagents. Both acetic anhydride and acetyl chloride, the two acetylation reagents, synergistically react with DMAP to form highly active acetylopyridinium intermediates, accelerating the acyl transfer process. For example, DMAP can efficiently activate the carbonyl carbon of acetic anhydride to form acetylopyridinium ions, thereby increasing the reaction rate. This allows the reaction to proceed rapidly under mild conditions of 10–100 °C, while selectively catalyzing the acetylation of the nitrogen atom at the 1-position of the indole ring while protecting the acetoxy group at the 3-position, resulting in stable and high yields. Furthermore, within the reaction temperature range of 10–100 °C, the activity of the 4-dimethylaminopyridine (DMAP) catalyst can be fully utilized, and the reaction system (e.g., tetrahydrofuran solvent) remains liquid, facilitating operation.

[0087] Optionally, in step S120, the N-acetylation reaction is carried out in the presence of the catalyst 4-dimethylaminopyridine, and the molar ratio of 5-bromo-chloro-3-acetoxyindole, the acetylation agent and 4-dimethylaminopyridine is 1:(5-15):(0.5-3).

[0088] Within this ratio range, the acetylation reagent (such as acetic anhydride or acetyl chloride) is in excess relative to the substrate 5-bromo-chloro-3-acetoxyindole, allowing for sufficient acetylation of the nitrogen atom in the indole ring. Simultaneously, the amount of catalyst 4-dimethylaminopyridine is moderate, effectively catalyzing the reaction while avoiding the introduction of impurities due to excess catalyst. This ratio range provides flexibility for those skilled in the art in adjusting the process, allowing for the selection of appropriate ratios based on actual reaction activity, reagent costs, and post-processing difficulty, all while achieving good N-acetylation yields. Specifically, the molar amount of 5-bromo-chloro-3-acetoxyindole is 1, the molar amount of the acetylation reagent can be any value within the range of 5 to 15, such as 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15, and the molar amount of catalyst 4-dimethylaminopyridine can be any value within the range of 0.5 to 3, such as 0.5, 1, 1.5, 2, 2.5, or 3.

[0089] Optionally, in step S120, the molar ratio of 5-bromo-chloro-3-acetoxyindole, the acetylation reagent, and 4-dimethylaminopyridine is 1:10:2.

[0090] At this ratio, 10 equivalents of the acetylation reagent can provide sufficient acyl source to drive the reaction to completion. 2 equivalents of DMAP can provide sufficiently high catalytic activity, enabling the reaction to be completed in a shorter time, improving preparation efficiency, and balancing reaction rate and conversion.

[0091] Optionally, in step S120, the N-acetylation reaction is carried out in the presence of the catalyst 4-dimethylaminopyridine, with acetic anhydride as the acetylation reagent, and the molar ratio of 5-bromo-chloro-3-acetoxyindole, acetic anhydride and 4-dimethylaminopyridine is 1:10:2.

[0092] Under these molar ratios, the amount of acetic anhydride used is 10 equivalents of the substrate 5-bromo-chloro-3-acetoxyindole, which provides a sufficient acetyl source to completely convert the nitrogen atom of the indole ring into the N-acetyl product. The amount of DMAP used is 2 equivalents, exhibiting high catalytic activity and effectively increasing the reaction rate.

[0093] Optionally, in step S120, after 5-bromo-chloro-3-acetoxyindole undergoes an N-acetylation reaction to introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, the reaction mixture of the N-acetylation reaction is further subjected to solvent evaporation, ethyl acetate is added to the residue, the resulting suspension is stirred, filtered, and the filter cake is collected.

[0094] After the N-acetylation reaction in step S120, a post-processing step is included. Taking advantage of the low solubility of the target product N-acetyl-5-bromo-chloro-3-acetoxyindole in ethyl acetate, solvent displacement is used to precipitate the product from the solution. Evaporation of tetrahydrofuran removes low-boiling-point solvents, preventing them from affecting subsequent crystallization. Upon addition of ethyl acetate, the target product N-acetyl-5-bromo-chloro-3-acetoxyindole precipitates slowly due to poor polarity matching, forming a suspension. Stirring promotes crystal growth, and filtration yields a high-purity solid product.

[0095] Optionally, in step S130, the acidic reagent used in the selective hydrolysis reaction is a dilute sulfuric acid solution with a concentration of 10% to 85%.

[0096] Dilute sulfuric acid can selectively hydrolyze the acetyl group (acetoxy group) on the oxygen atom of N-acetyl-5-bromo-chloro-3-acetoxyindole, while retaining the acetyl group (amide bond) on the nitrogen atom. The ester bond (acetoxy group) is much more reactive for hydrolysis under acidic conditions than the amide bond (N-acetyl group), thus selective deprotection can be achieved under mild conditions to generate N-acetyl-5-bromo-chloro-3-indole. The dilute sulfuric acid solution can be used at any concentration ranging from 10% to 85%.

[0097] Optionally, in step S130, the acidic reagent used in the selective hydrolysis reaction is an 80% dilute sulfuric acid solution.

[0098] At this concentration, the acidity and water content are well balanced, which can provide sufficient protons to catalyze the breaking of ester bonds, while avoiding the hydrolysis of amide bonds or carbonization of products due to excessive acidity.

[0099] Optionally, in step S130, the reaction temperature of the selective hydrolysis reaction is 10~50 °C.

[0100] Within a reaction temperature range of 10–50 °C, the hydrolysis rate of the ester bond (acetoxy group) is moderate, effectively breaking the acetyl group on the oxygen atom while maintaining the stability of the amide bond (N-acetyl group), achieving selective deprotection. A reaction temperature greater than or equal to 10 °C can increase the hydrolysis rate, shorten the reaction time, and improve production efficiency. A temperature less than or equal to 50 °C can avoid partial hydrolysis of the amide bond or product decomposition caused by overheating, thus ensuring high yield and high purity. The reaction temperature can be any value within the range of 10–50 °C, such as 10 °C, 15 °C, 20 °C, 25 °C, 30 °C, 35 °C, 40 °C, 45 °C, 50 °C, etc. Those skilled in the art can adjust the temperature within the 10–50 °C range according to the actual reaction progress; for example, using a lower temperature in the initial stage to control the exothermic reaction and then appropriately increasing the temperature in the later stage to promote complete reaction can both achieve good selective hydrolysis results.

[0101] Optionally, in step S130, the reaction temperature of the selective hydrolysis reaction is 20~30 ℃.

[0102] When the reaction temperature of selective hydrolysis is 20~30 ℃, it can balance the reaction rate and selectivity, avoiding the slow reaction caused by low temperature and preventing side reactions caused by high temperature. At the same time, it does not require strict temperature control and is easy to operate.

[0103] Optionally, in step S130, the reaction temperature of the selective hydrolysis reaction is 25 °C.

[0104] At a reaction temperature of 25 °C, the hydrolysis rate and selectivity of the ester bond reach an optimal balance. At this temperature, the amide bond is extremely stable, and no hydrolysis byproducts of the N-acetyl group are observed, thus improving the structural integrity of the target product. Furthermore, room temperature operation eliminates the need for additional heating or cooling equipment, resulting in low energy consumption, high safety, and suitability for industrial-scale production. Additionally, after the reaction at 25 °C, the reaction solution can be directly poured into ice water to precipitate the product, simplifying post-processing.

[0105] Optionally, in step S130, under acidic conditions, N-acetyl-5-bromo-chloro-3-acetoxyindole undergoes a selective hydrolysis reaction to remove the acetyl group on the oxygen atom, and after this process, the reaction solution from the selective hydrolysis reaction is poured into ice water to precipitate a solid compound.

[0106] After the selective hydrolysis reaction is complete, the above-mentioned post-treatment operation is performed by slowly pouring the reaction solution into ice water. The precipitated solid compound is N-acetyl-5-bromo-chloro-3-indolone. This post-treatment process utilizes the characteristic that the solubility of the target product is significantly reduced in a low-temperature aqueous phase. By diluting and cooling, the product is rapidly crystallized out, offering advantages such as simple operation, no need for organic extraction or column chromatography purification, and suitability for industrial production. Pouring the reaction solution into ice water rapidly lowers the system temperature and acid concentration, quickly terminating the hydrolysis reaction and preventing excessive hydrolysis that could lead to amide bond breakage or product decomposition. Furthermore, the target product, N-acetyl-5-bromo-chloro-3-indolone, has extremely low solubility in cold water, while byproducts such as acetic acid and excess sulfuric acid have good water solubility, thus promoting product crystallization and resulting in the precipitation of the product in a high-purity solid form. After dilution with ice water, impurities such as sulfuric acid and acetic acid in the reaction solution remain in the aqueous phase, and the product can be separated from the impurities by filtration, eliminating the need for cumbersome extraction or chromatographic purification. Using this post-processing method, the yields of the two isomers reached 88.0% and 89.0%, respectively, and the products were brown solids that could be directly dried for subsequent applications. This post-processing scheme effectively simplifies the operation process, shortens the production cycle, reduces costs, and ensures high yield and high purity, showing promising prospects for industrial applications.

[0107] This disclosure provides an N-acetyl-5-bromo-chloro-3-indolone, prepared using the preparation method described in any of the foregoing disclosure embodiments.

[0108] The N-acetyl-5-bromo-chloro-3-indolone provided in this disclosure has the chemical structural formula shown in Formula I: I.

[0109] N-acetyl-5-bromo-chloro-3-indolone can be used as a key intermediate in the preparation of 5-bromo-chloro-3-indolyl glycoside chromogenic substrates. In this N-acetyl-5-bromo-chloro-3-indolone, the substitution position of the chlorine atom on the indole ring is the same as that of the chlorine atom in Formula I of the aforementioned "Preparation Method of N-acetyl-5-bromo-chloro-3-indolone," and will not be repeated here. N-acetyl-5-bromo-chloro-3-indolone can be N-acetyl-5-bromo-4-chloro-3-indolone, N-acetyl-5-bromo-6-chloro-3-indolone, or N-acetyl-5-bromo-7-chloro-3-indolone.

[0110] This disclosure provides a 5-bromo-chloro-3-indole glycoside, which is prepared by glycosylation and deprotection reaction using N-acetyl-5-bromo-chloro-3-indole ketone as described in the foregoing disclosure.

[0111] This disclosure uses N-acetyl-5-bromo-chloro-3-indole ketone prepared in the aforementioned embodiments as a raw material. Through glycosylation and deprotection reactions, 5-bromo-chloro-3-indole glycosides can be efficiently prepared. The 5-bromo-chloro-3-indole glycosides provided in this disclosure can be used as highly sensitive chromogenic substrates. They can be hydrolyzed by specific enzymes of target microorganisms to release indolephenol chromogenic groups, producing characteristic colors. For example, the 4-chloro isomer (5-bromo-4-chloro-3-indole glycoside) is blue-green, and the 6-chloro isomer (5-bromo-6-chloro-3-indole glycoside) is purple-red. This enables rapid visual identification of colonies with low background interference, making it suitable for rapid detection of pathogenic microorganisms such as Escherichia coli and Staphylococcus aureus.

[0112] This disclosure provides a chromogenic culture medium using 5-bromo-chloro-3-indoleglycoside as a chromogenic substrate as described in the foregoing disclosure embodiments.

[0113] The chromogenic culture medium of this disclosure uses 5-bromo-chloro-3-indole glycoside from the aforementioned embodiments as the chromogenic substrate, enabling rapid and sensitive detection of target colonies through the specific chromogenic reaction between microbial metabolic enzymes and the substrate. The chromogenic culture medium possesses all the beneficial effects of the aforementioned 5-bromo-chloro-3-indole glycoside, which will not be elaborated further here.

[0114] The following specific embodiments illustrate the preparation method of N-acetyl-5-bromo-chloro-3-indolone, N-acetyl-5-bromo-chloro-3-indolone, 5-bromo-chloro-3-indoloside, and the colorimetric culture medium of this disclosure, to more clearly illustrate the technical problems solved by this application, the technical solutions, and the beneficial effects. Obviously, the described embodiments are only a part of the embodiments of this application, and not all of the embodiments. The following description of at least one exemplary embodiment is merely illustrative and is in no way intended to limit this application or its applications.

[0115] Where specific techniques or conditions are not specified in the examples, they shall be performed in accordance with the techniques or conditions described in the literature in this field or in accordance with the product instructions. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0116] Example 1 Example 1 describes the preparation of 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2) via oxidation reactions. Reaction conditions: potassium hydroxide was used as the basic reagent; DIB was used as the oxidant; acetonitrile was used as the first solvent; the molar ratio of 5-bromo-chloroindole, potassium hydroxide, and DIB was 1:2:1.5; the reaction temperature was 35°C; and the reaction time was 1.5 hours.

[0117] (1-1) Preparation of 5-bromo-4-chloro-3-acetoxyindole In a flask, 2.30 g (0.01 mol, 1 eq) of 5-bromo-4-chloroindole (Formula II-1), 1.12 g (0.02 mol, 2 eq) of potassium hydroxide, 4.83 g (0.015 mol, 1.5 eq) of DIB, and 100 mL of acetonitrile were added. The resulting reaction mixture was stirred at 35 °C for 1.5 h. After the reaction was complete, 300 mL of ethyl acetate was added, and the organic phase was washed three times with saturated brine, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure. The residue was separated by column chromatography (petroleum ether:ethyl acetate = 1:1) to give 5-bromo-4-chloro-3-acetoxyindole solid of Formula III-1 in 83.0% yield. The obtained 5-bromo-4-chloro-3-acetoxyindole solid was deep purple.

[0118] (1-2) Preparation of 5-bromo-6-chloro-3-acetoxyindole Under the same reaction conditions as (1-1), the starting material was replaced with 5-bromo-6-chloroindole (2.30 g, 0.01 mol, 1 eq) as shown in Formula II-2 to obtain 5-bromo-6-chloro-3-acetoxyindole solid as shown in Formula III-2, with a yield of 85.1%. The obtained 5-bromo-6-chloro-3-acetoxyindole solid was dark purple.

[0119] Under the reaction conditions of this embodiment, the yields of the two isomers, 5-bromo-4-chloro-3-acetoxyindole and 5-bromo-6-chloro-3-acetoxyindole, reached 83.0% and 85.1%, respectively, indicating that the reaction conditions have excellent reaction efficiency and substrate applicability, and can improve the reaction rate and yield of the oxidation reaction step.

[0120] Example 2 Example 2 describes the preparation of 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2). Reaction conditions: the molar ratio of 5-bromo-chloroindole, potassium hydroxide, and DIB was 1:1.5:1.2; the reaction temperature was 25 °C; the reaction time was 4 hours; and the rest were the same as in Example 1.

[0121] (2-1) Preparation of 5-bromo-4-chloro-3-acetoxyindole Oxidation reaction: In a flask, 2.30 g (0.01 mol, 1 eq) of 5-bromo-4-chloroindole, 0.84 g (0.015 mol, 1.5 eq) of potassium hydroxide, 3.86 g (0.012 mol, 1.2 eq) of DIB, and 100 mL of acetonitrile were added. The resulting reaction mixture was stirred at 25 °C for 4 hours. After the reaction was complete, 300 mL of ethyl acetate was added, and the organic phase was washed three times with saturated brine, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure. The residue was separated by column chromatography (petroleum ether:ethyl acetate = 1:1) to give solid 5-bromo-4-chloro-3-acetoxyindole of formula III-1, in a yield of 61.5%.

[0122] (2-2) Preparation of 5-bromo-6-chloro-3-acetoxyindole Under the same reaction conditions as (2-1), the starting material was replaced with 5-bromo-6-chloroindole (2.30 g, 0.01 mol, 1 eq) as shown in Formula II-2 to obtain 5-bromo-6-chloro-3-acetoxyindole solid as shown in Formula III-2, with a yield of 60.0%.

[0123] Compared with Example 1, in Example 2, the molar ratio of 5-bromo-chloroindole, potassium hydroxide, and DIB was reduced from 1:2:1.5 in Example 1 to 1:1.5:1.2, the reaction temperature was reduced from 35 °C to 25 °C, and the reaction time was extended to 4 hours. By reducing the amount of alkaline reagent and oxidant, lowering the reaction temperature, and extending the reaction time, the yields of 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2) were reduced to 61.5% and 60.0%, respectively, which were significantly lower than the 83.0% and 85.1% in Example 1.

[0124] This comparative result indicates that excess basic reagent plays a crucial promoting role in the reaction. Excess potassium hydroxide not only ensures the complete deprotonation of the indole ring nitrogen atom, generating a high concentration of nitrogen anions (nucleophilic species), but also effectively neutralizes the protons generated during the reaction, maintaining a stable basic environment and thus accelerating the forward direction of the oxidative acetoxylation reaction. When the amount of basic reagent is insufficient, the deprotonation equilibrium shifts to the left, the nitrogen anion concentration decreases, and both the reaction rate and conversion are inhibited. Excess oxidant has a driving effect. The oxidant DIB acts as both an oxidant and a source of acetoxy groups in the reaction. Reducing the amount of DIB from 1.5 eq to 1.2 eq decreases the concentration of reactive hypervalent iodine species, reduces the probability of oxidation of the CH bond at the 3-position of indole and acetoxy group capture, leading to incomplete conversion of some substrates. Excess oxidant ensures an effective collision frequency between substrate molecules and the oxidant, driving the reaction towards completion. Meanwhile, the reaction temperature has a significant impact on the reaction rate. Compared to 25 °C, the system energy is higher at 35 °C, with more molecules reaching the activation energy threshold, resulting in a significant increase in the reaction rate. Although Example 2 extended the reaction time (4 hours), the proportion of activated molecules was lower at low temperatures, making it difficult to compensate for the rate loss caused by the temperature reduction even with extended reaction time. Furthermore, appropriate heating can improve the homogeneity of the reaction mixture, reduce mass transfer resistance, and further improve conversion efficiency.

[0125] In summary, an appropriate excess of basic reagent and oxidant, along with a suitable reaction temperature (35 °C), can synergistically promote the oxidative acetoxylation reaction at the 3-position of the indole ring, achieving higher substrate conversion and target product yield.

[0126] Example 3 Example 3 describes the preparation of 5-bromo-4-chloro-3-acetoxyindole represented by Formula III-1 and 5-bromo-6-chloro-3-acetoxyindole represented by Formula III-2. Reaction conditions: The reaction temperature was 25 °C, and the rest were the same as in Example 1.

[0127] (3-1) Preparation of 5-bromo-4-chloro-3-acetoxyindole In a flask, 2.30 g (0.01 mol, 1 eq) of 5-bromo-4-chloroindole (Formula II-1), 1.12 g (0.02 mol, 2 eq) of potassium hydroxide, 4.83 g (0.015 mol, 1.5 eq) of DIB, and 100 mL of acetonitrile were added. The resulting reaction mixture was stirred at 25 °C for 1.5 h. After the reaction was complete, 300 mL of ethyl acetate was added, and the organic phase was washed three times with saturated brine, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure. The residue was separated by column chromatography (petroleum ether:ethyl acetate = 1:1) to give solid 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) in 67.2% yield.

[0128] (3-2) Preparation of 5-bromo-6-chloro-3-acetoxyindole Under the same reaction conditions as (3-1), the starting material was replaced with 5-bromo-6-chloroindole (2.30 g, 0.01 mol, 1 eq) as shown in Formula II-2 to obtain 5-bromo-6-chloro-3-acetoxyindole solid as shown in Formula III-2, with a yield of 63.3%.

[0129] Compared with Example 1, in Example 3, under the same ratio and reaction time, the reaction temperature was reduced from 35 °C to 25 °C. The decrease in reaction temperature resulted in a 15.8 and 21.8 percentage point decrease in the yields of the two isomers, 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2), respectively. This indicates that temperature has a significant impact on the reaction rate and yield of the oxidation reaction, and the yield at a reaction temperature of 35 °C is significantly better than that at a reaction temperature of 25 °C.

[0130] Example 4 Example 4 describes the preparation of 5-bromo-4-chloro-3-acetoxyindole as shown in Formula III-1 and 5-bromo-6-chloro-3-acetoxyindole as shown in Formula III-2. Reaction conditions: Dys-Martin reagent (DMP) was used instead of DIB as the oxidant; otherwise, the process was the same as in Example 1.

[0131] (4-1) Preparation of 5-bromo-4-chloro-3-acetoxyindole In a flask, 2.30 g (0.01 mol, 1 eq) of 5-bromo-4-chloroindole (Formula II-1), 1.12 g (0.02 mol, 2 eq) of potassium hydroxide, 6.36 g (0.015 mol, 1.5 eq) of DMP, and 100 mL of acetonitrile were added. The resulting reaction mixture was stirred at 35 °C for 1.5 h. After the reaction was complete, 300 mL of ethyl acetate was added, and the organic phase was washed three times with saturated brine, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure. The residue was separated by column chromatography (petroleum ether:ethyl acetate = 1:1) to give solid 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) in 76.4% yield.

[0132] (4-2) Preparation of 5-bromo-6-chloro-3-acetoxyindole Under the same reaction conditions as (4-1), the starting material was replaced with 5-bromo-6-chloroindole (2.30 g, 0.01 mol, 1 eq) as shown in Formula II-2 to obtain 5-bromo-6-chloro-3-acetoxyindole solid as shown in Formula III-2, with a yield of 79.1%.

[0133] In Example 4, DMP was used instead of DIB as the oxidant, and the remaining reaction conditions were the same as in Example 1. The yields of 5-bromo-4-chloro-3-acetoxyindole shown in Formula III-1 and 5-bromo-6-chloro-3-acetoxyindole shown in Formula III-2 were 76.4% and 79.1%, respectively. Although slightly lower than the yield in Example 1, they were still at a high level, proving that DMP is an effective alternative oxidant and broadening the range of oxidant choices.

[0134] Example 5 Example 1 describes the preparation of 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2). Reaction conditions: Triethylamine was used as the basic reagent; all other conditions were the same as in Example 4.

[0135] (5-1) Preparation of 5-bromo-4-chloro-3-acetoxyindole In a flask, 2.30 g (0.01 mol, 1 eq) of 5-bromo-4-chloroindole (Formula II-1), 2.02 g (0.02 mol, 2 eq) of triethylamine, 6.36 g (0.015 mol, 1.5 eq) of DMP, and 100 mL of acetonitrile were added. The resulting reaction mixture was stirred at 35 °C for 1.5 h. After the reaction was complete, 300 mL of ethyl acetate was added, and the organic phase was washed three times with saturated brine, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure. The residue was separated by column chromatography (petroleum ether:ethyl acetate = 1:1) to give solid 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) in 30% yield.

[0136] (5-2) Preparation of 5-bromo-6-chloro-3-acetoxyindole Under the same reaction conditions as (5-1), the starting material was replaced with 5-bromo-6-chloroindole (2.30 g, 0.01 mol, 1 eq) as shown in Formula II-2 to obtain 5-bromo-6-chloro-3-acetoxyindole solid as shown in Formula III-2, with a yield of 31%.

[0137] Compared to Example 4, Example 5 used triethylamine instead of potassium hydroxide as the basic reagent, resulting in a sharp drop in yield to 30% and 31%. Triethylamine is an organic tertiary amine, and its basicity is much weaker than that of potassium hydroxide. Furthermore, the steric hindrance of the triethylamine molecule is significant, hindering the rapid and complete abstraction of protons from the nitrogen atom of the indole ring. This leads to low efficiency in the formation of nitrogen anions, thus impeding the subsequent oxidative acetoxylation reaction. In contrast, the inorganic strong base potassium hydroxide has a stronger deprotonating ability and less steric hindrance, enabling it to efficiently promote the formation of indole nitrogen anions and thus achieve a higher reaction yield.

[0138] Example 6 Example 6 describes the preparation of 5-bromo-4-chloro-3-acetoxyindole as shown in Formula III-1 and 5-bromo-6-chloro-3-acetoxyindole as shown in Formula III-2.

[0139] In Example 6, sodium hydroxide (0.80 g, 0.02 mol, 2 eq) was used instead of potassium hydroxide as the alkaline reagent, and the remaining reaction conditions were the same as in Example 1. The yields of the target products, 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2), obtained in Example 6 were 57.3% and 58%, respectively.

[0140] In Example 6, sodium hydroxide was used instead of potassium hydroxide as the alkaline reagent. Under the same reaction conditions, the yield was significantly lower than that in Example 1, indicating that potassium hydroxide has a stronger deprotonation ability and can more efficiently promote the generation of indole ring nitrogen anions, thereby obtaining a better reaction yield.

[0141] Example 7 Example 7 describes the preparation of 5-bromo-4-chloro-3-acetoxyindole as shown in Formula III-1 and 5-bromo-6-chloro-3-acetoxyindole as shown in Formula III-2.

[0142] In Example 7, dioxane (100 mL) was used instead of acetonitrile as the first solvent, and the remaining reaction conditions were the same as in Example 1. The yields of the target products, 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2), obtained in Example 7 were 61.5% and 61.6%, respectively.

[0143] In Example 7, dioxane was used instead of acetonitrile as the solvent, and the yield was significantly lower than that in Example 1. This indicates that the polarity and solubility of acetonitrile are better matched with the reaction system, and it can more effectively dissolve the substrate, basic reagent and oxidant, providing a good homogeneous reaction environment, thereby obtaining a higher reaction yield.

[0144] Example 8 Example 8 describes the preparation of 5-bromo-4-chloro-3-acetoxyindole as shown in Formula III-1 and 5-bromo-6-chloro-3-acetoxyindole as shown in Formula III-2.

[0145] The reaction temperature in Example 8 was adjusted to 55 °C, while the remaining reaction conditions were the same as in Example 1. The yields of the target products obtained in Example 8, namely 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2), were 51.2% and 50.4%, respectively.

[0146] Comparing the results of Examples 1 (reaction temperature 35 °C), 8 (reaction temperature 55 °C), and 3 (reaction temperature 25 °C), all other reaction conditions were identical. The results showed that at a reaction temperature of 35 °C, the yields of the two isomers, 5-bromo-4-chloro-3-acetoxyindole (Formula III-1) and 5-bromo-6-chloro-3-acetoxyindole (Formula III-2), were 83.0% and 85.1%, respectively; when the reaction temperature decreased to 25 °C, the yields of the two isomers decreased to 67.2% and 63.3%, respectively; and when the reaction temperature increased to 55 °C, the yields significantly decreased to 51.2% and 50.4%. It is evident that excessively low temperatures (25 °C) lead to insufficient reaction rates and incomplete substrate conversion; while excessively high temperatures (55 °C) may accelerate the thermal decomposition of the high-valent iodine oxidant DIB, reducing the effective oxidant concentration. Simultaneously, side reactions such as polymerization and over-oxidation of the indole ring or reaction intermediates are prone to occur, reducing reaction selectivity. Therefore, 35 °C is the optimal reaction temperature for this oxidative acetoxylation reaction, ensuring a sufficient reaction rate while avoiding thermal decomposition and side reactions, thus achieving the highest yield.

[0147] Example 9 Example 9 describes the preparation of N-acetyl-5-bromo-4-chloro-3-acetoxyindole (Formula IV-1) and N-acetyl-5-bromo-6-chloro-3-acetoxyindole (Formula IV-2) via an N-acetylation reaction. Reaction conditions: acetylation reagent was acetic anhydride, catalyst was DMAP, second solvent was tetrahydrofuran, the molar ratio of 5-bromo-chloro-3-acetoxyindole, acetic anhydride, and DMAP was 1:10:2, the reaction temperature was the reflux temperature of tetrahydrofuran, and the reaction time was 3 hours.

[0148] (9-1) Preparation of N-acetyl-5-bromo-4-chloro-3-acetoxyindole In a round-bottom flask, 2.89 g (0.01 mol, 1.0 eq) of 5-bromo-4-chloro-3-acetoxyindole (Formula III-1), 150 mL of tetrahydrofuran, 10.21 g (0.1 mol, 10.0 eq) of acetic anhydride, and 2.44 g (0.02 mol, 2.0 eq) of DMAP were added. The resulting reaction mixture was heated under reflux for 3 hours. The solvent was evaporated, and 25 mL of ethyl acetate was added. The resulting suspension was stirred rapidly for 30 minutes, filtered, and the filter cake was collected to give N-acetyl-5-bromo-4-chloro-3-acetoxyindole solid (Formula IV-1) in 68.9% yield. The obtained N-acetyl-5-bromo-4-chloro-3-acetoxyindole solid was yellow.

[0149] (9-2) Preparation of N-acetyl-5-bromo-6-chloro-3-acetoxyindole Under the same reaction conditions as (9-1), the starting material was replaced with 5-bromo-6-chloro-3-acetoxyindole (2.89 g, 0.01 mol, 1.0 eq) as shown in Formula III-2 to obtain N-acetyl-5-bromo-6-chloro-3-acetoxyindole solid as shown in Formula IV-2, in a yield of 69.2%. The obtained N-acetyl-5-bromo-6-chloro-3-acetoxyindole solid was yellow.

[0150] In Example 9, an acetic anhydride / DMAP catalytic system was used with tetrahydrofuran as solvent, and the reaction was carried out at reflux temperature (approximately 66 °C) for 3 hours. The N-acetylation yields of the two intermediates, 5-bromo-4-chloro-3-acetoxyindole and 5-bromo-6-chloro-3-acetoxyindole, reached 68.9% and 69.2%, respectively. After the reaction, only the solvent needed to be evaporated, and the product could be obtained by crystallization with ethyl acetate without the need for column chromatography purification, making the operation simple and efficient.

[0151] Example 10 Example 10 describes the preparation of N-acetyl-5-bromo-4-chloro-3-indolone (Formula I-1) and N-acetyl-5-bromo-6-chloro-3-indolone (Formula I-2) via selective hydrolysis. Reaction conditions: 80% sulfuric acid as the acidic reagent, reaction temperature of 25 °C, and reaction time of 1 hour.

[0152] (10-1) Preparation of N-acetyl-5-bromo-4-chloro-3-indolone In a round-bottom flask, 35.6 mmol of N-acetyl-5-bromo-4-chloro-3-acetoxyindole (Formula IV-1) and 250 mL of 80% sulfuric acid were added. The resulting purplish-black solution was stirred at 25 °C for 1 hour. The reaction solution was then slowly poured into 300 g of ice water, precipitating N-acetyl-5-bromo-4-chloro-3-indole ketone solid (Formula I-1) in a yield of 88.0%. The obtained N-acetyl-5-bromo-4-chloro-3-indole ketone solid was brown.

[0153] The structure of the product obtained in this embodiment was verified by proton NMR spectroscopy, carbon NMR spectroscopy, and electrospray ionization mass spectrometry. The characterization experimental data are shown in Table 1, and the corresponding proton NMR spectrum, carbon NMR spectrum, and electrospray ionization mass spectrometry spectrum are shown in Table 1. Figures 5 to 7 As shown. This embodiment has verified that the compound N-acetyl-5-bromo-4-chloro-3-indolone, represented by Formula I-1, was obtained.

[0154] (10-2) Preparation of N-acetyl-5-bromo-6-chloro-3-indolone Under the same reaction conditions as (10-1), the starting material was replaced with N-acetyl-5-bromo-6-chloro-3-acetoxyindole (35.6 mol) as shown in Formula IV-2 to obtain N-acetyl-5-bromo-6-chloro-3-indolone solid as shown in Formula I-2, in yield of 89.0%. The obtained N-acetyl-5-bromo-6-chloro-3-indolone solid was brown.

[0155] The structure of the product obtained in this embodiment was verified by proton NMR spectroscopy, carbon NMR spectroscopy, and electrospray ionization mass spectrometry. The characterization experimental data are shown in Table 1, and the corresponding proton NMR spectrum, carbon NMR spectrum, and electrospray ionization mass spectrometry spectrum are shown in Table 1. Figures 8 to 10 As shown. This embodiment has verified that the compound N-acetyl-5-bromo-6-chloro-3-indolone, represented by Formula I-2, was obtained.

[0156] Table 1. 1H NMR spectra of the compounds shown in Formula I-1 and Formula I-2 ( 1 H NMR, carbon nuclear magnetic resonance (NMR) 13 C10 NMR and electrospray ionization high-resolution mass spectrometry (ESI-HRMS) data

[0157] In Example 10, using 80% sulfuric acid as the acidic medium and stirring at room temperature for 1 hour, the acetyl groups on oxygen atoms were selectively hydrolyzed. The yields of the two target products, N-acetyl-5-bromo-4-chloro-3-indolone and N-acetyl-5-bromo-6-chloro-3-indolone, were as high as 88.0% and 89.0%, respectively. The product structures were verified by 1H NMR, 1C NMR, and electrospray ionization high-resolution mass spectrometry. Moreover, the post-reaction processing only requires pouring the reaction solution into ice water for crystallization, without the need for column chromatography purification, fully demonstrating the outstanding advantages of high selectivity, high yield, and simple operation.

[0158] The target products N-acetyl-5-bromo-4-chloro-3-indolone and N-acetyl-5-bromo-6-chloro-3-indolone prepared in Example 10 of this invention were prepared using, respectively, [the following methods were employed]. 1 H NMR, 13 The structure was characterized by C NMR and ESI-HRMS.

[0159] Combination Figures 5 to 7 As shown, for the target product N-acetyl-5-bromo-4-chloro-3-indolone, 11H NMR (500MHz, CDCl3) characterization results show that δ8.43 (d, J=8.4 Hz, 1H) and δ7.82 (d, J=8.4 Hz, 1H) are characteristic doublets of two ortho aromatic hydrogens (6-H and 7-H) on the indole benzene ring, and the coupling constant J=8.4 Hz indicates that they are ortho, which is consistent with the 5-bromo-4-chloro substitution mode; δ4.36 (s, 2H) belongs to the methylene (-CH2-) of the indole ring; δ2.33 (s, 3H) is the proton signal of N-acetylmethyl (-COCH3); the chemical shifts, peak shapes, integrals and coupling constants of each hydrogen are consistent with the target structure. 13 C10 NMR (125 MHz, CDCl3) characterization results showed that δ190.6 was the carbonyl (C=O) signal of indoleone; δ168.1 was the carbonyl (C=O) signal of N-acetylamide; δ154.1, 140.9, 131.8, 122.8, 119.0, and 117.8 were attributed to quaternary and tertiary carbons of the benzene ring and fused ring moieties; δ56.4 was the carbon signal of methylene (-CH2-); and δ24.3 was the carbon signal of acetylmethyl. The number of carbon signals corresponded to the number of chemically inequivalent carbon atoms in the molecule, and the chemical shifts matched the target structure. ESI-HRMS characterization results showed that the measured deprotonated quasi-molecular ion peak was 285.9281 [MH]. - The molecular weight is highly consistent with the theoretically accurate molecular weight, confirming the molecular formula as C. 10 H7BrClNO2.

[0160] Combination Figures 8 to 10 As shown, for the target product N-acetyl-5-bromo-6-chloro-3-indolone, 1 1H NMR (500MHz, CDCl3) characterization results showed that δ8.75 (s, 1H) and δ7.97 (s, 1H) were characteristic singlets of two isolated meta aromatic hydrogens (4-H and 7-H) on the benzene ring, with no obvious ortho-coupled splitting, consistent with the 5-bromo-6-chloro substitution mode; δ4.33 (s, 2H) was the proton signal of the indole cyclic methylene (-CH2-); δ2.32 (s, 3H) was the proton signal of the N-acetylmethyl (-COCH3); the assignment of each hydrogen signal was clear and consistent with the target structure. 13C10 NMR (125 MHz, CDCl3) characterization results showed that δ192.1 was the carbonyl (C=O) signal of indoleone; δ168.1 was the carbonyl (C=O) signal of N-acetylamide; δ152.4, 143.6, 128.0, 124.6, 120.3, and 118.0 were carbon signals of the benzene ring; δ56.3 was the carbon signal of methylene (-CH2-); and δ24.1 was the carbon signal of acetylmethyl. The number of carbon signals and chemical shifts were highly consistent with the structure of the target product. ESI-HRMS characterization results showed that the quasi-molecular ion peak was measured at m / z = 285.9278 [MH]. - The precise molecular weight matches the theoretical calculation, confirming the molecular formula as C. 10 H7BrClNO2.

[0161] In summary, the results of proton NMR, carbon NMR, and high-resolution mass spectrometry corroborate each other, indicating that the products obtained in Example 10 are N-acetyl-5-bromo-4-chloro-3-indolone and N-acetyl-5-bromo-6-chloro-3-indolone, respectively, with correct structures and purity that meet the requirements for subsequent applications.

[0162] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0163] The embodiments described above are merely illustrative of several implementation methods of this application, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the invention patent. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of this application, and these all fall within the protection scope of this application. Therefore, the protection scope of this patent application should be determined by the appended claims.

Claims

1. A method for preparing N-acetyl-5-bromo-chloro-3-indolone, characterized in that, include: Under alkaline conditions, 5-bromo-chloroindole undergoes an oxidation reaction to introduce an acetoxy group at the 3-position of the indole ring, yielding 5-bromo-chloro-3-acetoxyindole; wherein the chlorine atom is located on the benzene ring of the indole ring; 5-Bromo-chloro-3-acetoxyindole undergoes an N-acetylation reaction to introduce an acetyl group onto the nitrogen atom at the 1-position of the indole ring, yielding N-acetyl-5-bromo-chloro-3-acetoxyindole; Under acidic conditions, N-acetyl-5-bromo-chloro-3-acetoxyindole undergoes selective hydrolysis to remove the acetyl group from the oxygen atom, yielding N-acetyl-5-bromo-chloro-3-indolone.

2. The preparation method according to claim 1, characterized in that, The chlorine atom is located at position 4 or 6 of the indole ring.

3. The preparation method according to claim 1, characterized in that, The oxidizing agent used in the oxidation reaction is one of iodophenylacetic acid, Dys-Martin reagent; and / or In the oxidation reaction, the alkaline reagent used under alkaline conditions is one of potassium hydroxide, sodium hydroxide, 1,8-diazabicyclo[5.4.0]undec-7-ene, 1,5-diazabicyclo[4.3.0]non-5-ene, or triethylamine; and / or The oxidation reaction is carried out in a first solvent, which is one or more of acetonitrile, dioxane, N,N-dimethylformamide, and N,N-dimethylacetamide; and / or The oxidation reaction is carried out at a temperature of 10~80 ℃; or, the oxidation reaction is carried out at a temperature of 35 ℃.

4. The preparation method according to claim 1, characterized in that, In the oxidation reaction, the molar ratio of 5-bromo-chloroindole, the basic reagent, and the oxidant is 1:(1.5~3):(1.2~2); or, In the oxidation reaction, potassium hydroxide is used as the alkaline reagent, iodophenyl diacetic acid is used as the oxidant, and the molar ratio of 5-bromo-chloroindole, potassium hydroxide and iodophenyl diacetic acid is 1:2:1.

5.

5. The preparation method according to any one of claims 1 to 4, characterized in that, The acetylation reagent used in the N-acetylation reaction is one of acetic anhydride and acetyl chloride; and / or The N-acetylation reaction is carried out at a temperature of 10–100 °C; or, the N-acetylation reaction is carried out in a second solvent at a temperature equal to the reflux temperature of the second solvent; or, the N-acetylation reaction is carried out at a temperature of 66 °C; and / or The N-acetylation reaction is carried out in the presence of a catalyst, namely 4-dimethylaminopyridine; and / or The N-acetylation reaction is carried out in a second solvent, which is tetrahydrofuran.

6. The preparation method according to any one of claims 1 to 4, characterized in that, The N-acetylation reaction is carried out in the presence of the catalyst 4-dimethylaminopyridine, and the molar ratio of 5-bromo-chloro-3-acetoxyindole, the acetylation reagent, and 4-dimethylaminopyridine is 1:(5–15):(0.5–3); or, The N-acetylation reaction is carried out in the presence of the catalyst 4-dimethylaminopyridine, with acetic anhydride as the acetylation reagent, and the molar ratio of 5-bromo-chloro-3-acetoxyindole, acetic anhydride and 4-dimethylaminopyridine is 1:10:

2.

7. The preparation method according to any one of claims 1 to 4, characterized in that, In the selective hydrolysis reaction, the acidic reagent used under acidic conditions is 10%~85% sulfuric acid; or, in the selective hydrolysis reaction, the acidic reagent used under acidic conditions is 80% sulfuric acid; and / or The reaction temperature of the selective hydrolysis reaction is 10~50 ℃; or, the reaction temperature of the selective hydrolysis reaction is 25 ℃.

8. An N-acetyl-5-bromo-chloro-3-indolone, characterized in that, It is prepared by the preparation method according to any one of claims 1 to 7.

9. A 5-bromo-chloro-3-indole glycoside, characterized in that, It was prepared by using N-acetyl-5-bromo-chloro-3-indolone as described in claim 8 as a raw material, via glycosylation and deprotection reactions.

10. A chromogenic culture medium, characterized in that, The 5-bromo-chloro-3-indole glycoside as described in claim 9 was used as the chromogenic substrate.

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