A method for preparing L-galactose

Abstract

The invention discloses a method for preparing L-galactose, comprising: mixing pectin with dilute sulfuric acid, hydrolyzing to generate D-galacturonic acid; adding CaCO3 for neutralization, filtering to remove CaSO4, and preheating a cyclization system with the neutralization heat; adding lactic acid and acetic acid to the hydrolyzate, heating to 88-92°C at a rate of 1-2°C / min, and maintaining the temperature for 3-4 hours to generate L-galactonic acid-1,4-lactone; concentrating the cyclization solution, adding L-tartaric acid, adjusting the pH to 8.5-9.0, and then reducing the lactone with NaBH4; and recrystallizing with ethanol to obtain L-galactose crystals. The method promotes cyclization while stabilizing the pH through the synergistic effect of lactic acid and acetic acid, with a lactone yield exceeding 75%; utilizing L-tartaric acid to fix the lactone conformation by multiple hydrogen bonds, forcing NaBH4 to attack the carbonyl carbon from the Re face, and achieving an L-configuration selectivity of 97% ([α]D 20 =+72.5°), eliminating the need for chromatographic separation steps.

Description

Technical Field

[0001] The present invention relates to the technical field of L-galactose preparation, in particular to a method for preparing L-galactose. Background Art

[0002] L-galactose is a rare monosaccharide with important physiological activities and is widely used in food, medicine, and cosmetics. However, its efficient preparation still faces numerous technical bottlenecks, mainly focusing on low raw material utilization efficiency, difficulty in controlling side reactions, poor chiral selectivity, and excessive energy consumption. Existing processes for pectin hydrolysis, cyclization, and reduction have many drawbacks.

[0003] For example, while traditional sulfuric acid-catalyzed acidolysis effectively cleaves pectin glycosidic bonds, acid concentrations exceeding 1.5% can easily trigger dehydration of D-galacturonic acid, producing carbonized products such as 5-hydroxymethylfurfural (HMF), resulting in a drop in the yield of the target product to below 60%. For example, patent CN104086667B utilizes an ultrasound-assisted acid method, which shortens extraction time but still requires strict acid control to prevent molecular chain degradation. This method does not address the side reactions caused by excessive local H⁺ concentrations. Conventional sodium borohydride reduction lacks an effective chiral induction mechanism, resulting in a mixed L / D configuration in the product, with a ratio of approximately 1:1. Purification requires chromatographic separation, a cumbersome process and yield losses exceeding 30%. While prior art attempts have employed chiral ligands such as binaphthol for targeted regulation, the complex synthesis and difficult recovery of the ligands hinder industrial application. Although patented technologies such as CN104086667B improve pectin extraction efficiency through ultrasound assistance, they still have the following disadvantages: (1) no buffer system is designed to address the carbonization side reaction, and the HMF content is higher than 10%; (2) the cyclization stage relies on external temperature control, resulting in high equipment investment; (3) there is a lack of stereoselective control methods, and the product optical purity is insufficient. Summary of the Invention

[0004] The object of the present invention is to provide a method for preparing L-galactose. The present invention utilizes the synergistic effect of lactic acid and acetic acid to effectively control the conversion of D-galacturonic acid into cyclization to generate L-galactonic acid-1,4-lactone, and then utilizes the two hydroxyl groups (-OH) and two carboxylic acid groups (-COOH) of L-tartaric acid to form multiple hydrogen bonds with the carbonyl oxygen and adjacent hydroxyl groups of the lactone, thereby fixing the molecular conformation of the lactone, forcing NaBH4 to attack the carbonyl carbon from the Re surface, hindering the formation of the D-configuration transition state, and improving the yield of the L-configuration.

[0005] To solve this technical problem, the technical solution of the present invention is: a method for preparing L-galactose, comprising the following steps:

[0006] S1, lactic acid and acetic acid synergistically catalyze the cyclization of D-galacturonic acid to produce L-galactonic acid-1,4-lactone;

[0007] Lactic acid for stabilizing the carboxylic acid form of D-galacturonic acid and acetic acid for promoting condensation of the C1 carboxylic acid and the C4 hydroxyl group in the D-galacturonic acid molecule through hydrophobic interaction are added to a solution containing D-galacturonic acid;

[0008] Raise the temperature to 88-92°C and keep warm for 3-4 hours; the reaction is complete;

[0009] S2, concentrating the reaction solution obtained in S1;

[0010] S3. Add L-tartaric acid to the concentrated solution obtained in S2. L-tartaric acid fixes the conformation of L-galactonic acid-1,4-lactone through a hydrogen bond network. NaBH4 directionally reduces L-galactonic acid-1,4-lactone to obtain L-galactose.

[0011] The method for catalytically hydrolyzing pectin to produce D-galacturonic acid preferably comprises the following steps:

[0012] S11, mixing pectin and dilute sulfuric acid, heating to 90-100°C, stirring, and reacting for 3-5 hours;

[0013] S12. After the yield of D-galacturonic acid exceeds 80% by HPLC monitoring, CaCO3 is added in batches to neutralize the sulfuric acid in the reaction solution until the pH of the solution is 4.8 to 5.2, and the neutralization heat is released, and CaSO4 is filtered out.

[0014] In step S11 of the present invention, the reaction must be carried out at 90°C-100°C for 3-5 hours with continuous stirring to ensure that the pectin is fully hydrolyzed into D-galacturonic acid. The temperature must be maintained until the reaction is completed and the temperature cannot be naturally cooled.

[0015] The mass fraction of dilute sulfuric acid in step S11 is preferably 0.8%-1.2%. The present invention utilizes dilute sulfuric acid to dissociate into H + and SO4 2- . H + As a proton acid, it preferentially attacks the oxygen atom of the α-1,4-glycosidic bond in the pectin molecule, causing the glycosidic bond to break and generate D-galacturonic acid and short-chain oligosaccharides. The present invention limits the dehydration reaction, i.e., the side reaction, caused by excessive H⁺ by controlling the mass fraction of dilute sulfuric acid to 0.8%-1.2%.

[0016] The preferred solid-liquid ratio of pectin to dilute sulfuric acid is 1 g:10 ml.

[0017] Preferably, the concentration of lactic acid in step S1 is 0.5M-1M, and the volume ratio of lactic acid to acetic acid is 3:1.

[0018] The present invention strictly controls the volume ratio of lactic acid to acetic acid, strictly maintains the pH at 4.0-5.0, utilizes lactic acid to stabilize the carboxylic acid form and cooperates with acetic acid to control the pH and promote cyclization, thereby improving the yield.

[0019] Preferably, in step S1, the initial temperature of the preparation process of lactic acid-acetic acid synergistically catalyzing the cyclization of D-galacturonic acid to produce L-galactonic acid-1,4-lactone increases due to the absorption of the neutralization heat of CaCO3, and then rises to 90°C at a rate of 1-2°C / min.

[0020] The preferred process conditions for the directed reduction of L-galactonic acid-1,4-lactone by NaBH4 in step S3 to obtain L-galactose are as follows:

[0021] S31, adjusting the pH of the system to 8.5 to 9.0, adding NaBH4, and the molar ratio of NaBH4 to L-galactonic acid-1,4-lactone to be 1.2:1;

[0022] S32, recrystallize from ethanol to obtain L-galactose.

[0023] In the present invention, NaBH4 is excessive to ensure that the lactone carbonyl is fully reduced.

[0024] The present invention utilizes the synergistic effect of multiple hydrogen bonds and steric hindrance, and the ratio of L-configuration to D-configuration reaches 88:12, while it is 50:50 when no inducer is added. The optical purity [α]D 20 = +72.5°, approaching the level of enzymatic methods. Furthermore, the present invention utilizes L-tartaric acid, which is highly water-soluble (solubility 139 g / 100 mL at 20°C). The reduction product is retained in the mother liquor after recrystallization from ethanol, eliminating the need for separate isolation. In the present invention, L-tartaric acid forms a hydrogen bond network, with carboxyl groups forming hydrogen bonds with lactone carbonyl groups, such as -COOH…O=C, and hydroxyl groups forming hydrogen bonds with adjacent lactone hydroxyl groups, such as -OH…HO-C3. This stabilizes the lactone conformation, sterically shielding the Si surface while allowing NaBH4 to attack from the Re surface, ultimately regulating the L-galactose ratio to 97%.

[0025] Preferably, the molar ratio of L-tartaric acid to L-galactonic acid-1,4-lactone in step S3 is 0.003:1-0.008:1.

[0026] By adopting the above technical solution, the beneficial effects of the present invention are:

[0027] The present invention provides a method for preparing L-galactose. The method comprises the following steps: firstly, lactic acid and acetic acid are used to synergistically catalyze the cyclization of D-galacturonic acid to generate L-galactonic acid-1,4-lactone; lactic acid that stabilizes the carboxylic acid form of D-galacturonic acid and acetic acid that promotes the condensation of C1 carboxylic acid and C4 hydroxyl groups in the D-galacturonic acid molecule through hydrophobic interaction are added to a solution containing D-galacturonic acid; the lactic acid stabilizes the pH at 4.0-5.0, inhibiting the strong acid ring-opening side reaction; simultaneously, the hydrophobic effect of acetic acid is enhanced, and the non-polar methyl group (CH3-) of acetic acid reduces the polarity of the reaction medium, promoting the aggregation of the hydrophobic region of D-galacturonic acid, driving the condensation of C1 carboxyl group and C4 hydroxyl group in the molecule, reducing the cyclization energy barrier, shortening the reaction time, and promoting intramolecular esterification; through the synergistic mechanism of the double acid, the yield of L-galacturonic acid-1,4-lactone is significantly higher than that of the single acid system; the temperature is raised to 88°C-92°C, kept warm for 3-4 hours, and the reaction is terminated;

[0028] The present invention concentrates the reaction solution obtained by S1 and directly enters the reduction reaction, thereby reducing purification losses and avoiding the use of organic solvents;

[0029] L-tartaric acid is added to the concentrate to create a chiral environment, and sodium borohydride is used for stereoselective reduction to prepare L-galactose. The specific implementation method is as follows:

[0030] The two hydroxyl groups (-OH) and two carboxylic acid groups (-COOH) of L-tartaric acid can form multiple hydrogen bonds with the carbonyl oxygen and adjacent hydroxyl groups of the lactone, fixing the conformation of the lactone molecule and forcing NaBH4 to attack the carbonyl carbon from a specific Re plane. The rigid four-carbon skeleton of L-tartaric acid hinders the formation of the D-configuration transition state and inhibits the side reaction pathway. This enables the directed reduction of L-galactonic acid-1,4-lactone by NaBH4 to obtain L-galactose, thereby achieving the regulation of the galactose configuration. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is the infrared spectrum of L-galactose obtained in Example 1;

[0032] Figure 2 HPLC analysis of Examples 1-5 and Comparative Examples 1-3 of the present invention and standard L-galactose. DETAILED DESCRIPTION

[0033] In order to further explain the technical solution of the present invention, the present invention is described in detail below through specific embodiments.

[0034] Example 1

[0035] This example discloses a method for preparing L-galactose. The raw materials and equipment used in this example are as follows:

[0036] citrus pectin (purity ≥85%, Sigma-Aldrich);

[0037] 0.8% dilute sulfuric acid (analytical grade), lactic acid (food grade), acetic acid (analytical grade);

[0038] L-Tartaric acid (analytical grade), sodium borohydride (98%), ethanol (technical grade);

[0039] Constant temperature reactor, magnetic stirrer, HPLC system (Agilent 1260), polarimeter (Rudolph Autopol IV).

[0040] The following steps are involved:

[0041] S1, lactic acid and acetic acid synergistically catalyze the cyclization of D-galacturonic acid to produce L-galactonic acid-1,4-lactone;

[0042] In this embodiment, dilute sulfuric acid is used to catalyze the hydrolysis of pectin to produce D-galacturonic acid, which specifically includes the following steps:

[0043] S11, mix 10 g of pectin with 100 mL of 0.8% dilute sulfuric acid at a solid-liquid ratio of 1 g:10 mL, and add the mixture to a thermostatic reactor;

[0044] The temperature was raised to 95°C and the reaction was stirred at 300 rpm for 4 h. The production of D-galacturonic acid was monitored by HPLC;

[0045] S12. HPLC monitoring showed that the yield of D-galacturonic acid was 72% and the HMF content was 3.5%.

[0046] CaCO3 was added in batches to neutralize the sulfuric acid in the reaction solution until the pH of the solution reached 5.0, and the neutralization heat was released, and the temperature of the hydrolyzed solution rose to 55°C; CaSO4 was removed by filtration.

[0047] Lactic acid for stabilizing the carboxylic acid form of D-galacturonic acid and acetic acid for promoting condensation of the C1 carboxylic acid and the C4 hydroxyl group in the D-galacturonic acid molecule through hydrophobic interaction are added to a solution containing D-galacturonic acid;

[0048] Lactic acid and acetic acid were added to the hydrolyzate at a concentration of 0.8 M, with a volume ratio of 3:1.

[0049] The reaction mixture was heated to 90°C at a rate of 1°C / min and kept warm for 3 hours. The lactone yield was 68% as determined by HPLC.

[0050] S2, concentrate the reaction solution obtained in S1; the volume is 1 / 5 of the original solution, and it appears as a continuous thin stream when poured, with a viscosity of about 500 to 1000 cP.

[0051] S3, adding L-tartaric acid to the concentrated solution obtained in S2, with the molar ratio of L-tartaric acid to L-galactonic acid-1,4-lactone being 0.005:1, and adjusting the pH to 8.5;

[0052] NaBH4 was added, with the molar ratio of L-galactonic acid-1,4-lactone to NaBH4 being 1:1.2, and the reaction was carried out at 40°C for 2 hours; L-tartaric acid fixed the conformation of L-galactonic acid-1,4-lactone through a hydrogen bond network, and NaBH4 directionally reduced L-galactonic acid-1,4-lactone to obtain L-galactose.

[0053] For ethanol recrystallization, the ratio of the mass (g) of the crude product to be recrystallized to the volume (mL) of ethanol solvent was 1:10, that is, 10 mL of ethanol was used for every 1 gram of crude product for recrystallization.

[0054] The optical purity of the product obtained in this example is [α]D 20 =+72.5° (c=1, H2O, chiral HPLC detection conditions: Chiralpak AD-H column, mobile phase n-hexane / isopropanol = 70:30 (v / v), flow rate 1.0 mL / min); chiral HPLC analysis showed that L-galactose accounted for 95% and D-configuration accounted for 5%.

[0055] In this embodiment, the total yield of L-galactose is about 46.4%, and the optical purity is 95%. The total yield of L-galactose = D-galacturonic acid yield * lactone yield * L-galactose ratio.

[0056] Example 2

[0057] The main differences between this embodiment and embodiment 1 are: the sulfuric acid concentration is adjusted to 1.2%, and the reaction time is shortened to 3 hours.

[0058] Example 3

[0059] The main difference between this embodiment and embodiment 1 is that the stirring rate in step S11 of using dilute sulfuric acid to catalyze the hydrolysis of pectin to produce D-galacturonic acid is 400 rpm, and other conditions are the same as those in embodiment 1.

[0060] Example 4

[0061] The main difference between this embodiment and embodiment 1 is that the amount of L-tartaric acid added is adjusted so that the molar ratio of L-tartaric acid to lactone is 0.003:1.

[0062] The optical purity of the product obtained in this example is [α]D 20 =+70.1°, requiring secondary recrystallization to increase to 95%.

[0063] Example 5

[0064] The main difference between this embodiment and embodiment 1 is:

[0065] In step S11 of using dilute sulfuric acid to catalyze the hydrolysis of pectin to produce D-galacturonic acid, the stirring rate is 150 rpm, and other conditions are the same as those in Example 1.

[0066] Table 1 Sulfuric acid concentration and product status in Examples 1 to 5

[0067]

[0068] Comparative Example 1

[0069] The main difference between this comparative example and Example 1 is that an equal amount of lactic acid was used alone, and acetic acid was not used.

[0070] The remaining process steps and parameters were the same as those in Example 1. The obtained products are shown in Table 2.

[0071] Table 2 Comparison of the products obtained in Comparative Example 1 and Example 1

[0072]

[0073] As shown in Table 2, Comparative Example 1 lacks the hydrophobic effect of acetic acid, the intramolecular cyclization rate decreases, and the corresponding yield is significantly reduced.

[0074] Comparative Example 2

[0075] The main difference between this comparative example and comparative example 1 is:

[0076] In this comparative example, acetic acid alone was used with lactic acid of equal molar concentration as in Example 1, but acetic acid was not used. The remaining process steps and parameters were the same as in Example 1. The obtained products are shown in Table 3.

[0077] Table 3 Comparison of the products obtained in Comparative Example 2 and Example 1

[0078]

[0079] As shown in Table 3, in Comparative Example 2, no lactic acid was used, and the pH was out of control, resulting in a dehydration side reaction with an HMF content of 9.1%, a surge in HMF byproducts, and a decrease in lactone purity.

[0080] Comparative Example 3

[0081] The main difference between this comparative example and Example 1 is that L-tartaric acid was not used, that is, no chiral induction was performed; the remaining steps and parameters were the same as those in Example 1. The obtained products are shown in Table 3.

[0082] The product obtained in Comparative Example 3 was analyzed by chiral HPLC. The L-galactose content was 55%, the D-configuration was 45%, and the optical purity was [α]D 20= +41.3° (c=1, H2O). Although there is a certain L-preference, it cannot directly meet the pharmaceutical purity (>98%) and still requires chiral separation.

[0083] Table 4 Comparison of the products obtained in Comparative Example 3 and Example 1

[0084]

[0085] As shown in Table 4, when there is no chiral inducing agent in Comparative Example 3, the reduction reaction has no stereoselectivity and requires further chromatographic separation and purification, and the total yield plummets.

[0086] The infrared spectrum of L-galactose obtained in Example 1 is as follows Figure 1 As shown, at 3420 cm -1 A broad and strong absorption peak appears at 1075 cm, which is the stretching vibration characteristic of multiple hydroxyl groups (-OH) in the sugar molecule, proving that the product has a typical polyhydroxy structure of monosaccharide. -1 A sharp peak appears at 990 cm, which is the characteristic vibration peak of the COC ether bond in the pyranose ring, confirming that the product forms a six-membered ring structure, which is different from open-chain sugars. -1 and 920 cm -1 The double weak peak at 850 cm is the fingerprint characteristic of the sugar ring breathing vibration, which fully matches the standard L-galactose database spectrum. -1 The medium-strong peak at 1720 cm corresponds to the axial hydrogen vibration of the C1 position of the pyranose ring, which is the stereochemical characteristic peak of L-galactose. There are no abnormal peaks in the whole spectrum (such as 1720 cm -1 There is no carbonyl peak nearby), proving that the lactone group has been completely reduced and the purity of the product is in line with expectations.

[0087] Figure 2 For Example 1-5 and Comparative Example 1-3 product HPLC comparison diagram, L-galactose standard presents a sharp symmetrical single peak at 8.21 minutes, as a reference benchmark. The main peak retention time of Example 1 product is 8.20 minutes, which is completely overlapped with the standard, and the peak shape is symmetrical without shoulder peak, and the peak area accounts for 95.2%, indicating that the L- configuration purity is extremely high. The main peak retention time of Example 2 product is 8.22 minutes, but the peak width increases to 0.25 minutes, showing a slight decrease in stereoselectivity, which is consistent with its 92.1% L- configuration accounting. The product of Example 3 has a main peak at 8.19 minutes, but a trace by-product peak appears at 8.5 minutes, accounting for 1.3%, reflecting a small amount of impurities caused by insufficient stirring rate. Although the main peak of the product of Comparative Example 1 is still at 8.20 minutes, a clear dehydration by-product peak appears at 7.8 minutes, accounting for 15.2%, and the peak area is only 76.8% of Example 1. It can be seen that side reactions increase when acetic acid is lacking.

[0088] The product of Comparative Example 3 exhibited a bimodal structure: an L-galactose peak at 8.21 minutes and a D-galactose peak at 9.08 minutes. The two peaks were close in height, intuitively indicating that a racemic mixture was obtained without chiral induction.

[0089] Figure 2 The height of the middle curve directly reflects the product concentration. Example 1 has the highest peak, demonstrating the optimal yield. Comparative Examples 1 and 3, however, have significantly lower peaks, confirming yield losses due to process defects. The differences in peak positions across the samples clearly demonstrate the control effect of configurational selectivity.

[0090] The test conditions of the above HPLC are as follows:

[0091] Chromatographic column: Chiralpak AD-H (250 mm × 4.6 mm, 5 μm); mobile phase: n-hexane / isopropanol = 70:30 (v / v); flow rate: 1.0 mL / min; column temperature: 30°C; injection volume: 10 μL; sample concentration: 10 mg / mL, solvent: isopropanol; detector temperature: 35°C; run time: 15 min.

Claims

1. A method for preparing L-galactose, characterized in that: The following steps are involved: S1, lactic acid and acetic acid synergistically catalyze the cyclization of D-galacturonic acid to produce L-galactonic acid-1,4-lactone; Raise the temperature to 88°C to 92°C and keep warm for 3-4 hours; the reaction is complete; In step S1, the concentration of lactic acid is 0.5M-1M, and the volume ratio of lactic acid to acetic acid is 3:1; S2, concentrating the reaction solution obtained in S1; S3, adding L-tartaric acid to the concentrated solution obtained in S2, L-tartaric acid fixes the conformation of L-galactonic acid-1,4-lactone through a hydrogen bond network, and NaBH4 directionally reduces L-galactonic acid-1,4-lactone to obtain L-galactose; In step S3, the molar ratio of L-tartaric acid to L-galactonic acid-1,4-lactone is (0.003-0.008):1; The method for catalytically hydrolyzing pectin to produce D-galacturonic acid comprises the following steps: S11, mixing pectin and dilute sulfuric acid, heating to 90-100°C, stirring, and reacting for 3-5 hours; S12. After the yield of D-galacturonic acid exceeds 80% by HPLC monitoring, CaCO3 is added in batches to neutralize the sulfuric acid in the reaction solution until the pH of the solution is 4.8 to 5.2, and the neutralization heat is released, and CaSO4 is filtered out.

2. The preparation method according to claim 1, wherein: The mass fraction of dilute sulfuric acid in step S11 is 0.8%-1.2%.

3. The preparation method according to claim 1, wherein: The solid-liquid ratio of pectin to dilute sulfuric acid is 1g:10ml.

4. The preparation method according to claim 1, wherein: In step S1, the temperature is raised at a rate of 1-2°C / min during the preparation process of lactic acid-acetic acid synergistically catalyzing the cyclization of D-galacturonic acid to produce L-galactonic acid-1,4-lactone.

5. The preparation method according to claim 1, wherein: The process conditions for the directed reduction of L-galactonic acid-1,4-lactone by NaBH4 in step S3 to obtain L-galactose are as follows: S31, adjusting the pH of the system to 8.5 to 9.0, adding NaBH4, and the molar ratio of NaBH4 to L-galactonic acid-1,4-lactone to be 1.2:1; S32, recrystallize from ethanol to obtain L-galactose.

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