A method for producing multi-stage functionalized humus products using amino acid-based deep eutectic solvents

Abstract

This invention provides an integrated method for producing multi-stage functionalized humic products using an amino acid-based deep eutectic solvent, belonging to the field of high-value utilization of biomass resources. The method uses a specific amino acid combined with a weak acid to form an amino acid-based deep eutectic solvent, which is then used for ultrasonic pretreatment and controlled hydrothermal reaction of biomass raw materials. Through natural cooling combined with pH gradient fractional precipitation, high-molecular-weight humic acid A, suitable for soil improvement, and low-molecular-weight, highly water-soluble humic acid B, suitable for foliar fertilizer, are obtained. Rapid cooling and concentration yield a highly bioactive fulvic acid concentrate. This invention achieves the targeted conversion and precise separation of a single biomass raw material into multiple functional humic acid products, significantly increasing the nitrogen content of the products. Furthermore, the amino acid-based deep eutectic solvent can be efficiently recycled, combining high efficiency, environmental friendliness, and high value.

Description

Technical Field

[0001] This invention relates to the field of comprehensive utilization technology of biomass resources, and in particular to an intelligent and controllable integrated production method for converting biomass waste into a variety of high-value-added humic products. Background Technology

[0002] The main active substances in humic substances include humic acid and fulvic acid, and humic substances play a crucial role in soil remediation and plant growth promotion. Hydrothermal humification technology is an efficient method for the directional conversion of biomass raw materials (such as straw and sludge) into humic substances based on a subcritical hydrothermal environment. Its core lies in achieving the recrystallization of insoluble inorganic substances and the directional conversion of organic matter through the coordinated regulation of temperature and pressure and the optimization of mineralizing agents.

[0003] To improve the utilization efficiency and effectiveness of hydrothermal humic materials, accelerating the hydrothermal humification reaction efficiency of biomass and improving the quality of humic acid and fulvic acid are crucial factors. Chinese invention patent application CN 113563603A proposes a method for enhancing the hydrothermal humification of lignocellulosic biomass through acid-hydrothermal pretreatment. However, while the acid-alkali two-step method improves the humic acid conversion efficiency of lignocellulosic waste, the product is singular and the reaction temperature is high, resulting in huge energy consumption. Furthermore, the nitrogen content in the humic acid is low due to the limited nitrogen content in the raw materials. Chinese invention patent application CN 119371680A proposes a method for catalyzing the hydrothermal conversion of plant waste into mineral-like humic acid using iron-doped nitrogen-phosphorus modified montmorillonite. However, this technology involves catalyst preparation and functional regulation under high temperature and multiple media, making the process complex and costly, and difficult to meet the demands of modern agriculture for efficient and environmentally friendly fertilizers.

[0004] Deep eutectic solvents (DES), as green and efficient reaction solvents, have shown significant advantages in biomass conversion in recent years. Current research indicates that lignin in straw undergoes deep depolymerization at 120°C in an AA-DES system with alanine as a hydrogen donor. Patent CN113556776A uses DES to pretreat seaweed residue to reduce crude fiber for use in animal feed; however, its target product and process route differ significantly from this application, and it does not involve fine-grained fractionation and functional-oriented regulation of the product molecules. Currently, no technology can flexibly switch process parameters using the same set of raw materials to produce humic products with drastically different properties and functions, based on the specific needs of downstream agricultural applications (such as whether rapid action or improved physical structure is required). Summary of the Invention

[0006] The present invention aims to provide an integrated method that has a lower reaction temperature, lower energy consumption, higher nitrogen content in the product, and can stably obtain a variety of different products by controlling a small number of process parameter changes, thereby significantly improving the yield of humic acid and the nitrogen content of the product in biomass hydrothermal humification.

[0007] The present invention utilizes a core reaction system, employing amino acid-based deep eutectic solvent AA-DES as a multifunctional reaction medium, coupled with ultrasonic pretreatment and a controllable hydrothermal reaction, to form a stable and efficient core conversion unit. This AA-DES not only serves as a pretreatment agent and reaction medium, but its components also participate in product construction as reactants, particularly as nitrogen sources and functional group donors. Humic acid and fulvic acid are then obtained through two controllable product-directing pathways.

[0008] Based on this idea, the present invention provides a method for producing multi-stage functionalized humic products using an amino acid-based deep eutectic solvent, the method comprising the following steps:

[0009] (1) Preparation of amino acid-based deep eutectic solvent

[0010] Amino acids and organic acids are thoroughly mixed in a molar ratio of 1:1 to 1:3 to obtain an amino acid-based deep eutectic solvent, wherein the amino acid is selected from proline, alanine, or glycine, and the organic acid is selected from citric acid, malic acid, or lactic acid.

[0011] (2) Raw material pretreatment

[0012] After drying, crushing and sieving, biomass raw materials are obtained as biomass raw material powder. The amino acid-based deep eutectic solvent described in step (1) is mixed with the biomass raw material powder at a mass ratio of 1:5 to 1:10. After homogenization, pretreated material is obtained.

[0013] (3) Hydrothermal reaction

[0014] The pretreated material is subjected to hydrothermal treatment at 140~220℃;

[0015] (4.1) Humic acid was isolated and obtained

[0016] The product obtained in step (3) was naturally cooled, and after solid-liquid separation, the liquid phase was recovered for later use. The solid phase product was then subjected to alkaline extraction and fractional precipitation: first, the pH was adjusted to 3.0-4.0, and the first solid product, humic acid A, was obtained; then, the pH of the supernatant was adjusted to 1.0-2.0, and the second solid product, humic acid B, was obtained; or

[0017] (4.2) Fulvic acid was separated and obtained

[0018] The product obtained in step (3) is rapidly cooled, and after solid-liquid separation, the liquid phase is concentrated to obtain humic acid concentrate.

[0019] In this invention, particularly preferred combinations of amino acids and organic acids are proline and citric acid in a molar ratio of 1:1 to 1:3, or alanine and malic acid, or glycine and lactic acid. Experiments have confirmed that these three combinations of amino acids and organic acids, when used in subsequent hydrothermal humification, can synergistically increase the humic acid yield by 10%-30% and the nitrogen content in the product by more than 60%.

[0020] In this invention, the biomass raw materials are selected from one or more of crop straw, vegetable waste, and livestock manure, all of which are readily available in the art. After these raw materials are thoroughly mixed with an amino acid-based deep eutectic solvent, the conventional homogenization process involves treating the mixture at 40-60°C and an ultrasonic power of 200-500W for 30-60 minutes.

[0021] In this invention, the hydrothermal treatment in step (3) involves thoroughly mixing the pretreated material with deionized water at a solid-liquid ratio of 1:9 and then placing the mixture in a stainless steel high-pressure reactor. The mixture is then heated to 140-220°C in a sealed environment and maintained for 1-4 hours. Those skilled in the art can also adjust the temperature and time conditions required for the hydrothermal treatment as appropriate.

[0022] After the hydrothermal reaction is complete, technicians can choose different cooling conditions to obtain different products. Typically, to obtain humic acid, the solid phase in step (4.1) is extracted with alkali, and the resulting solid product is added to a mixed alkali solution at a solid-liquid ratio of 1:50~100 and mixed thoroughly. Subsequently, it is placed in a shaker and mixed thoroughly for 1 hour. The mixed alkali solution is a NaOH / Na4P2O7 mixed alkali solution with a molar ratio of 1:0~1:1 and a concentration of 0.1 mol / L.

[0023] Subsequently, different precipitates can be obtained by changing the pH value of the reaction system. Specifically, when the pH is controlled at 3.0-4.0, the separated precipitate yields humic acid A, with a number-average molecular weight greater than 2000 Da, which is typically used to prepare soil conditioners or slow-release fertilizer carriers that improve soil physical structure and increase cation exchange capacity. When the pH of the reaction system is further controlled to 1.0-2.0, the separated precipitate yields humic acid B, with a number-average molecular weight less than 2000 Da, which is typically used to prepare liquid fertilizers, biostimulants, or pesticide adjuvants for foliar spraying, drip irrigation, or seed coating.

[0024] If fulvic acid is desired, it can be obtained through rapid cooling (i.e., step (4.2)). For example, the product is cooled at a rate of 20-50 °C / min, followed by solid-liquid separation and concentration to obtain a fulvic acid concentrate with a fulvic acid content of not less than 40%, and the concentrate also contains organic acids and amino acid derivatives derived from amino acid-based deep eutectic solvents. These fulvic acid concentrates can be used to prepare water-soluble fertilizers or biostimulants that promote plant root growth, improve nutrient absorption efficiency, or alleviate abiotic stress.

[0025] In this invention, preferably, in step (4.1), the recovered liquid phase is subjected to vacuum distillation to recover the amino acid-based eutectic solvent component and reuse it in step (1). Vacuum distillation can be carried out at a temperature of 60~80℃ and a pressure of -0.08~-0.1MPa.

[0026] This invention is the first in the field of biomass humification to propose and realize a process that allows for switching between the production of completely different functional products by controlling a simple parameter (cooling rate), resulting in extremely high production flexibility. Through this graded process and parameter control, this invention clearly differentiates the traditionally ambiguous "humic acid" into several standardized products (including humic acid A, B, and fulvic acid concentrate) with specific molecular weight ranges, functional group compositions, and varying solubilities, and establishes quantifiable quality indicators for them, resulting in a high degree of product standardization.

[0027] This invention integrates DES chemistry, reaction engineering (cooling control), and separation science (pH fractionation) to solve the technical challenge of "targeted production" with a holistic solution. It can directly produce products that meet the needs of different markets through a single method (production process). Attached Figure Description

[0028] Figure 1 This is a process flow diagram of the method of the present invention;

[0029] Figure 2 Experimental diagrams illustrating the effects of different humic substances on promoting seed germination;

[0030] Figure 3 Peak segmentation analysis of N 1s spectra of humic acid B produced by different AA-DES combinations. Detailed Implementation

[0031] The following examples are used to explain the technical solutions of the present invention in a non-limiting manner.

[0032] Example 1 Preparation of humic acid

[0033] Weigh proline and citric acid in a molar ratio of 1:2 and mix them evenly to obtain an amino acid-based deep eutectic solvent.

[0034] Take corn stalks, dry, crush and sieve them, mix them with amino acid-based deep eutectic solvent at a weight ratio of 8:1, pre-treat them with ultrasound, and then hydrothermally react them in a reactor at 180°C for 2 hours.

[0035] The reactor was allowed to cool naturally to room temperature, and the solid and liquid phases were obtained by filtration. The obtained solid phase was added to the mixed alkaline solution at a solid-liquid ratio of 1:100 and mixed thoroughly. The mixed alkaline solution used was a NaOH / Na4P2O7 mixed alkaline solution with a molar ratio of 1:1, and the final concentrations of both NaOH and Na4P2O7 in the mixed alkaline solution were 0.5 mol / L.

[0036] Then, hydrochloric acid with a pH of 1-2 is added to the reaction system until the pH reaches 3.0-4.0. After the reaction is complete, the mixture is filtered to obtain the first precipitate. Then, acid is added to the liquid phase until the pH reaches 1.0-2.0. After the reaction is complete, the mixture is separated to obtain the second precipitate.

[0037] Example 2 Preparation of fulvic acid

[0038] The same batch of raw materials as in Example 1 were used, and the procedure was carried out in the same manner as in Example 1.

[0039] The difference lies in that after the hydrothermal reaction is completed, the reactor is placed in an ice-water bath for rapid cooling until the temperature of the reaction system is below 50°C. The liquid phase obtained by filtration is concentrated under reduced pressure at 60°C and -0.1 MPa to 1 / 5 of the original volume to obtain a concentrated solution.

[0040] The average molecular weights of the first precipitate and the second precipitate in Example 1 and the concentrate in Example 2 were determined by gel permeation chromatography (GPC) using sodium polystyrene sulfonate as a standard and 0.1 mol / L NaCl solution as the mobile phase. The results are as follows:

[0041] Table 1. Molecular weights of humic substances obtained from different pathways

[0042]

[0043] The results showed that by controlling the cooling conditions after the hydrothermal reaction, the mixed humic acid could be successfully separated into several components with significantly different molecular weights using a fractional precipitation process: humic acid A1 > humic acid B1 > fulvic acid concentrate > blank CK group.

[0044] A comparative experiment was conducted to investigate the effects of blank control, humic acid A1, humic acid B1, and concentrated fulvic acid on seed germination and growth.

[0045] Using Chinese cabbage as the experimental crop, a pot experiment was conducted to test its growth-promoting effect under the same conditions. Four treatments were set up: humic acid A1, humic acid B1, concentrated fulvic acid solution, and a blank control group (CK) without any additives. Each pot contained 2 kg of soil and was moistened with an appropriate amount of water to approximately 30% humidity. The HA used in the pot experiment was prepared as a liquid fertilizer by first dissolving it in an alkaline solution and then diluting it to a concentration of 0.1%. 10 mL of the diluted solution was sprayed onto the soil surface of each pot. Five Chinese cabbage seeds were evenly planted in the soil at a depth of approximately 1 cm in each pot. Throughout the growing season, the plants were sprayed with 50 mL of tap water every 3 days.

[0046] The results are as follows: Figure 3 As shown.

[0047] The results showed that on the 4th day of germination, the effect on promoting seed germination was: fulvic acid concentrate > humic acid B1 > humic acid A1. The fulvic acid concentrate had the most comprehensive and significant effect; this invention, through rapid cooling, locked in highly active small molecules, resulting in the best growth-promoting effect. Humic acid B1 exhibited excellent root-promoting properties, indicating that it contains a large number of small molecules and highly active components. It is noteworthy that humic acid A had a relatively small effect in the seed germination experiment, reflecting its large molecular weight and slow-acting characteristics, complementing the distinct functions and uses of humic acid B and fulvic acid.

[0048] Example 3: Effects of different AA-DES combinations on the functional properties of the product

[0049] The procedure was carried out in the same manner as in Example 1, except that different combinations of amino acids and organic acids were used: DES-1 (glycine-lactic acid in a molar ratio of 1:2), DES-2 (alanine-malic acid in a molar ratio of 1:2), DES-3 (proline-citric acid in a molar ratio of 1:2), and HA (without AA-DES, as a control). The method of Example 1 was repeated to obtain three different first precipitates and second precipitates.

[0050] The humic acid yield, nitrogen content, and functional group distribution of the three second precipitates were determined.

[0051] Humic acid yield = Mass of humic acid produced (g) / Mass of biomass feedstock (g) × 100%

[0052] Nitrogen content was determined using an elemental analyzer to measure total nitrogen content and X-ray photoelectron spectroscopy (XPS) to analyze the 1s spectrum of nitrogen (pyrrole nitrogen, amino nitrogen, quaternary ammonium nitrogen, etc.). The results are shown below:

[0053] Table 2. Humic acid B yield and elemental content of different AA-DES combinations

[0054]

[0055] Comparative Example 1: Traditional Single-Path Process

[0056] The same raw materials and hydrothermal conditions as in Example 1 were used. After the hydrothermal reaction was completed, the reactor was naturally cooled to room temperature, and the solid and liquid phases were obtained by filtration. The obtained solid phase was added to the mixed alkaline solution at a solid-liquid ratio of 1:100 and mixed thoroughly. After the reaction was complete, the pH of the reaction system was adjusted to 1-2 with acid, and then the precipitate sample was obtained by precipitation.

[0057] The obtained sample was tested and compared with the precipitates of Examples 1 and 2:

[0058] Organic matter retention rate: Calculated by measuring and comparing the change in total organic carbon in the soil before and after cultivation. Steps:

[0059] Initial value determination: Take the original soil sample without added humus and determine its background total organic carbon content.

[0060] Cultivation experiment: Different humus treatments were mixed evenly with soil according to the effective concentration in the seed experiment, and the moisture content was adjusted to the optimal level. The mixture was then cultivated indoors for 30 days under constant temperature and humidity conditions.

[0061] Endpoint value determination: After the culture is completed, the total organic carbon content of the culture soil is measured.

[0062] calculate:

[0063] Organic matter retention rate (%) = (C t -C0) / C a ×100%

[0064] C t Soil organic carbon content (g / kg) in the treatment groups after cultivation.

[0065] C0: Organic carbon content (g / kg) of the original soil (blank control group)

[0066] C a : Amount of organic carbon added from exogenous humus (g / kg)

[0067] Total organic carbon (TOC) was measured using a total organic carbon (TOC) analyzer.

[0068] Aggregate stability index: The wet sieving method was used for determination. Procedure:

[0069] Air-dried soil samples were pre-passed through a 2mm sieve, and agglomerates with a particle size of 1-2mm were collected as test samples. A certain amount (10g) of the 1-2mm agglomerates was placed on the top sieve (0.25mm sieve) of a wet sieve apparatus and vibrated up and down in water at a fixed frequency and amplitude for 30 minutes. After vibration, the agglomerates remaining on each sieve (2mm, 1mm, 0.5mm, 0.25mm) were rinsed into containers of known weight, dried at 60℃ to constant weight, and weighed.

[0070] R 0.25 (%) = Sum of weights of aggregates at all levels >0.25mm / Total weight of aggregates × 100%

[0071] Testing equipment: wet sieve analyzer / aggregate analyzer; sieve set: aperture sizes of 2mm, 1mm, 0.5mm, and 0.25mm; constant temperature drying oven; analytical balance.

[0072] Table 3. Comparison of properties between mixed humic acid and fractionated products from Example 1

[0073]

[0074] The results show that the mixed humic acid falls between humic acid A1 and B1 in terms of molecular weight distribution and functional group composition. In soil improvement applications, its ability to promote aggregate formation and retain organic matter for a long time is significantly lower than that of specially isolated humic acid A1 (37% and 51% lower, respectively).

[0075] It is evident that while traditional single-path processes can produce humic acid, their overall yield is lower than that achieved by this invention, and the product's properties are limited, failing to meet the demands of modern agriculture for specialized, high-efficiency fertilizers. The graded extraction process of this invention, through simple pH gradient control, successfully achieves functional differentiation of the products, yielding two high-value products with complementary properties and distinct advantages. This is precisely the inventive aspect of this invention compared to existing technologies.

[0076] Comparative Example 2: Using standard DES (choline chloride / urea)

[0077] Using the exact same biomass raw materials as in Example 1, the difference was that the eutectic solvent was a conventional choline chloride / urea molar ratio of 1:2. Humic acid A and humic acid B were prepared under identical process conditions (raw materials, ratios, hydrothermal conditions, and fractionation steps). The products were tested using the same method, and the results are shown in the table below.

[0078] Table 4. Comparison of properties between mixed humic acid and fractionated products from Example 1

[0079]

[0080] The results showed that conventional DES is relatively unstable under hydrothermal conditions and has decomposed, leading to a reduced recovery rate. The humic acid nitrogen content obtained from conventional DES is low (<34%), and its performance in soil improvement applications lags behind that of AA-DES products. Therefore, conventional DES cannot replace the specific AA-DES combination of this invention. The inventiveness of this invention lies not only in the use of DES, but also in the discovery that a specific combination of amino acids and weak acids can produce a synergistic effect in a hydrothermal humification system, achieving efficient product transformation and functional differentiation—a phenomenon not previously revealed in existing technologies.

[0081] The above data demonstrates that the specific AA-DES combination and process selected in this invention can stably and significantly improve the yield of humic acid and the nitrogen content of the products simultaneously, solving the problems of low product yield and insufficient nitrogen nutrients in humic acid. It successfully transforms traditional homogeneous humic acid into two leading products with well-defined molecular structures and precise functional positioning: humic acid A (high molecular weight, focusing on long-term soil structure improvement) and humic acid B (low molecular weight, high activity, focusing on foliar nutrition and biostimulation). This invention, using the same set of raw materials and core reactor, allows for flexible switching of process paths (natural cooling / rapid cooling) to produce functional product combinations suitable for different high-end agricultural scenarios, maximizing resource value. Furthermore, this invention confirms that the AA-DES used has superior high-temperature stability and recyclability (recovery rate > 86%) compared to conventional DES. By screening and optimizing amino acid / weak acid combinations, the specific chemical form of nitrogen in the products is directionally regulated. The glycine system is beneficial for retaining active amino groups, the alanine system promotes the formation of stable heterocyclic structures, while the proline system easily generates quaternary ammonium structures with high charge density. This discovery demonstrates that by selecting different AA-DES, humic acid products with different functions can be designed and produced at the molecular structure level, which is a key technological advancement that cannot be achieved by existing single processes or universal DES.

Claims

1. A method for producing multi-stage functionalized humus products using amino acid-based deep eutectic solvents, characterized by The method includes the following steps: (1) Preparation of amino acid-based deep eutectic solvent Amino acids and organic acids are thoroughly mixed in a molar ratio of 1:1 to 1:3 to obtain an amino acid-based deep eutectic solvent, wherein the amino acid is selected from proline, alanine, or glycine, and the organic acid is selected from citric acid, malic acid, or lactic acid. (2) Raw material pretreatment After drying, crushing and sieving, biomass raw materials are obtained as biomass raw material powder. The amino acid-based deep eutectic solvent described in step (1) is mixed with the biomass raw material powder at a mass ratio of 1:5 to 1:

10. After homogenization, pretreated material is obtained. (3) Hydrothermal reaction The pretreated material is subjected to hydrothermal treatment at 140~220℃; (4.1) Humic acid was isolated and obtained The product obtained in step (3) is naturally cooled, and after solid-liquid separation, the liquid phase is recovered for later use. The obtained solid phase product is subjected to alkaline extraction, and then acid is added to the reaction system until the pH value is 3.0-4.

0. The precipitate is separated to obtain the first solid product, humic acid A; then the pH value of the remaining supernatant is adjusted to 1.0-2.0, and the precipitate is separated to obtain the second solid product, humic acid B; or (4.2) Fulvic acid was separated and obtained The product obtained in step (3) is rapidly cooled, and after solid-liquid separation, the liquid phase is concentrated to obtain humic acid concentrate.

2. The method of claim 1, wherein In step (1), proline is thoroughly mixed with citric acid, or alanine with malic acid, or glycine with lactic acid in a molar ratio of 1:1 to 1:

3.

3. The method of claim 1, wherein The alkaline extraction in step (4.1) involves adding the obtained solid product to a mixed alkaline solution at a solid-liquid ratio of 1:50 to 100 and mixing thoroughly. Then, the mixture is placed in a shaker and mixed thoroughly for 1 hour. The mixed alkaline solution is a NaOH / Na4P2O7 mixed alkaline solution with a molar ratio of 1:0 to 1:1 and a concentration of 0.1 mol / L.

4. The method of claim 1, wherein The method further includes step (4.1), in which the recovered liquid phase is subjected to vacuum distillation to recover the amino acid-based deep eutectic solvent component and reuse it in step (1).

5. The method according to claim 4, characterized in that... The vacuum distillation described in step (4.1) is carried out at a temperature of 60~80℃ and a pressure of -0.08~-0.1MPa.

6. The method according to claim 1, characterized in that... The acid in step (4.1) is selected from HCl and / or H2SO4 with a pH of 1 to 2.

7. The method according to claim 1, characterized in that... The number-average molecular weight of humic acid A obtained in step (4.1) is greater than 2000 Da.

8. The method according to claim 1, characterized in that... The humic acid B obtained in step (4.1) is characterized in that the number average molecular weight of humic acid B is less than 2000 Da.

9. The method according to claim 1, characterized in that... The rapid cooling in step (4.2) involves cooling the resulting product at a rate of 20~50℃ / min.

10. The method according to claim 1, characterized in that... The fulvic acid content in the fulvic acid concentrate obtained in step (4.2) shall not be less than 40%.

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