A method for preparing fiber from pineapple leaves through bio-degumming without scraping pretreatment

Abstract

This invention discloses a method for preparing pineapple leaf fiber through biological degumming without pre-treatment of scraping, belonging to the field of pineapple leaf fiber preparation technology. It utilizes *Bacillus subtilis* XW-18, a pectinase-producing bacterium isolated and screened from pineapple plantations, to directly impregnate and ferment pineapple leaves to obtain pineapple leaf fiber. The screened strain used in this invention effectively removes hemicellulose, lignin, and gum substances tightly bound to pineapple leaves, resulting in soft, extensible fibers with high yield, low residual gum content, clean, neat, bundled fibers, and strong thermal stability. Compared to the current process of "manual / mechanical scraping of pineapple leaves → pineapple leaf raw fiber → physicochemical degumming → pineapple leaf finished fiber," this invention can complete the preparation of pineapple leaf to finished fiber in one step. The operation process is simple, green, and environmentally friendly. It not only avoids the damage to the raw fiber caused by manual or mechanical scraping but also overcomes the problems of high energy consumption, environmental pollution, and damage to the finished fiber caused by physicochemical degumming methods.

Description

Technical Field

[0001] This invention relates to the field of pineapple leaf fiber preparation technology, specifically a method for preparing fiber from pineapple leaves through biological degumming without scraping pretreatment. Background Technology

[0002] After pineapples are harvested, a large number of pineapple leaves are generated and discarded directly in the fields or burned as waste, which not only threatens the environment but also wastes resources.

[0003] Pineapple leaf fiber is a natural plant fiber with high toughness, tensile strength, elasticity, white color, abrasion resistance, and antibacterial and biodegradable properties. It has significant potential application value and market development prospects in the textile and paper industries, and in improving the performance of certain composite materials. Fiber extraction and degumming are the two most important processes in pineapple leaf fiber preparation, significantly impacting fiber quality and yield. Currently, most methods involve manually or mechanically scraping away the pectin from pineapple leaves to extract the pineapple leaf raw fiber; this process is also known as scraping. Because the extracted pineapple leaf raw fiber still retains a high content of hemicellulose, lignin, and pectin, degumming is necessary. Currently, pineapple leaf fiber degumming commonly employs chemical methods based on acid and alkali reactions, which can damage the fiber and pollute the environment.

[0004] Microbial degumming refers to the direct inoculation of degumming bacteria into pineapple leaf fibers, utilizing enzymes such as pectinase, xylanase, and mannanase produced during the growth and reproduction of these bacteria for degumming. Because microbial degumming does not require steaming or the addition of acids or alkalis, it is a mild, environmentally friendly, and ideal method for fiber degumming. Currently, microbial degumming is mostly applied to pineapple leaf fibers that have undergone scraping treatment; there is no direct application to pineapple leaf fibers in a single step for fiber production. Summary of the Invention

[0005] The purpose of this invention is to at least solve one of the technical problems existing in the prior art, and to provide a method for preparing fiber from pineapple leaves through biological degumming without scraping the hemp pretreatment.

[0006] The technical solution of the present invention is as follows:

[0007] A method for preparing pineapple leaf fiber without scraping pretreatment involves first isolating a degumming strain XW-18 that can produce pectinase from pineapple plantations, and then directly inoculating pineapple leaves with this degumming strain for soaking, fermentation, and degumming to obtain pineapple leaf fiber.

[0008] As a preferred embodiment of the present invention, the following steps are included:

[0009] S1: Degumming strain XW-18 was isolated and screened from the soil of pineapple plantation.

[0010] S2: Cultivate the degummed strain XW-18 to obtain seed culture;

[0011] S3: Add sterile water to the seed liquid to prepare the fermentation solution; add pineapple leaves to the fermentation solution for soaking, fermentation and degumming.

[0012] As a preferred embodiment of the present invention, in step S3, the volume ratio of the bacterial solution to the sterile water is 1:9-14.

[0013] As a preferred embodiment of the present invention, in step S3, the mass-to-volume ratio of pineapple leaves to fermentation liquid is 1:5-10.

[0014] As a preferred embodiment of the present invention, in step S2, the degummed strain XW-18 is cultured in LB liquid medium.

[0015] As a preferred embodiment of the present invention, the LB liquid culture medium comprises: 10 g / L tryptone, 5 g / L yeast extract, 10 g / L NaCl, pH 7.4, and autoclaved at 121°C for 20 min.

[0016] The beneficial effects of this invention are as follows: This invention uses screened strains to prepare pineapple leaf fibers, which can effectively remove hemicellulose, lignin, and gum substances that are tightly bound to pineapple leaves. The prepared fibers are soft and extensible, with high fiber yield, low residual gum content, clean and neat fibers in bundles, and strong thermal stability. It also proves that the process of this invention can complete the preparation of finished pineapple leaf fibers in one step, without the need for hemp scraping pretreatment. It has the advantages of simple operation and environmental friendliness. It not only avoids the damage to pineapple leaf fibrils caused by manual or mechanical hemp scraping, but also overcomes the problems of high energy consumption, environmental pollution, and damage to finished fibers caused by physicochemical degumming methods. Attached Figure Description

[0017] Figure 1 This is a diagram illustrating the process of preparing pineapple leaf fiber using microorganisms obtained through screening and isolation.

[0018] Figure 2 Images of pineapple leaves and pineapple leaf fibers prepared by different methods, along with their scanning electron microscope (SEM) images: (A, F, K) are pineapple leaves and their SEM images; (B, G, L) are pineapple leaf fibers prepared by manual scraping and their SEM images; (C, H, M) are pineapple leaf fibers prepared by commercially available pectinase and their SEM images; (D, I, N) are pineapple leaf fibers prepared by crude enzyme solution of strain XW-18 and their SEM images; (E, J, O) are pineapple leaf fibers prepared by immersion fermentation of strain XW-18 and their SEM images.

[0019] Figure 3 These are infrared spectra of pineapple leaves and pineapple leaf fibers prepared by different methods; where (A) to (E) represent, respectively, the blank group of pineapple leaves, pineapple leaf fibers prepared by manual scraping, pineapple leaf fibers prepared by commercially available pectinase, pineapple leaf fibers prepared by crude enzyme solution of strain XW-18, and pineapple leaf fibers prepared by immersion fermentation of strain XW-18.

[0020] Figure 4 These are thermogravimetric (TG) curves of pineapple leaves and pineapple leaf fibers prepared by different methods in nitrogen; where (A) to (E) represent the blank group of pineapple leaves, pineapple leaf fibers prepared by manual scraping method, pineapple leaf fibers prepared by commercially available pectinase method, pineapple leaf fibers prepared by crude enzyme method of strain XW-18, and pineapple leaf fibers prepared by soaking and fermentation method of strain XW-18, respectively.

[0021] Figure 5 These are the thermogravimetric (DTG) curves of pineapple leaves and pineapple leaf fibers prepared by different methods in nitrogen; where (A) to (E) represent the blank group of pineapple leaves, pineapple leaf fibers prepared by manual scraping method, pineapple leaf fibers prepared by commercially available pectinase method, pineapple leaf fibers prepared by crude enzyme method of strain XW-18, and pineapple leaf fibers prepared by soaking and fermentation method of strain XW-18, respectively. Detailed Implementation

[0022] The technical solution of the present invention will be further explained below through the applicant's specific experiments.

[0023] 1. Materials and Methods

[0024] 1.1 Strains, culture media and raw material sources

[0025] 1.1.1 Strains

[0026] The strain that produces pectinase is Bacillus subtilis XW-18, which was isolated from the soil of a pineapple field in Xuwen, Zhanjiang, China. The selected strain was resuspended in 30% glycerol and stored in an ultra-low temperature freezer at -80℃.

[0027] 1.1.2 Culture medium

[0028] LB agar medium: tryptone 10 g / L, yeast extract 5 g / L, NaCl 10 g / L, agar 20 g / L, pH 7.4, autoclaved at 121°C for 20 min.

[0029] LB liquid medium: tryptone 10 g / L, yeast extract 5 g / L, NaCl 10 g / L, pH 7.4, autoclaved at 121°C for 20 min.

[0030] Enzyme-producing medium: sucrose 40 g / L, peptone 20 g / L, sodium nitrate 2.5 g / L, MgSO4·7H2O 2 g / L, K2HPO4 1 g / L, FeSO4·7H2O 0.01 g / L, pH 7.0-7.2, sterilized at 121℃ for 30 min.

[0031] 1.1.3 Raw material sources

[0032] The pineapple leaves used in this study were collected from pineapple plantations in Xuwen, Zhanjiang, China.

[0033] 1.2 Preparation method of pineapple leaf fiber

[0034] (1) Manual scraping of hemp

[0035] The pineapple leaf fibers are extracted manually. Freshly picked pineapple leaves are scraped to remove surface substances with a scraper, and the fibers are extracted directly from the leaves. The leaves are then washed multiple times with clean water until the fibers are white, and then naturally dried.

[0036] (2) Commercially available pectinase method

[0037] Pineapple leaf fiber was prepared by degumming using commercially available pectinase (Shanghai Yuanye Biotechnology Co., Ltd., product specifications: biological reagent purity, enzyme activity 50000 U / g). Pineapple leaves were directly added to a 3 g / L pectinase solution at a mass-to-volume ratio of 1:5. After adjusting the pH to 5.0, the solution was left to stand at 50℃ for 7 days. After degumming, the temperature was raised to 100℃ and maintained for 5 minutes to inactivate the pectinase. The pineapple leaves were then removed, washed with water to remove impurities, and the remaining fiber was washed again and air-dried.

[0038] (3) XW-18 strain crude enzyme method

[0039] The crude enzyme solution method utilizes the crude enzyme solution produced by fermentation of strain XW-18 to degumm and prepare pineapple leaf fiber. First, a single colony of XW-18 grown on LB agar plates is inoculated into a 250mL tertiary flask containing 50mL of LB liquid medium and cultured at 30℃ and 170rpm for 16 hours to obtain a seed culture. The seed culture is then inoculated at a rate of 2% (v / v) into enzyme-producing medium in 40mL / 250mL Erlenmeyer flasks and cultured at 30℃ and 170rpm for 24 hours. The culture is then centrifuged at 5000rpm for 20 minutes, and the supernatant is collected; this is the crude enzyme solution produced by fermentation of strain XW-18. The crude enzyme solution of strain XW-18 is diluted 1:1 with sterile water, and pineapple leaves are added directly to the diluted enzyme solution at a mass-to-volume ratio of 1:5. The mixture is then incubated at 50℃ for 7 days. Remove the pineapple leaves, wash away impurities with clean water, and wash the remaining fibers again before letting them air dry.

[0040] (4) XW-18 strain immersion fermentation method

[0041] The XW-18 impregnation-fermentation method refers to the direct preparation of pineapple leaf fiber by inoculating with strain XW-18 and degumming it. First, a single colony of XW-18 grown on an LB agar plate is inoculated into a 250mL Erlenmeyer flask containing 50mL of LB liquid medium and cultured at 30℃ and 170rpm for 16 hours to obtain a seed culture. The seed culture is then diluted with sterile water at a volume ratio of 1:9 to prepare an impregnation-fermentation degumming solution. Pineapple leaves are then directly added to this solution, with a leaf mass to solution volume ratio of 1:5 (w / v). The solution is incubated at 30℃ with shaking (170rpm) for 7 days. The pineapple leaves are then removed, washed with clean water to remove impurities, and the remaining fiber is washed again and air-dried.

[0042] 1.3 Analysis of fiber yield and residual glue rate

[0043] 1.3.1 Fiber Yield

[0044] The yield of pineapple leaf fiber was calculated based on the mass before and after preparation, as shown in formula (1).

[0045] Equation (1) M W = (M0 / M1) × 100%

[0046] Where: M W —Fiber yield, %; M0—Mass of the prepared fiber, g; M1—Mass of the pineapple leaves before preparation, g.

[0047] 1.3.2 Fiber Yield

[0048] The yield of pineapple leaf fiber was calculated based on the mass before and after preparation, as shown in formula (1).

[0049] Equation (1) M W = (M0 / M1) × 100%

[0050] Where: M W —Fiber yield, %; M0—Mass of the prepared fiber, g; M1—Mass of the pineapple leaves before preparation, g.

[0051] 1.4 Structural characterization of pineapple leaves and pineapple leaf fibers

[0052] The pineapple leaves and the prepared pineapple leaf fibers were characterized by scanning electron microscopy, infrared spectroscopy, and thermogravimetric analysis.

[0053] 1.4.1 Scanning Electron Microscope

[0054] The surface morphology of pineapple leaves and pineapple leaf fibers was observed using a scanning electron microscope.

[0055] 1.4.2 Fourier Infrared Spectroscopy

[0056] Using Fourier transform infrared spectroscopy, a certain amount of pineapple leaves and pineapple leaf fibers were mixed with KBr, ground, and then pressed into thin sheets. The structure of the pineapple leaves and pineapple leaf fibers was characterized using a scanning range of 4000 cm⁻¹. -1 ~500cm -1 The resolution is 4cm. -1 Each wavelength is scanned 32 times.

[0057] 1.4.3 Thermogravimetric Analysis

[0058] Thermogravimetric analysis (TGA) of pineapple leaves and pineapple leaf fibers was performed using a simultaneous thermal analyzer. The heating range was 30–700℃, the heating rate was 10℃ / min, and the airflow rate was controlled at 20 mL / min. The analysis was conducted in both nitrogen and ordinary air environments to determine the thermal stability of pineapple leaves and pineapple leaf fibers.

[0059] 2. Results and Analysis

[0060] 2.1 Fiber yield and residual glue rate

[0061] Pineapple leaf fiber was prepared using different methods, and the fiber yield and residual gum rate were calculated for each method. Table 1 shows that the manual scraping method yielded the lowest fiber yield (0.71%) and the highest residual gum rate (24.00%), indicating that scraping not only causes fiber loss but also requires degumming. The XW-18 impregnation and fermentation method yielded the highest fiber yield and the lowest residual gum rate, at 2.29% and 3.50%, respectively, indicating that this method had the highest degumming efficiency and the least fiber loss among all preparation methods.

[0062] Table 1. Fiber yield and residual gum content of pineapple leaf fiber under different preparation methods.

[0063]

[0064] 2.2 Observation of the appearance and morphology of pineapple leaves and pineapple leaf fibers

[0065] The morphology of pineapple leaves and pineapple leaf fibers was observed using a scanning electron microscope, such as... Figure 2As shown in (A-E). The pineapple leaves in the control group turned pale yellow after dehydration and drying, while the pineapple leaf fibers prepared by different methods all exhibited the morphology expected of natural fibers. Under scanning electron microscopes at 600× and 1000×, the surface of the pineapple leaves in the control group was rough and uneven, with many small grooves. The surface of the pineapple leaf fibers was covered with hemicellulose, lignin, and gum substances. In contrast, the pineapple leaf fibers prepared by manual scraping, commercially available pectinase, XW-18 enzyme crude solution, and XW-18 impregnation and fermentation methods showed the disappearance of surface grooves, a smooth and clean fiber surface, and the removal of most of the hemicellulose, lignin, and gum substances, resulting in fiber bundles. However, the pineapple leaf fibers extracted by the manual scraping method still had a significant amount of gum surrounding their surface. Figure 2 -G,L) indicates that the pineapple leaf fiber extracted by manual scraping still needs further degumming. Compared with the commercially available pectinase method and the crude enzyme solution method of strain XW-18, the pineapple leaf fiber prepared by the XW-18 impregnation and fermentation method has a smoother surface, a significantly finer fiber diameter, and larger gaps and pores between fibers. Figure 2 -J,O), indicating that the XW-18 strain's immersion fermentation method helps improve the appearance and structure of fibers.

[0066] 2.3 Infrared Spectroscopic Characterization Analysis of Pineapple Leaves and Pineapple Leaf Fibers

[0067] The functional group changes in pineapple leaves and pineapple leaf fibers were characterized using Fourier transform infrared spectroscopy. The infrared spectra are shown below. Figure 3 As shown in Table 2, the characteristic absorption peaks and their corresponding bonds / functional groups are also shown. Analysis of the main characteristic absorption peaks of pineapple leaves and pineapple leaf fibers reveals that the peak at 3416 cm⁻¹ is... -1 It is the -OH functional group band of cellulose, the main component of pineapple leaf fiber; 2920cm -1 This is the CH functional group band on the saturated carbon of the fiber, which is the stretching vibration peak of the -CH (-CH3, -CH2) unit of cellulose and lignin phenylpropane; 1734 cm⁻¹ -1 The peak represents the stretching vibration of hemicellulose C=O. However, compared to pineapple leaves, the absorption peak at this point is relatively weaker in pineapple leaf fibers prepared by commercially available pectinase method, XW-18 enzyme crude liquid method, and XW-18 impregnation fermentation method, indicating a relative reduction in lignin and hemicellulose content in the fibers; 1052 cm⁻¹ -1 The absorption peaks are the C-OH functional group bands and β-1,4-glycosidic bond stretching vibrations in cellulose and lignin. Pineapple leaves and pineapple leaf fibers prepared by various methods all showed strong absorption peaks, indicating that the degumming treatment did not change the cellulose structure.

[0068] Table 2. Infrared characteristic absorption peaks of pineapple leaves and pineapple leaf fibers and their corresponding bonds / functional groups.

[0069]

[0070] 2.4 Thermogravimetric analysis results of pineapple leaves and pineapple leaf fibers

[0071] The thermogravimetric (TG) and micro-quotient (DTG) curves of pineapple leaves and pineapple leaf fibers are shown below. Figure 4 A~E and Figure 5 As shown in Figures A through E, the thermal decomposition process of pineapple leaves in the control group can be divided into three stages (…). Figure 4 A and Figure 5 A): The first stage is between 40 and 230°C, with a weight loss rate of 9.76%, corresponding to the temperature stage of slow decomposition in the DTG curve. The weight loss here is mainly water, which is the external water absorbed by hemicellulose and lignin in the pineapple leaves, as well as the internal water released by the breakage of unstable side chains in the structure. The second stage is between 240 and 360°C, with a weight loss rate of 75.19%, corresponding to the stage of sharp decline in the DTG curve. This indicates that the pineapple leaves lose the most mass at this stage and are converted into gaseous compounds at around 360°C. This is the process of some biomass macromolecules breaking down into small molecule monomers. The last stage is between 450 and 700°C, which is the degradation process of the small molecule monomers formed after the degradation of the pineapple leaves.

[0072] The thermal decomposition diagrams of pineapple leaf fibers prepared by different methods can also be roughly divided into three stages ( Figure 4 B~E and Figure 5 B~E): The first stage is between 40~230℃, with a weight loss rate of 6.5~4.46%. However, compared with pineapple leaves, the prepared fiber has a lower weight loss rate in this stage due to its lower water content. The second stage is between 260~380℃, with a weight loss rate of 75.19~80.7%. However, the DTG curve shows that the temperature at which pineapple leaf fiber decomposes rapidly is higher than that of pineapple leaves, indicating that the prepared pineapple leaf fiber has a relatively lower impurity content. The third stage is between 360~700℃, with a smaller weight loss rate.

[0073] Compared to manual scraping, commercially available pectinase, and crude enzyme solution of strain XW-18, the pineapple leaf fiber prepared by the XW-18 impregnation and fermentation method had the highest weight loss rate, reaching 96.18%. This indicates that the XW-18 impregnation and fermentation method can more efficiently remove the gum attached to the fiber during fiber preparation, increase the cellulose content in the pineapple leaf fiber, and reduce the lignin and hemicellulose content, resulting in fewer impurities and consequently less residue remaining after the thermal decomposition of cellulose.

[0074] 3. Conclusion

[0075] The world produces a vast quantity of pineapple leaves annually, providing a significant resource advantage for the production of natural plant fibers. However, pineapple leaves are composed of cellulose, hemicellulose, lignin, and pectin, which are tightly bound together. Simply peeling the leaves manually or mechanically is insufficient to separate the pectinous substances, necessitating further degumming. Chemical degumming methods using acids and alkalis not only damage the properties of the extracted fibers but also generate waste alkalis and acids that harm the environment. Degumming using microorganisms from the natural environment is uncontrollable, and purchasing commercially available pectinase for degumming is too expensive, with subsequent processing steps being quite complex.

[0076] In contrast, screening specific strains of bacteria capable of degrading gum substances from nature, which is rich in microorganisms and enzymes, offers advantages such as high efficiency, environmental friendliness, and high specificity. The soil in pineapple plantations is rich in pineapple leaf waste, thus fostering many microorganisms closely related to the degradation and utilization of pineapple leaf gum substances. Specific degumming microorganisms can be screened from these microorganisms for the production of pineapple leaf fibers. Furthermore, because the microbial degumming process is conducted under mild conditions, it can significantly improve the physical properties of pineapple leaf fibers, including fineness, moisture regain, luster, brightness, and whiteness.

[0077] This invention applies the screened Bacillus subtilis XW-18 to the preparation of pineapple leaf fiber, discovering that it can be directly used for degumming pineapple leaf fiber, resulting in fibers that exhibit the characteristics of natural plant fibers. This preparation process eliminates the need for scraping pretreatment, simplifying the pineapple leaf fiber preparation process. Furthermore, fiber yield, residual gum rate, and fiber structure characterization results indicate that Bacillus subtilis XW-18 can effectively remove the gum substances tightly bound to the fibers in pineapple leaves. The prepared pineapple leaf fiber has advantages such as high fiber yield, low residual gum rate, and strong thermal stability. This invention provides a method for preparing pineapple leaf fiber without scraping pretreatment, which not only has the advantages of simple operation and environmental friendliness, but also effectively increases fiber content and avoids fiber damage, helping to improve the quality of pineapple leaf fiber, reduce fiber production costs, and promote the high-value utilization of pineapple leaf waste.

[0078] The embodiments described above are merely preferred embodiments of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art can make various other corresponding changes and modifications based on the technical solutions and concepts described above, and all such changes and modifications should fall within the protection scope of the claims of the present invention.

Claims

1. A method for preparing fiber from pineapple leaves through biological degumming without scraping pretreatment, characterized in that, First, a strain of Bacillus subtilis XW-18 that can produce pectinase was isolated from the pineapple plantation. Then, this degumming strain was used to directly inoculate pineapple leaves for soaking, fermentation and degumming to obtain pineapple leaf fiber. The steps are as follows: S1: Degumming strain XW-18 was isolated and screened from the soil of pineapple plantation. S2: Cultivate the degummed strain XW-18 to obtain seed culture; S3: Add sterile water to the seed liquid to prepare the fermentation solution; add pineapple leaves to the fermentation solution for soaking, fermentation and degumming; the pineapple leaves are harvested from pineapple plantations in Xuwen, Zhanjiang, China; In step S3, the mass-to-volume ratio of pineapple leaves to fermentation liquid is 1:5-10.

2. The method for preparing fiber from pineapple leaves through biological degumming without scraping pretreatment according to claim 1, characterized in that, In step S3, the volume ratio of the bacterial solution to sterile water is 1:9-14.

3. The method for preparing fiber from pineapple leaves through biological degumming without scraping pretreatment according to claim 1, characterized in that, In step S2, the degummed strain XW-18 is cultured in LB liquid medium.

4. The method for preparing fiber from pineapple leaves through biological degumming without scraping pretreatment according to claim 3, characterized in that, The LB liquid culture medium comprises: 10 g / L tryptone, 5 g / L yeast extract, 10 g / L NaCl, pH 7.4, and autoclaved at 121°C for 20 min.

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