A METHOD FOR PRODUCTION OF FUNGAL INULINASE

Abstract

The present invention discloses a method for production of fungal inulinase. More particularly, a novel method for the efficient and cost-effective production of fungal inulinase, aiming to address the challenges associated with bioethanol production from renewable sources, particularly inulin-rich substrates. The said method includes various steps such as sampling (110), isolation (120), screening (130), identification (140), and optimization (150). This invention offers a comprehensive approach from strain selection to application, significantly contributing to the advancement of sustainable bioeconomy.The invention begins with the sampling, isolation, screening, and identification of potent fungal strains capable of producing high levels of inulinase, focusing on strains indigenous to Punjab, India. Overall, this invention represents a significant advancement in the field of bioethanol production, offering a scalable, cost-effective, and environmentally sustainable solution for harnessing the potential of inulin-rich biomass towards the development of a bio-based economy.

Specification

Description:The following specification particularly describes the invention and the manner in which it is to be performed.
CROSS-REFERENCE TO RELATED APPLICATIONS AND PRIORITY
[001] The present application does not claim priority from any patent application.
PREAMBLE
[002] The following specification particularly describes the invention and the manner in which it is to be performed.
TECHNICAL FIELD
[003] The present subject matter described herein, in general, relates to a method for production of fungal inulinase, and more particularly, a novel method for the efficient and cost-effective production of fungal inulinase, aiming to address the challenges associated with bioethanol production from renewable sources, particularly inulin-rich substrates.
BACKGROUND
[004] Inulin has tremendous potential as a renewable energy source and there are a number of plant sources that are underutilised. Even though inulin is employed as a nutritional supplement in the food business, there is still a long way to go in developing cost-effective methods for large-scales synthesis of inulinase enzymes, which may be used to produce various forms of fructooligosaccharides (FOS). As mentioned in the previous section fructose can also be derived from inulin, which is rapidly becoming a commercially relevant constituent in both the pharmaceutical and food industries. It is particularly important in biofuels since bioethanol can be recovered in large quantities from it and inulinase can enhance the process by breaking down inulin polymer into fructose which can further be fermented to form ethanol. The most significant research gap in bioethanol production is scarcity of data using fungal inulinase, detailed investigation of the parameter which can aid in increasing output and its scale up studies. As a result, it is critical to identify these research gaps in order to progress towards a sustainable bioeconomy and to investigate new ideas for producing bioethanol from these sources.
[005] The prior art TR2023018347A2 discloses the method of obtaining microbial inulinase enzyme using Cryphonectria parasitica. The invention relates to the production of inulinase enzyme, which provides enzymatic fructose production, by subjecting inulin to a single fermentation, and thus to obtain inulinase enzyme at a low cost, by Cryphonectria parasitica, a microbial fungus species found in high numbers in Turkey. However, prior art fails to mention the method of production of fungal inulinase contributing to the advancement of sustainable bioeconomy.
[006] The prior art CN111235073A discloses Lactobacillus plantarum producing inulinase and application thereof. The strain has the capability of producing acid inulinase by fermentation and degrading fructan substances to prepare products such as fructose and the like. The object of the present invention is the use of Lactobacillus plantarum for the production of inulinase. However, prior art fails to mention the method of production of fungal inulinase contributing to the advancement of sustainable bioeconomy.
[007] The present invention addresses the above shortcomings of the prior art. However, the invention is entirely different from the prior art in terms of novelty and technological advancements.
OBJECT
[008] The main object of the present invention is sampling, isolation, screening, and identification of inulinase-producing fungal strain from Punjab, India and optimization of fungal inulinase production and application in biorefinery.
SUMMARY
[009] The disclosure of the present invention discloses a method for production of fungal inulinase. More particularly, a novel method for the efficient and cost-effective production of fungal inulinase, aiming to address the challenges associated with bioethanol production from renewable sources, particularly inulin-rich substrates. A method for producing inulinase comprising the steps of sampling, isolating, screening, and identifying potent fungal strains from inulin-rich substrates, particularly those indigenous to Punjab, India. A method for optimizing the media components for fungal inulinase. The application of crude fungal inulinase in the biorefinery industry for the production of bioethanol from inulin-rich biomass. Fungal inulinase presents a promising solution for enhancing the efficiency of biorefineries by enabling the conversion of inulin-rich biomass into valuable products. While challenges remain, ongoing research and development are likely to optimize its use, making it a key player in sustainable biotechnological applications.
[0010] Embodiments may disclose the method for production of fungal inulinase. Fungal inulinase is an enzyme produced by certain fungi that catalyzes the hydrolysis of inulin, a fructan polysaccharide found in various plants. This enzyme plays a significant role in biotechnological applications, particularly in the production of fermentable sugars, biofuels, and other valuable biochemicals. Inulinase cleaves the glycosidic bonds in inulin, resulting in the release of fructose and glucose. Fungal inulinase is a valuable enzyme with diverse applications, particularly in biofuel production and food processing. Fungal inulinase is a key enzyme in the sustainable production of bioethanol, facilitating the conversion of non-food biomass into renewable energy. By addressing challenges and leveraging its advantages, the integration of inulinase in bioethanol production can contribute significantly to a more sustainable energy future.
[0011] In these implementations a method for production of fungal inulinase is described. The above-mentioned method comprises various steps such as a) sampling (110) wherein, soil samples collected from various locations in Punjab, ensuring diversity in the substrate type and geographic area, b) isolation (120) wherein, isolation of inulinolytic fungi from soil samples surrounding inulin-rich plants followed by serial dilution and spread plating, c) screening (130) wherein, axenic cultures are screened for inulinase activity using Lugol's iodine solution on Czapek-Dox agar containing inulin, d) identification (140) wherein, identification of most active isolates by morphological and molecular characterization using 18S rRNA gene sequencing, and e) optimization (150) wherein, inulinase production is optimized by selecting the fungal strain in submerged fermentation also key parameters like inoculum size, pH, and aeration will be optimized using minimal media containing inulin as the carbon source.
BRIEF DESCRIPTION
[0012] The foregoing detailed description of embodiments is better understood when read in conjunction with the appended drawings. For the purpose of illustrating of the present subject matter, an example of construction of the present subject matter is provided as figures; however, the invention is not limited to the specific method disclosed in the document and the figures.
[0013] Figure. 1 is a block diagram illustrating the process, according to some embodiments of the present disclosure.
[0014] Figure. 2 is an image of the strain identified as A. terreus, according to some embodiments of the present disclosure.
DETAILED DESCRIPTION
[0015] Some of the embodiments of this disclosure, illustrating all its features will now be discussed in detail. The words "comprising," "having," "containing," and "including," and other forms thereof, are intended to be equivalent in meaning and be open-ended in that an item or items following any one of these words is not meant to be an exhaustive listing of such item or items or meant to be limited to only the listed item or items. It must also be noted that as used herein and in the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise.
[0016] Before the present method for production of fungal inulinase is described, it is to be understood that this method is not limited to the particular system(s), and methodologies described, as there can be multiple possible embodiments that are not expressly illustrated in the present disclosure. It is also to be understood that the terminology used in the description is to describe the particular implementations or versions or embodiments only, and is not intended to limit the scope of the present application. This summary is provided to introduce aspects involved in the method for production of fungal inulinase. This summary is not intended to identify essential features of the claimed subject matter nor is it intended for use in determining or limiting the scope of the claimed subject matter.
[0017] The present invention discloses a novel method for the efficient and cost-effective production of fungal inulinase, aiming to address the challenges associated with bioethanol production from renewable sources, particularly inulin-rich substrates. This invention offers a comprehensive approach from strain selection to application, significantly contributing to the advancement of sustainable bioeconomy. The invention begins with the sampling, isolation, screening, and identification of potent fungal strains capable of producing high levels of inulinase, focusing on strains indigenous to Punjab, India. Subsequently, the identified strain media components will be optimized. Once optimized, the application of crude fungal inulinase will be explored in the field of biorefinery. Overall, this invention represents a significant advancement in the field of bioethanol production, offering a scalable, cost-effective, and environmentally sustainable solution for harnessing the potential of inulin-rich biomass towards the development of a bio-based economy.
[0018] In one embodiment, the method for producing fungal inulinase comprising the steps of sampling, isolating, screening, and identifying potent fungal strains from inulin-rich substrates, particularly those indigenous to Punjab, India. A method for optimizing the media components for fungal inulinase. The application of crude fungal inulinase in the biorefinery industry for the production of bioethanol from inulin-rich biomass. Fungal inulinase presents a promising solution for enhancing the efficiency of biorefineries by enabling the conversion of inulin-rich biomass into valuable products. While challenges remain, ongoing research and development are likely to optimize its use, making it a key player in sustainable biotechnological applications
[0019] In another embodiment, the method for production of fungal inulinase is disclosed. Fungal inulinase is an enzyme produced by certain fungi that catalyzes the hydrolysis of inulin, a fructan polysaccharide found in various plants. This enzyme plays a significant role in biotechnological applications, particularly in the production of fermentable sugars, biofuels, and other valuable biochemicals. Inulinase cleaves the glycosidic bonds in inulin, resulting in the release of fructose and glucose. Fungal inulinase is a valuable enzyme with diverse applications, particularly in biofuel production and food processing. Fungal inulinase is a key enzyme in the sustainable production of bioethanol, facilitating the conversion of non-food biomass into renewable energy. By addressing challenges and leveraging its advantages, the integration of inulinase in bioethanol production can contribute significantly to a more sustainable energy future
[0020] In yet another embodiment, the crude inulinase will be assessed for its ability to convert pre-treated inulin-rich agro-residues followed by high-Performance Liquid Chromatography (HPLC) will be used to monitor fructose production. Finally, Gas Chromatography-Mass Spectrometry (GC-MS) will quantify the bioethanol yield from the inulinase-mediated conversion of fructose. The invention described offers several advantages in the context of bioethanol production from inulin-rich substrates. These advantages can be summarized as follows: the method focuses on isolating and identifying fungal strains that produce high levels of inulinase, ensuring efficient enzyme production, the optimization of media components, the use of fungal strains indigenous to Punjab, India, potentially lowers the dependency on imported strains and leverages local biodiversity, the invention supports the production of bioethanol from renewable, inulin-rich biomass, contributing to a sustainable bioeconomy, utilizing inulin-rich biomass and producing bioethanol reduces reliance on fossil fuels, thereby decreasing greenhouse gas emissions, and by providing a method that supports the production of bioethanol and other bioproducts, the invention contributes to the development and growth of a bio-based economy. Overall, this research aims to develop a cost-effective and scalable fungal inulinase production process for bioethanol generation from underutilized inulin sources, contributing to a sustainable bioeconomy.
[0021] In one embodiment, referring to figure 1, the method for production of fungal inulinase comprises various steps such as: a) Sampling (110) where soil samples collected from various locations in Punjab, ensuring diversity in the substrate type and geographic area, b) Isolation (120) where isolation of inulinolytic fungi from soil samples surrounding inulin-rich plants followed by serial dilution and spread plating, c) Screening (130) where axenic cultures are screened for inulinase activity using Lugol's iodine solution on Czapek-Dox agar containing inulin, d) Identification (140) where identification of most active isolates by morphological and molecular characterization using 18S rRNA gene sequencing, and e) Optimization (150) where inulinase production is optimized by selecting the fungal strain in submerged fermentation also key parameters like inoculum size, pH, and aeration will be optimized using minimal media containing inulin as the carbon source.
[0022] In one embodiment of the present invention, experimental data shows that in a study involving six sampling sites, collected 24 samples and isolated 46 cultures. Among these, one strain exhibited the highest enzyme activity, leading to its identification. Additionally, the strain labeled N5S3-b showed significant genetic similarity to Aspergillus terreus based on nucleotide homology and phylogenetic analysis. Figure 2 shows the image of the strain identified as A. terreus.
, Claims:1. A method for production of fungal inulinase comprises:
a) sampling (110);
b) isolation (120);
c) screening (130);
d) identification (140); and
e) optimization (150).
2. The method as claimed in claim 1, wherein soil samples collected from various locations in Punjab, ensuring diversity in the substrate type and geographic area.
3. The method as claimed in claim 1, wherein isolation of inulinolytic fungi from soil samples surrounding inulin-rich plants followed by serial dilution and spread plating.
4. The method as claimed in claim 1, wherein axenic cultures are screened for inulinase activity using Lugol's iodine solution on Czapek-Dox agar containing inulin.
5. The method as claimed in claim 1, wherein identification of most active isolates by morphological and molecular characterization using 18S rRNA gene sequencing.
6. The method as claimed in claim 1, wherein inulinase production is optimized by selecting the fungal strain in submerged fermentation also key parameters like inoculum size, pH, and aeration will be optimized using minimal media containing inulin as the carbon source.

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