CN116349700B - Application of noni fruit extract in inhibiting aspergillus flavus - Google Patents

Application of noni fruit extract in inhibiting aspergillus flavus Download PDF

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CN116349700B
CN116349700B CN202310180175.4A CN202310180175A CN116349700B CN 116349700 B CN116349700 B CN 116349700B CN 202310180175 A CN202310180175 A CN 202310180175A CN 116349700 B CN116349700 B CN 116349700B
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noni
powder
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aspergillus flavus
freeze
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CN116349700A (en
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陈旭玉
杨美华
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Institute of Medicinal Plant Development of CAMS and PUMC
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    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N65/00Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
    • A01N65/08Magnoliopsida [dicotyledons]
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N65/00Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01PBIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
    • A01P3/00Fungicides
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02ATECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

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  • Preparation Of Fruits And Vegetables (AREA)

Abstract

The invention relates to application of noni fruit extract in inhibiting aspergillus flavus, which is prepared by the following steps: slicing noni, freeze-drying and crushing to obtain noni powder; mixing noni powder with water, adjusting pH to 5-6, and performing ultrasonic-assisted extraction at 60-80deg.C. The noni fruit extract can obviously inhibit the growth of aspergillus flavus, can effectively control the aspergillus flavus to produce aflatoxin, is a plant extract with good biocontrol effect and no pollution, and has wide development and application prospects.

Description

Application of noni fruit extract in inhibiting aspergillus flavus
Technical Field
The invention relates to the technical field of plant extracts, in particular to application of noni fruit extracts in the aspect of inhibiting aspergillus flavus.
Background
Aspergillus flavus (Aspergillus flavus) is Aspergillus fungus. Aspergillus flavus is a common saprophytic fungus which is widely distributed in nature and is mostly found on soil, moldy grains and other moldy organic matters. Aspergillus flavus can produce aflatoxin, has carcinogenicity, and can cause animal poisoning. After eating foods containing aflatoxin, animals can cause liver damage, causing toxic diseases characterized by liver degeneration, swelling, cirrhosis and bleeding.
Noni refers to Morinda citrifolia, the chemical name Morinda citrifolia Linn, the English name Noni, and also the names Morinda citrifolia, and Noni, which are plants of the genus Morinda of the family Rubiaceae. Noni is the fruit of Morinda citrifolia. Noni belongs to traditional medicinal plants, has good curative effects on cancers, hypertension, diabetes mellitus and the like, has research and considers that noni has antibacterial effects, for example Liu Aisha and the like examine the antibacterial activity of noni fruit fermentation juice, seed ethanol extract, seed water distilled extract and noni leaf extract on 4 pathogenic bacteria of pseudomonas aeruginosa, escherichia coli, staphylococcus aureus and streptococcus mutans, and the result shows that the noni fruit fermentation juice with the concentration of 75% -100% has an inhibitory effect on three pathogenic bacteria and has no inhibitory effect on streptococcus mutans, and the seed water distilled extract with the concentration of 10% -2% has an inhibitory effect on four pathogenic bacteria and the seed ethanol extract with the concentration of 10% -4% has no inhibitory effect on escherichia coli, staphylococcus aureus and streptococcus mutans. Min and other researches show that the noni fresh fruit extract and noni fermentation juice have certain inhibition effect on escherichia coli. The bacteriostatic action of noni plants has obtained a certain research result, but the current research is mainly focused on bacteria such as escherichia coli, reports on the inhibition of fungi by noni fruit extracts are rarely seen, and reports on the resistance of aspergillus flavus are not seen.
Disclosure of Invention
In view of the defects of the prior art, the invention provides application of noni fruit extract in inhibiting aspergillus flavus, and the invention provides that the application of noni fruit can be further widened, and the medicinal value of noni fruit is improved.
The technical scheme of the invention mainly comprises the following contents:
The invention relates to application of noni fruit extract in inhibiting aspergillus flavus, which is prepared by the following steps: slicing noni, freeze-drying and crushing to obtain noni powder; mixing noni powder with water, adjusting pH to 5-6, and performing ultrasonic-assisted extraction at 60-80deg.C. The method strictly controls the water extraction conditions to obtain the high-efficiency antibacterial part.
Preferably, the noni extract is prepared by the following method: slicing noni, freeze-drying and crushing to obtain noni powder; mixing noni powder with acetone, soaking at 30-50deg.C for 15-30 min, filtering, removing filtrate, and lyophilizing to obtain pretreated noni powder; mixing the pretreated noni fruit powder with water, adjusting pH to 5-6, and performing ultrasonic-assisted extraction at 60-80deg.C. Through pretreatment, the release of active ingredients can be promoted, the influence of other ingredients on the antibacterial effect is avoided, and meanwhile, the purity of the final product is improved, so that the antibacterial effect is improved.
Noni changes color from dark green to light green, then to yellowish green, then to pale yellow, and finally to white as maturity increases. The invention discovers that the Aspergillus flavus inhibition effect of the extract obtained by taking the noni fruits with green yellow and immature seeds as raw materials is obviously better than that of the noni fruits with complete ripeness.
Preferably, the noni fruits are crushed to have a particle size of 10-30 μm.
Preferably, the mass ratio of noni fruit powder to water is 1:10-30.
Preferably, the mass ratio of noni fruit powder to acetone is 1:5-10.
Preferably, the ultrasonic frequency is 20kHz-40kHz, the ultrasonic power is 50mW/mL-100mW/mL, and the ultrasonic time is 60min-90min.
More preferably, the noni extract is prepared by the following method: taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; mixing noni powder and acetone according to a mass ratio of 1:10, soaking for 15min at 50 ℃, filtering, removing filtrate, and freeze-drying to obtain pretreated noni powder; mixing the pretreated noni fruit powder with water according to a mass ratio of 1:20, adjusting pH to 5.2, performing ultrasonic-assisted extraction at 80 ℃ for 90min, centrifuging, collecting supernatant, and freeze-drying; the ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Compared with the prior art, the invention has the beneficial effects that:
the preparation method of the extract is simple and easy to operate, and is easy to popularize and use in industrialization.
The noni fruit extract prepared by the invention can obviously inhibit the growth of aspergillus flavus, can effectively control the aspergillus flavus to produce aflatoxin, is a plant extract with good biocontrol effect and no pollution, and has wide development and application prospects.
The noni extract provided by the invention can show obvious aspergillus flavus inhibition effect at a lower concentration, and the minimum inhibition concentration is as low as 1.25mg/mL.
Drawings
Fig. 1: the inhibitory effect of noni fruit extract on aspergillus flavus; upper graph: aspergillus flavus, the following diagram: the growth effect of Aspergillus flavus in the culture medium containing noni fruit extract is shown to be inhibited.
Detailed Description
In order to better understand the technical content of the present invention, the following provides specific examples to further illustrate the present invention.
Example 1: preparation of noni fruit extract
Taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; the noni fruit powder and water are mixed according to the mass ratio of 1:20, adjusting pH to 5.5, performing ultrasonic-assisted extraction at 80deg.C for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Example 2: preparation of noni fruit extract
Taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; the noni fruit powder and water are mixed according to the mass ratio of 1:20, adjusting pH to 5.3, performing ultrasonic-assisted extraction at 80deg.C for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze drying. The ultrasonic frequency is 40kHz, and the ultrasonic power is 50mW/mL.
Example 3: preparation of noni fruit extract
Taking white peel and completely cooked noni fruits, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; the noni fruit powder and water are mixed according to the mass ratio of 1:20, adjusting pH to 6.8, performing ultrasonic-assisted extraction at 80deg.C for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Example 4: preparation of noni fruit extract
Taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; mixing noni powder and acetone according to a mass ratio of 1:10, soaking for 15min at 50 ℃, filtering, removing filtrate, and freeze-drying to obtain pretreated noni powder; mixing the pretreated noni fruit powder with water according to a mass ratio of 1:20, adjusting pH to 5.0, performing ultrasonic-assisted extraction at 80 ℃ for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze-drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Example 5: preparation of noni fruit extract
Taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; mixing noni powder and acetone according to a mass ratio of 1:10, soaking for 15min at 50 ℃, filtering, removing filtrate, and freeze-drying to obtain pretreated noni powder; mixing the pretreated noni fruit powder with water according to a mass ratio of 1:20, adjusting pH to 6.0, performing ultrasonic-assisted extraction at 60 ℃ for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze-drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Example 6: preparation of noni fruit extract
Taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; mixing noni powder and acetone according to a mass ratio of 1:5, soaking for 30min at 50 ℃, filtering, removing filtrate, and freeze-drying to obtain pretreated noni powder; mixing the pretreated noni fruit powder with water according to a mass ratio of 1:20, adjusting pH to 6.0, performing ultrasonic-assisted extraction at 60 ℃ for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze-drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Experimental example 1: influence of noni extracts obtained by different extraction processes on Aspergillus flavus activity
1.1 Preparation of extracts
Extract a (hot water ultrasound assisted extraction): taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; the noni fruit powder and water are mixed according to the mass ratio of 1:20, adjusting pH to 5.2, performing ultrasonic-assisted extraction at 80deg.C for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Extract B (methanol extraction): taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; mixing noni fruit powder and methanol according to a mass ratio of 1:20, soaking for 6h at 50 ℃, centrifuging for 10min at 5000rpm, collecting supernatant, and freeze-drying.
1.2 Preparation of bacterial liquid:
Aspergillus flavus strain (CGMCC No.18545, information of which is described in patent document CN110982703B filed earlier by the inventor) is activated to prepare spore suspension with the concentration of 10 6 cfu/mL.
1.3 Inhibition zone and Minimum Inhibitory Concentration (MIC) determination
Taking filter paper sheets with the diameter of 6mm, and sterilizing for later use. The concentration of the extract (water as solvent) was 20mg/mL, 10mg/mL, 5mg/mL, 2.5mg/mL, 1.25mg/mL, and 0.625mg/mL, respectively, by double dilution. 25mL of PDA solid medium was added to the petri dish and condensed into a solid plate. 0.1mL of bacteria are dripped into a culture medium and uniformly coated, and a bacteria-containing flat plate is prepared. And (3) sticking the filter paper sheets subjected to soaking treatment of the extracts with different concentrations on the surface of a culture medium, and taking sterile water as a blank control group. The petri dish was sealed with a preservative film, cultured at 28℃for 48 hours, and the diameter of the inhibition zone (including the diameter of the filter paper sheet) was measured. Judging the antibacterial effect of the extract according to the diameter of the antibacterial ring. The test was repeated 3 times and the results averaged.
1.5 Experimental results
TABLE 1
20mg/mL 10mg/mL 5mg/mL 2.5mg/mL 1.25mg/mL 0.625mg/mL
Extract A 15.43±0.42 12.40±0.29 9.47±0.41 7.50±0.41 6.43±0.17 -
Extract B 7.21±0.22 6.25±0.20 - - - -
Blank control - - - - - -
"-" Means no inhibition.
The results in Table 1 show that the minimum inhibitory concentration of extract A was 1.25mg/mL and the minimum inhibitory concentration of extract B was 10mg/mL. The antibacterial effect of the extract A is obviously better than that of the extract B.
Experiment 2 study of noni extracts with different maturity to inhibit Aspergillus flavus Activity
Extract a: taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; the noni fruit powder and water are mixed according to the mass ratio of 1:20, adjusting pH to 5.2, performing ultrasonic-assisted extraction at 80deg.C for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Extract B: taking white peel and completely cooked noni fruits, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; the noni fruit powder and water are mixed according to the mass ratio of 1:20, adjusting pH to 5.2, performing ultrasonic-assisted extraction at 80deg.C for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
According to the method of experiment 1, the diameter of the inhibition zone and the minimum inhibition concentration were determined. The results are shown in Table 2.
TABLE 2
20mg/mL 10mg/mL 5mg/mL 2.5mg/mL 1.25mg/mL 0.625mg/mL
Extract A 15.43±0.42 12.40±0.29 9.47±0.41 7.50±0.41 6.43±0.17 -
Extract B 7.94±0.34 6.83±0.24 - - - -
Blank control - - - - - -
The results in Table 2 show that the minimum inhibitory concentration of extract A was 1.25mg/mL and the minimum inhibitory concentration of extract B was 10mg/mL.
Experiment 3 Effect of pretreatment on antibacterial Effect of noni extract
Extract a: taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; the noni fruit powder and water are mixed according to the mass ratio of 1:20, adjusting pH to 5.2, performing ultrasonic-assisted extraction at 80deg.C for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
Extract B: taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; mixing noni powder and acetone according to a mass ratio of 1:10, soaking for 15min at 50 ℃, filtering, removing filtrate, and freeze-drying to obtain pretreated noni powder; mixing the pretreated noni fruit powder with water according to a mass ratio of 1:20, adjusting pH to 5.2, performing ultrasonic-assisted extraction at 80 ℃ for 90min, centrifuging at 5000rpm for 10min, collecting supernatant, and freeze-drying. The ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
According to the method of experiment 1, the diameter of the inhibition zone and the minimum inhibition concentration were determined. The results are shown in Table 3.
TABLE 3 Table 3
The results in Table 3 show that the minimum inhibitory concentration of both extract A and extract B is 1.25mg/mL, and at concentrations above 2.5mg/mL, the inhibitory effect of extract B is significantly higher than that of extract A.
The foregoing description of the preferred embodiments of the invention is not intended to limit the invention to the precise form disclosed, and any such modifications, equivalents, and alternatives falling within the spirit and scope of the invention are intended to be included within the scope of the invention.

Claims (6)

1. Use of noni fruit extract for the preparation of a product for inhibiting aspergillus flavus, characterized in that the noni fruit extract is prepared by the following method:
Slicing noni, freeze-drying and crushing to obtain noni powder; mixing noni powder with acetone, soaking at 30-50deg.C for 15-30 min, filtering, removing filtrate, and lyophilizing to obtain pretreated noni powder; mixing the pretreated noni fruit powder with water, adjusting the pH to 5-6, and carrying out ultrasonic-assisted extraction at 60-80 ℃;
the noni fruits are yellowish green, and the seeds of the noni fruits are not mature.
2. Use of noni extract according to claim 1 for the preparation of a product for inhibiting aspergillus flavus, characterized in that the noni is crushed to a particle size of 10-30 μm.
3. Use of noni extract according to claim 1 for the preparation of a product for inhibiting aspergillus flavus, characterized in that the mass ratio of noni powder after pretreatment to water is 1:10-30.
4. Use of noni extract according to claim 1 for the preparation of a product for inhibiting aspergillus flavus, characterized in that the mass ratio of noni powder to acetone is 1:5-10.
5. Use of noni extract according to claim 1 for the preparation of a product for inhibiting aspergillus flavus, characterized in that the ultrasound frequency is 20kHz-40kHz, the ultrasound power is 50mW/mL-100mW/mL, the ultrasound time is 60min-90min.
6. Use of noni extract according to claim 1 for the preparation of a product for inhibiting aspergillus flavus, characterized in that the noni extract is prepared by the following method: taking noni fruits with yellow-green peel and immature seeds, slicing, freeze-drying and crushing to the particle size of 10-30 mu m to obtain noni fruit powder; mixing noni powder and acetone according to a mass ratio of 1:10, soaking for 15min at 50 ℃, filtering, removing filtrate, and freeze-drying to obtain pretreated noni powder; mixing the pretreated noni fruit powder with water according to a mass ratio of 1:20, adjusting pH to 5.2, performing ultrasonic-assisted extraction at 80 ℃ for 90min, centrifuging, collecting supernatant, and freeze-drying; the ultrasonic frequency is 20kHz, and the ultrasonic power is 100mW/mL.
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