A kind of preparation method and application of the black angle orchid extract of corona treatment
Technical field
The invention belongs to antibacterial agent and food additives fields, and in particular to a kind of black angle orchid extraction of corona treatment
The preparation method and application of object.
Background technique
Black angle blue (Origanum majorana) also known as Ma Yucao, are under the jurisdiction of Labiatae Origanum, originate in Mediterranean
Area is distributed in Turkey, the ground such as Egypt.Its blue warm-natured, mildly bitter flavor of black angle, has invigorating qi and refreshing, and heal invigorating the spleen, and cough-relieving is flat
Asthma, aid digestion, dissipate-swelling gas, lissive effect.In addition, black angle orchid fragrant odour, the fragrance with sweet tea pine and citrus is common
Spice.Stem and Ye Jun contain volatile oil, and main component is Thymol (Thymol) and carvacrol (Carvacrol).
Currently used ink angle orchid extracting method is mainly water extraction and alcohol extracting method, such as methanol, ethyl alcohol.However these extracting methods are deposited
The disadvantages of extraction efficiency is low, serious waste of resources, weak safety.Therefore, present invention proposition is mentioned using pulsed magnetic field technology
The extraction efficiency of high ink angle orchid, reduces the wasting of resources, enhances its safety.
Pulsed magnetic field technology is a kind of very promising new technique.Pulsed magnetic field is the narrow arteries and veins an of low frequency, high field intensity
Magnetic field, generally 1Hz~1.25Hz or so are rushed, head surface highest field strength is up to 3000Gs or more, pulse width 7ms or so.
In medical field, pulsed magnetic field is using the intracorporal blood of people, body fluid, cell medium etc. as conductor, by magnetic field strength, pulse frequency
Rate, polar variation act on body, and without fuel factor, will not generate tissue inflammatory exudation, general magnetic fields are up to body
Interior 150~200mm.Pulsed magnetic field can be with sedation-analgesia, reduction blood viscosity etc., clinical application many years, securely and reliably, patient
Without sense of discomfort.In field of food, pulsed magnetic field can destroy the eucaryotic cell structure of bacterium as non-thermal sterilization technology at a lower temperature
And the microorganism in food is killed, therefore be able to maintain the original flavor of food, color and nutritional ingredient.
Plasma is the aggregate of approximate electroneutral, is made of ion and electronic population, and can be to electric and magnetic fields
It responds.Plasma technique is raw using the certain industry of partially ionized plasma completion of plasma generator generation
Produce the means of target.Research, which has shown that corona treatment not only, has the energy mechanism of ionising radiation, but also there are also quality
Sunk-fund effect and charge-exchange effect.Corona treatment is rapid, and reaction temperature is low, and environment friendly and pollution-free.Surface in 2008 etc.
Gas ions photocatalysis is also formally suggested, this series of proof all shows formedness of the surface plasma in photocatalysis
Energy.Using plasma high temperature or in which active particle and radiation facilitate certain chemical reactions, to obtain new substance
It is more popular research hotspot.
Summary of the invention
It is an object of the invention to overcome technological deficiency existing in the prior art, such as: extraction efficiency is low, and the wasting of resources is tight
It is heavy, safety is weak etc., the present invention provides a kind of preparation methods of the black angle orchid extract of corona treatment.
Specific preparation method of the invention, comprising the following steps:
(1) preparation of black angle orchid alcohol extract:
The leaf cleaning of black angle orchid is clean, and soaked in absolute ethyl alcohol is added in shearing, drying, and it is blue to obtain black angle for shaking extraction
Alcohol extract;
(2) preparation of black angle orchid extract:
It by black angle orchid alcohol extract, is placed in pulsed magnetic field instrument and carries out pulse magnetic field treatment, filter, concentration obtains ink
Angle orchid extract;
(3) corona treatment ink angle orchid extract:
Black angle orchid extract is placed in plasma spectrometer, is handled, corona treatment ink angle orchid is obtained and mentions
Take object.
In step (1), the temperature of the shaking extraction is 25 DEG C, and the time of the shaking extraction is for 24 hours;
In step (2), when the pulse magnetic field treatment, temperature is 4~25 DEG C, and pulse strength is 1.0~4.0T, umber of pulse
It is 10~40;
In step (3), when the processing, protective gas is nitrogen, and nitrogen flow 100sccm or protective gas are
Air;Plasma spectrometer power is 300~600W, handles 2~6min of time.
The present invention also provides a kind of application of corona treatment ink angle orchid extract, corona treatment ink angles
Preservation and freshness of the blue extract for fruit and vegetable and meat.
Compared with prior art, beneficial effects of the present invention embody as follows:
(1) black angle orchid extract is prepared by pulsed magnetic field technology in the present invention, with existing water extraction and alcohol extracting method phase
Compare, have extraction efficiency high, the advantages that the wasting of resources can be effectively reduced, increase black angle orchid extract safety.Utilize arteries and veins
Rush magnetic field action temperature and, short processing time, break born of the same parents' high efficiency and it is without side-effects the features such as, the present invention is used to accelerate black angle blue
The extraction process of ingredient obtains more effective, safer black angle orchid extract.
(2) the black angle orchid extract in the present invention Jing Guo corona treatment is to foodborne bacterial pathogens and nosocomial infection cause of disease
Bacterium has more efficient, more significant antibacterial effect.Be embodied in its with smaller minimal inhibitory concentration (0.0625%~
And minimum bactericidal concentration (0.125%~0.25%) 0.25%).The minimum suppression of not plasma-treated black angle orchid extract
Bacteria concentration is 0.25%~0.5%, and minimum bactericidal concentration is 0.25%~1.0%.
(3) corona treatment ink prepared by the present invention angle orchid extract has in the preservation and freshness of fruit and vegetable and meat
There is good fungistatic effect, such as the application in cucumber, beef.In embodiment 4, corona treatment ink angle orchid extract is answered
In beef, compared with black angle orchid extract, Escherichia coli after the orchid extract-treated of corona treatment ink angle and golden yellow
The staphylococcic thalline quantity decline of color is obvious.In 96h, the sterilizing rate of Escherichia coli and staphylococcus aureus is respectively reached
76.55% and 85.19%.
(4) present invention combines black angle orchid extract with plasma technique, and the antibacterial for improving black angle orchid extract is living
Property, expand its application in food.Corona treatment ink obtained angle orchid extract can effectively inhibit food-borne
The growth of opportunistic pathogen and pathogenic bacteria of nosocomial infection, effectively prevention food posioning, ensuring food safety property extend food shelf
Phase.
Detailed description of the invention
Fig. 1 is the antibacterial of corona treatment ink angle orchid extract of the invention to Escherichia coli and staphylococcus aureus
Effect;
Fig. 2 is corona treatment ink angle orchid extract of the invention to Escherichia coli in cucumber and staphylococcus aureus
Antibacterial effect;
Fig. 3 is corona treatment ink angle orchid extract of the invention to Escherichia coli in beef and staphylococcus aureus
Antibacterial effect.
Specific embodiment
Illustrate a specific embodiment of the invention, but protection content of the invention by following Examples, is not only limited to this.
Embodiment 1:
Experimental material:
(1) preparation of black angle orchid alcohol extract:
10g ink angle orchid blade is weighed, is cleaned up, is sheared, dries, with 40mL soaked in absolute ethyl alcohol, wherein black angle orchid leaf
The ratio between quality volume of piece and dehydrated alcohol is 2:8, is placed in shaking extraction in 25 DEG C of shaking baths and for 24 hours, obtains black angle orchid second
Alcohol leaching liquor.
(2) preparation of black angle orchid extract:
Black angle orchid alcohol extract is placed in pulsed magnetic field instrument, temperature is 4 DEG C, pulse strength 1.0T, and umber of pulse is
10;Then black angle orchid alcohol extract is filtered, and filtrate is concentrated in Rotary Evaporators, black angle orchid extract is obtained, mentions
Taking rate is 61.38%.
(3) corona treatment ink angle orchid extract:
Black angle orchid extract is placed in small beaker, then black angle orchid extract is placed in plasma spectrometer, into
Row processing, nitrogen are protective gas, and nitrogen flow 100sccm, plasma power 300W handle time 2min, obtain
The black angle orchid extract of gas ions processing.
The present invention promotees to propose effect by the broken born of the same parents of pulsed magnetic field, 42.19% promotion that recovery rate is extracted by ethyl alcohol to
61.38%, obtain more comprehensively, deeper into extraction;Black angle orchid extract is placed in plasma spectrometer, by it is equal from
Daughter processing, improves antibacterial activity.
Embodiment 2:
(1) preparation of black angle orchid alcohol extract:
10g ink angle orchid blade is weighed, is cleaned up, is sheared, dries, with 40mL soaked in absolute ethyl alcohol, wherein black angle orchid leaf
The ratio between quality volume of piece and dehydrated alcohol is 2:8, is placed in shaking extraction in 25 DEG C of shaking baths and for 24 hours, obtains black angle orchid second
Alcohol leaching liquor.
(2) preparation of black angle orchid extract:
Black angle orchid alcohol extract is placed in pulsed magnetic field instrument, temperature is 12 DEG C, pulse strength 2.0T, and umber of pulse is
30;Then black angle orchid alcohol extract is filtered, and filtrate is concentrated in Rotary Evaporators, black angle orchid extract is obtained, mentions
Taking rate is 75.26%.
(3) corona treatment ink angle orchid extract:
Black angle orchid extract is placed in small beaker, then black angle orchid extract is placed in plasma spectrometer, into
Row processing, nitrogen are protective gas, and nitrogen flow 100sccm, plasma power 500W handle time 4min, obtain
The black angle orchid extract of gas ions processing.
The present invention promotees to propose effect by the broken born of the same parents of pulsed magnetic field, 42.19% promotion that recovery rate is extracted by ethyl alcohol to
75.26%, obtain more comprehensively, deeper into extraction;Black angle orchid extract is placed in plasma spectrometer, by it is equal from
Daughter processing, improves antibacterial activity.
Embodiment 3:
(1) preparation of black angle orchid alcohol extract:
10g ink angle orchid blade is weighed, is cleaned up, is sheared, dries, with 40mL soaked in absolute ethyl alcohol, wherein black angle orchid leaf
The ratio between quality volume of piece and dehydrated alcohol is 2:8, is placed in shaking extraction in 25 DEG C of shaking baths and for 24 hours, obtains black angle orchid second
Alcohol leaching liquor.
(2) preparation of black angle orchid extract:
Black angle orchid alcohol extract is placed in pulsed magnetic field instrument, temperature is 25 DEG C, pulse strength 4.0T, and umber of pulse is
40;Then black angle orchid alcohol extract is filtered, and filtrate is concentrated in Rotary Evaporators, black angle orchid extract is obtained, mentions
Taking rate is 96.61%.
(3) corona treatment ink angle orchid extract:
Black angle orchid extract is placed in small beaker, then black angle orchid extract is placed in plasma spectrometer, with
Air is that protective gas is handled, and plasma power 600W handles time 6min, obtains corona treatment ink angle orchid
Extract.
The present invention promotees to propose effect by the broken born of the same parents of pulsed magnetic field, 42.19% promotion that recovery rate is extracted by ethyl alcohol to
96.61%, obtain more comprehensively, deeper into extraction;Black angle orchid extract is placed in plasma, plasma is passed through
Processing improves antibacterial activity.
Embodiment 4:
(1) fungistatic effect of the corona treatment ink angle orchid extract to foodborne bacterial pathogens and pathogenic bacteria of nosocomial infection
And bactericidal effect (1) experimental strain (the entitled italic of strain)
A. foodborne bacterial pathogens:
Escherichia coli: Escherichia coli IFO;
Staphylococcus aureus: Staphyloccocus aureus;
Bacterium enteritidis: Salmonella Enteritidis;
Bacillus subtilis: Bacillus cereus;
Hay bacillus: Bacillus subtilis;
Listeria monocytogenes: Listeria monocytogenes;
Clostridium botulinum: Clostridium botulinum.
B. pathogenic bacteria of nosocomial infection:
Methicillin-resistant staphylococcus aureus: MRSA (Methicillin-resistant Staphylococcus
aureus)。
(2) experimental method
It adds corona treatment ink angle orchid extract respectively into sterile vegetative meat soup (NB) culture solution and black angle orchid mentions
Object is taken, making its mass fraction is respectively 0.0625%, 0.125%, 0.25%, 0.5%, 1.0% (w/v).Use micropipettor
The bacterium solution of culture to logarithmic phase, initial concentration 10 is added5-6CFU/mL.The culture solution of inoculation is placed in 37 DEG C of incubators and cultivates
24~48h, wherein Listeria monocytogenes are placed in 25 DEG C of incubators and cultivate 48h.
Minimal inhibitory concentration (Minimum Inhibitory Concentration, MIC) refers to be incubated in certain circumstances
Afterwards, the Cmin of bacterial reproduction can not be distinguished with the naked eye.The culture solution for the minimal inhibitory concentration that will be greater than or equal to is respectively with inoculation
Ring dips a ring, crosses on sterile NA culture medium, is equally placed in 24~48h of culture in 37 DEG C and 25 DEG C of incubators.In nutrition
The concentration that can't see bacterium colony growth on agar (NA) culture medium completely is set to minimum bactericidal concentration (Minimum Bactericidal
Concentration, MBC).
(3) experimental result
Sample of the present invention is corona treatment ink angle orchid extract, is a kind of novel antibacterial agent.In order to judge etc. from
Daughter handles the anti-microbial property of black angle orchid extract, as a control group by black angle orchid extract, in foodborne bacterial pathogens and institute
The minimal inhibitory concentration and minimum bactericidal concentration of infection pathogen are tested.As shown in Table 1, black angle orchid extract is to each disease
The minimal inhibitory concentration of opportunistic pathogen is 0.25%~0.5%, and minimum bactericidal concentration is 0.25%~1.0%;And corona treatment
Black angle orchid extract is 0.0625%~0.25% to the minimal inhibitory concentration of each pathogen, minimum bactericidal concentration 0.125%
~0.25%.Therefore, corona treatment ink angle orchid extract has more significant antibacterial and sterilization effect than black angle orchid extract
Fruit illustrates that corona treatment can be improved the antibacterial activity of black angle orchid extract.
1 corona treatment ink angle orchid extract of table is to the fungistatic effect of food-borne germ and pathogenic bacteria of nosocomial infection and kills
Bacterium effect
(2) antibacterial experiment of corona treatment ink angle orchid extract
(1) experimental strain (the entitled italic of strain)
Escherichia coli: Escherichia coli;
Staphylococcus aureus: Staphyloccocus aureus;
(2) experimental method
Using the method for plate culture count, with Escherichia coli (Escherichia coli) and staphylococcus aureus
(Staphylococcus aureus) is type culture, measures the anti-microbial property of corona treatment ink angle orchid extract.
Phosphate buffer (PBS) solution is dispensed into test tube in equal volume, high pressure sterilization is spare.By corona treatment ink angle orchid
Extract and black angle orchid extract are added in test tube with the concentration that final mass score is 0.50% (w/v), and access culture
To the Escherichia coli and staphylococcus aureus of logarithmic phase, initial concentration 105CFU/mL, not add the test tube of sample as sky
White control group is uniformly mixed, is placed on 37 DEG C of shaking table culture.In 0h, for 24 hours, when 48h, take out test tube, step by step dilution spread in
On NA culture medium, remaining bacterium number is measured.Measuring method is total according to 4789.2-2010 food hygiene microbiologic inhibition tests bacterium colony of GB
Number carries out.
(3) experimental result
It will be seen from figure 1 that in 48h, by the Escherichia coli and staphylococcus aureus of black angle orchid extract-treated
Remaining bacterium number have dropped 59.15%, 67.11% respectively;Meanwhile corona treatment ink angle orchid extract to Escherichia coli and
Staphylococcus aureus has good bactericidal effect, and remaining bacterium number has dropped 91.83%, 99.999% respectively.It follows that waiting
The black angle orchid extract of gas ions processing has significantly more bactericidal effect, will have wider purposes in the food industry.
(3) application of the corona treatment ink angle orchid extract in cucumber
(1) experimental strain (the entitled italic of strain)
Escherichia coli: Escherichia coli;
Staphylococcus aureus: Staphyloccocus aureus;
(2) experimental method
Using the method for plate culture count, with Escherichia coli (Escherichia coli) and staphylococcus aureus
(Staphylococcus aureus) is type culture, measures application of the corona treatment ink angle orchid extract in cucumber.
Fresh cucumber is bought, cleans, dices.Cucumber is mixed with phosphate buffer (PBS) with mass ratio 1:9, is transferred to
Sterile homogenizing bag, homogeneous 5min, filtering, Aspirate supernatant to get mass fraction 10% (w/w) cucumber culture solution, in equal volume
Test tube is dispensed, it is spare after high pressure sterilization.By corona treatment ink angle orchid extract and black angle orchid extract with final mass point
Number is that the concentration of 0.50% (w/v) is added in test tube, and accesses the Escherichia coli cultivated to logarithmic phase and Staphylococcus aureus
Bacterium, initial concentration 103CFU/mL is uniformly mixed using not adding the test tube of sample as blank control group, is placed on 37 DEG C and shakes
Bed culture.In 0h, for 24 hours, 48h, 72h when 96h, take out test tube, and dilution spread is on NA culture medium, measuring remaining bacterium number step by step.
Measuring method is carried out according to 4789.2-2010 food hygiene microbiologic inhibition tests total plate count of GB.
(3) experimental result
Figure it is seen that the Escherichia coli and staphylococcus aureus of blank assay group are rapid in cucumber culture solution
Breeding, total clump count increase to 7.91Log CFU/mL by 3.20Log CFU/mL.In black angle orchid extract-treated groups, large intestine bar
The growth of bacterium and staphylococcus aureus receives a degree of inhibition, the phenomenon that rapid growth breeding does not occur.In 96h
When, the clump count of Escherichia coli and staphylococcus aureus is respectively 4.25Log CFU/mL and 3.45Log CFU/mL.Equally
Ground, in corona treatment ink angle orchid extract group, the growth and breeding of bacterium has obtained further inhibition.In 96h, large intestine
The clump count of bacillus and staphylococcus aureus is respectively that 1.1Log CFU/mL and 0.72Log CFU/mL, sterilizing rate reach
85.53% and 90.79%.Therefore, corona treatment ink angle orchid extract has significantly more bactericidal effect in cucumber,
Help to ensure food safety, extends Food Shelf-life.
(4) application of the corona treatment ink angle orchid extract in beef
(1) experimental strain (the entitled italic of strain)
Escherichia coli: Escherichia coli;
Staphylococcus aureus: Staphyloccocus aureus;
(2) experimental method
Using the method for plate culture count, with Escherichia coli (Escherichia coli) and staphylococcus aureus
(Staphylococcus aureus) is type culture, measures application of the corona treatment ink angle orchid extract in beef.
Fresh beef is bought, is cleaned.Beef is mixed with phosphate buffer (PBS) with mass ratio 1:9, is transferred to sterile equal
Matter bag, homogeneous 5min, filtering, beef culture solution of the Aspirate supernatant to get mass fraction 10% (w/w), isometric packing examination
It manages, it is spare after high pressure sterilization.It is with final mass score by corona treatment ink angle orchid extract and black angle orchid extract
The concentration of 0.50% (w/v) is added in test tube, and accesses the Escherichia coli and staphylococcus aureus of culture to logarithmic phase, just
Beginning concentration is 103CFU/mL is uniformly mixed using the test tube for not adding antibacterial agent as blank control group, is placed on 37 DEG C of shaking table
Culture.In 0h, for 24 hours, 48h, 72h when 96h, take out test tube, and dilution spread is on NA culture medium, measuring remaining bacterium number step by step.It surveys
Determine method to carry out according to 4789.2-2010 food hygiene microbiologic inhibition tests total plate count of GB.
(3) experimental result
From figure 3, it can be seen that the Escherichia coli and staphylococcus aureus of blank assay group are rapid in beef culture solution
Breeding, total clump count increase to 8.15Log CFU/mL by 3.24Log CFU/mL.In black angle orchid extract-treated groups, large intestine bar
The speed of growth of bacterium and staphylococcus aureus slows down.In 96h, the clump count of Escherichia coli and staphylococcus aureus is distinguished
For 4.59Log CFU/mL and 4.23Log CFU/mL.Similarly, in corona treatment ink angle orchid extract group, the life of bacterium
Long breeding has obtained further inhibition.In 96h, the clump count of Escherichia coli and staphylococcus aureus is respectively
1.94Log CFU/mL and 1.23Log CFU/mL, sterilizing rate reach 76.55% and 85.19%.Therefore, corona treatment ink
Angle orchid extract has significantly more bactericidal effect in beef, can effectively prevent food posioning, ensures food
Safety extends Food Shelf-life.