CN101711969B - Device and method for biological processing with pulsed multi-frequency ultrasonic waves in sweep-frequency mode and use thereof - Google Patents
Device and method for biological processing with pulsed multi-frequency ultrasonic waves in sweep-frequency mode and use thereof Download PDFInfo
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Abstract
The invention discloses a device and a method for biological processing with pulsed multi-frequency ultrasonic waves in a sweep-frequency mode and use thereof, which relate to the field of agricultural product deep processing. The device consists of an ultrasonic wave processing system, an ultrasonic wave generating system and a computer control system, which are sequentially connected by cables, wherein the ultrasonic wave processing system consists of an upper ultrasonic wave transducer, a lower ultrasonic wave transducer, an ultrasonic wave processing tank body, a circulating pump and an upper oscillatory plate height adjusting screw rod; an ultrasonic wave generator set consists of a group of pulsed ultrasonic wave generators at different frequencies; and the control system consists of an ultrasonic wave controller, a temperature controller and a computer. The equipment can be used in ultrasonic wave-assisted extraction of active ingredients of natural products and the extraction efficiency is improved 10 to 90 percent. The equipment can be used in ultrasonic wave-assisted enzymolysis of biological macromolecules, wherein the ultrasonic wave-assisted enzymolysis includes preprocessing raw materials, preprocessing hydrolases and performing enzymolysis in an ultrasonic wave field; and the enzymolysis efficiency is improved by 25 to 65 percent.
Description
Technical field
The present invention relates to agricultural product intensive processing field, relate in particular to a kind of pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode and in the application in agricultural product intensive processing field.
Background technology
Usually be frequency that the ripple of 20kHz～10MHz is called ultrasonic wave; It is a kind of elastic mechanical ripple of in medium, propagating; Ultrasonic technology is as a kind of physical means and instrument, can in chemical reaction medium commonly used, produce a series ofly to approach extreme condition, as produce local and high temperature moment, high pressure etc.; This energy not only can excite or promote chemical reaction, accelerate chemical reaction velocity, even can also change the direction of some chemical reaction.Fall at extraction separation, the preparation of biologically active peptide, the polysaccharide of active ingredient of natural product in recent years and the field such as gather and obtained using comparatively widely.
A large amount of researchs show that ultrasonic wave promotes that chemical reaction is relevant with acoustic cavitation effectiveness, mechanical effect and fuel factor.Acoustic cavitation effectiveness is meant when the ultrasonic wave of certain intensity is propagated in liquid medium; Be present in the micro-bubble in the liquid; Under the effect of ultrasonic field, be activated; Show as a series of dynamic processes such as formation, vibration, growth, contraction and even collapse of complex, and the physics and the chemical effect that cause.
Hu Aijun etc. (" the phonochemistry yield progress of multiple frequency ultrasonic irradiation "; " acoustic technique " 22 volume 1 phases (2003), 62 pages) be described in the frequency range such between 20kHz～60kHz, supersonic frequency increases; The sonochemistry reaction power strengthens, and the sonochemistry yield increases thereupon; The phonochemistry yield that the composite ultraphonic radiation of double frequency or multifrequency produces can significantly promote chemical reaction, accelerate chemical reaction velocity much larger than the phonochemistry yield sum of two single-frequency difference radiation.Research is also found to select for use batch (-type) to launch hyperacoustic mode, can strengthen ultrasonic treatment effect.In running order when ultrasonic transmitter, biological cell receives strong ultrasonic vibration; When remaining static, system homogenizing, cell relax.The step stressed destruction of strengthening pair cell, help the carrying out of mass transport process, also help lend some impetus to chemical reaction, accelerate chemical reaction velocity.
Chinese invention patent " triple-frequency ultrasonic extraction device and method for using thereof " (ZL200610124003.1) adopts that three kinds of frequencies are ultrasonic works simultaneously, confirms that really it is to the extraction efficiency of the Chinese herbal medicine effective ingredients ultrasonic Extraction equipment apparently higher than single-frequency.This patent has been used the probe type ultrasonic wave producer, and the weak point of this generator is needing in the laboratory only to be suitable for ultrasound intensity extraction material big, that treating capacity is few; Detecting head surface can receive cavitation corrosion, can reduce hyperacoustic luminous efficiency, and the cavitation corrosion thing can be fallen in the volume, can pollute extract solution; There is ultrasonic field problem pockety in liquid.
In order to overcome single-frequency and the multiple deficiency that exists frequently; Zhang Shuce has designed a kind of sweep-frequency B ultrasonic generator (swept frequency range 28～73kHz); Be used for research (Shandong Textile science and technology to the ramee degumming effect; 2006 the 3rd phases), said here " frequency sweep " refers to operate carrying out a frequency sweep in a certain bigger frequency band at the appointed time, promptly sends the continuous ultrasound of different frequency at the appointed time." sweep-frequency B " ultrasonic wave of this article definition is because the frequency range wide coverage; Purpose is to make most cavitation nucleus reach best cavitation effect in the short period of time; Thereby can play the removal effect to the glue impurity in the big small gaps of difference simultaneously; Shorten the ultrasonic wave usually time greatly, improved degumming effect.
Although present ultrasonic treatment unit used multifrequency compound or (with) the service intermittent mode, it is most that what use is fixed frequency.For the biological processing system of a complicacy, fixing supersonic frequency is difficult to excite treatment system to produce the resonant frequency that is complementary with himself intrinsic frequency, can not give full play to the due action effect of ultrasonic wave.Present " sweep-frequency B " ultrasonic wave in principle, also just the frequency spectrum of vibrations object wideer, effect is more, does not reach the effect that farthest reaches resonance.
Summary of the invention
The objective of the invention is in order to overcome the deficiency in the above-mentioned existing ultrasonic generator; Multifrequency combination, the pulsating ultrasound of developing a kind of frequency sweep mode of operation takes place, key parameter is monitored in real time and the multifunction ultrasonic bioprocess apparatus of new generation of control; The ultrasonic assistance enzymolysis and the modification that can be applicable to ultrasonic assisted extraction, the large biological molecule of active ingredient of natural product are handled, and to the ultrasonotomography of residues of pesticides etc.
" sweep-frequency B " ultrasonic wave of the present invention's design is meant that ultrasonic wave fluctuates up and down (shown in Figure of description 1) around centre frequency with certain frequency sweep cycle in a narrower scope, represent (a is a centre frequency, and δ is amplitude of variation up and down) with a ± δ kHz.In a frequency sweep cycle, frequency of ultrasonic increases to a+ δ kHz with certain speed from a-δ kHz, reduces to a-δ kHz with identical speed from a+ δ kHz again, is a kind of frequency sweep mode of isosceles triangle.No matter sweep on being; Still sweep under; When ultrasonic wave all can produce the peak value of an output energy during through centre frequency a to the effect system, will produce a batch (-type) effect energy pulse that doubles frequency sweep cycle like this, when the resonant frequency of ultrasonication object equals or approach 2 times of frequency sweep cycles; System will produce the vibration of large amplitude, causes that effective object produces strong vibration.Need seek best frequency sweep cycle according to the difference of ultrasonication object at work, purpose is to make system realize maximum sonic oscillation with the minimal power input.The present invention realizes through following dual mode the change of frequency sweep cycle: the size of scanning constant rapid change change of frequency amplitude δ, fixedly the δ value changes sweep speed.The sweep-frequency B ultrasonic wave of the present invention design helps exciting treatment system to produce the resonant frequency that is complementary with himself resonant frequency, has given full play to the effect of ultrasonic wave effect, has overcome fixedly supersonic frequency in the defective that exists aspect this.The present invention also comprise in addition a kind of from high frequency to low frequency and a kind of from low frequency to high frequency, carry out the frequency sweep mode (like Fig. 2 and Fig. 3) of the dull scanning of right angled triangle.
The ultrasonic wave combination of two different center frequency of the present invention's design is to shaking, and the stack of transmitted beam ripple produces the beat frequency effect, and the effect (shown in Figure of description 4) of amplitude stack is arranged.So not only can improve the sound intensity, but also can avoid the generation of standing wave, also help lend some impetus to chemical reaction, accelerate chemical reaction velocity.
The pulsating ultrasound emergence pattern of the present invention's design not only helps strengthening the destruction to biological cell; Also help reducing the rising of temperature and the energy consumption of the system of saving; Can make ultrasonic treatment unit avoid being in the long duty, influence the life-span of supersonic generator.
The dynamic tracking of major parameter, monitoring, adjustment and control can accurately be adjusted running parameter in real time according to the working condition of system, guarantee optimum working efficiency.
Technical scheme of the present invention:
The pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode: be made up of ultrasonic Treatment system, ultrasonic wave generation systems and computer control system, this three part is connected in sequence by cable.
Described ultrasonic Treatment system by last ultrasonic transducer promptly go up vibration plate, ultrasonic transducer promptly descend vibration plate down, ultrasonic Treatment casing, circulating pump, circulation valve, discharging valve, water-bath heat-conductive water, electrothermal tube, transducer height adjustment screw rod composition.Wherein install electrothermal tube in the ultrasonic Treatment casing, can satisfy sonicated liquid different temperatures and set; Following vibration plate is positioned at the bottom of ultrasonic Treatment casing; The symmetry placement up and down of last vibration plate and following vibration plate, last vibration plate is fixed through transducer height adjustment screw rod, and carries out up and down when shaking; Distance between the vibration plate is adjusted through transducer height adjustment screw rod up and down; Circulating pump is positioned at the outside of ultrasonic Treatment casing, and the inside through conveyance conduit and valve and ultrasonic Treatment casing links together, and in the lower right corner of ultrasonic Treatment casing liquid outlet is set; From the upper left corner of casing inlet is set, the lower right corner part of ultrasonic Treatment casing 3 is equipped with circulation valve and discharging valve.
Wherein said ultrasonic transducer (going up vibration plate) and following ultrasonic transducer (following vibration plate), its centre frequency is 24kHz～68kHz, the amplitude δ that changes up and down is 0～5kHz; Carry out up and down when shaking, last vibration plate is different with the centre frequency of following vibration plate;
Circulating pump is positioned at the outside of ultrasonic Treatment casing; Link together through conveyance conduit and valve and ultrasonic Treatment cell body inside; The lower right corner at the ultrasonic Treatment casing is provided with liquid outlet; From the upper left corner of casing inlet is set,, returns from the upper left corner of cell body from the lower right corner fluid of ultrasonic Treatment cell body.The lower right corner part of ultrasonic Treatment cell body is equipped with circulation valve and discharging valve, when after hours, closes circulation valve, opens the discharging valve, emits material solution.Circulating pump is used to realize being processed circulating of material solution, strengthens convective mass transfer, improves the sonicated effect.
Described ultrasonic wave generation systems is exactly the supersonic generator group; The supersonic generator group is made up of one group of different frequency supersonic generator; Its corresponding ultrasonic transducer of the supersonic generator of each frequency is corresponding, is used to control the excusing from death wave transducer and sends ultrasonic wave according to ultrasonic time and intermittent time, the whole ultrasonic processing time of the power that sets, frequency sweep cycle, frequency sweep fabric width (± δ kHz), impulse ultrasound; When carrying out the sweep parameters setting, be example with δ=2kHz, change of frequency amplitude (δ) can be set arbitrarily between 0～2kHz; Frequency sweep cycle (promptly accomplish a secondary frequencies and increase to a+ δ kHz from a-δ kHz, reduce to the required time of a-δ kHz from a+ δ kHz again) can change arbitrarily between 1ms～1000ms.When δ=0, then be common fixed frequency mode of operation.The main setup parameter of pulsating ultrasound mode of operation comprises ultrasonic time and intermittent time, and ultrasonic time and intermittent time all can be adjusted between 1～1000s.
Described computer control system is by the ultrasonic wave controller, and temperature controller and computer are formed; Link together by cable; The ultrasonic time of the power of computer through ultrasonic wave controller control each particular centre frequency (a kHz) supersonic generator, frequency sweep cycle, frequency sweep fabric width (± δ kHz), impulse ultrasound and intermittent time, whole ultrasonic processing time.Computer is through the bath temperature of temperature controller control ultrasonic Treatment cell body.Can set above-mentioned all parameters through computer.
Method based on the pulsed multi-frequency ultrasonic bioprocess apparatus of above-mentioned frequency sweep mode:
Can be according to the different choice of the amount of production scale in the actual production and the material handling logarithm of vibration plate up and down, the combination of four vibration plates can realize following working method up and down:
(1) single-frequency frequency sweep mode: i.e. two identical following vibration plates of placement frequency at the bottom of the case, inject the material solution of certain depth, carry out the frequency sweep sonicated;
(2) multifrequency frequency sweep mode: i.e. placement frequency different two and with vibration plate up and down at the bottom of the case, inject the material solution of certain depth, carry out double frequency and multifrequency frequency sweep sonicated;
(3) double frequency is to the frequency sweep mode of shaking: i.e. two identical following vibration plates of placement frequency at the bottom of the case; Inject the material solution of certain depth; Be close to vibration plate on identical two of solution surface placement frequency, last vibration plate frequency is different from vibration plate down, carries out double frequency to the frequency sweep sonicated of shaking.
Wherein said alternative centre frequency (a) is 24kHz～68kHz, and the amplitude (δ) that changes up and down is 0～5kHz; When carrying out the sweep parameters setting, change of frequency amplitude (δ) can be set arbitrarily between 0～5kHz; Frequency sweep cycle is promptly accomplished a secondary frequencies and is increased to a+ δ kHz from a-δ kHz, reduces to the required time of a-δ kHz from a+ δ kHz again and can be between 1ms～1000ms changes arbitrarily; When δ=0, then be common fixed frequency mode of operation; The main setup parameter of pulsating ultrasound mode of operation comprises ultrasonic time and intermittent time, and ultrasonic time and intermittent time all can be adjusted between 1～1000s.
The application of frequency sweep mode pulsed multi-frequency ultrasonic bioprocess apparatus of the present invention comprises: the ultrasonic assisted extraction of active ingredient of natural product, the ultrasonic assistance enzymolysis of large biological molecule etc., the ultrasonic assistance enzymolysis of described large biological molecule comprise preliminary treatment to raw material, carry out enzyme digestion reaction to the preliminary treatment of hydrolase with in ultrasound field.In the extraction of active ingredient of natural product, the frequency sweep mode pulsed multi-frequency ultrasonic can be realized the maximum extraction efficiency of active ingredient in the situation of minimum power input.About the ultrasonic assistance enzymolysis of large biological molecule, in preliminary treatment to raw material, ultrasonic wave through improving raw material dissolubility and to the influence of raw material mix, improve follow-up enzymolysis efficiency; In the preliminary treatment to hydrolase, suitable ultrasonic wave can significantly improve the activity of enzyme; In ultrasonic field, carry out enzymolysis, help quickening the touch opportunity of enzyme and large biological molecule, improve the reacting dynamics characteristic of enzymolysis process, improve reaction efficiency.
The course of work of the pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode of the present invention is following: put into vibration plate 2 down in treatment trough bottom, the material solution that injects mixings to cell body 3 is behind certain depth; According to the programme of work of setting, when needs, be close to the liquid level of material solution and upward vibration plate 1 of placement, go up the distance between vibration plate 1 and the following vibration plate 2 through last vibration plate height adjustment screw rod 15 adjustment; The ultrasonic time of the power through the computer settings supersonic generator, frequency sweep cycle, frequency sweep fabric width (± δ kHz), impulse ultrasound and idle hours, whole ultrasonic processing time, the temperature of water-bath, the rotating speed of circulating pump; Open electrothermal tube 6, material solution is heated through water-bath; After being heated to the operating temperature of setting, opening circulation valve 7 and carry out the liquid circulation, open supersonic generator and begin to carry out sonicated with ON cycle pump 4.Behind the end-of-job, close circulation valve 7, open discharging valve 8, emit material solution.
This equipment shows in the application study of agricultural product intensive processing invention, when being applied to the extraction of active ingredient of natural product, compares with present similar technology, and extraction efficiency improves 10-90%; When being applied to proteolysis and preparing polypeptide, with present similar technology relatively, enzymolysis efficiency improves 25-65%.
Description of drawings
Fig. 1 is the supersonic frequency change curve (the frequency sweep waveform is isosceles triangle) of frequency sweep mode;
Fig. 2 is the supersonic frequency change curve (the frequency sweep waveform is dull and descends) of frequency sweep mode,
Fig. 3 is the supersonic frequency change curve (the frequency sweep waveform is dull and rises) of frequency sweep mode,
Fig. 4 is the supersonic frequency change curve of double frequency combination to shaking,
Fig. 5 is the structural representation of frequency sweep mode pulsed multi-frequency ultrasonic bioprocess apparatus, and wherein 1-goes up ultrasonic transducer (going up vibration plate), ultrasonic transducer under the 2-(following vibration plate), 3-ultrasonic Treatment casing; The 4-circulating pump, 5-water-bath heat-conductive water, 6-electrothermal tube, 7-circulation valve; 8-discharging valve, 9, the 14-cable, 10-ultrasonic wave controller, 11-temperature controller; The 12-computer, 13-supersonic generator group, 15-transducer height adjustment screw rod.
The specific embodiment
Below in conjunction with specific embodiment and accompanying drawing the present invention is described in further detail, but the embodiment of describing among the present invention only is used to explain the present invention, and does not limit the application of this device.
The pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode: be made up of ultrasonic Treatment system, ultrasonic wave generation systems and computer control system, this three part is connected in sequence by cable.Ultrasonic Treatment system shown in Figure of description 5 is promptly gone up vibration plate 1, is descended ultrasonic transducer promptly to descend vibration plate 2, ultrasonic Treatment casing 3, circulating pump 4 by last ultrasonic transducer; Circulation valve 7, discharging valve 8, water-bath heat-conductive water 5; Electrothermal tube 6, transducer height adjustment screw rod 15 is formed.Wherein installing electrothermal tube 6 in the ultrasonic Treatment casing 3 can satisfy sonicated liquid different temperatures and set; Following vibration plate 2 is positioned at the bottom of ultrasonic Treatment casing 3; Last vibration plate and the symmetry placement up and down of following vibration plate; Last vibration plate 1 is fixed through transducer height adjustment screw rod 15; When carrying out up and down when shaking, the distance between the vibration plate can be adjusted through transducer height adjustment screw rod 15 up and down, satisfies the needs of differing heights liquid amount;
Wherein said ultrasonic transducer (going up vibration plate) 1 and following ultrasonic transducer (following vibration plate) 2, its centre frequency is 24kHz～68kHz, the amplitude (δ) that changes up and down is 0～5kHz; Carry out up and down when shaking, last vibration plate is different with the centre frequency of following vibration plate;
Can be according to the different choice of the amount of production scale in the actual production and the material handling logarithm of vibration plate up and down, the combination of four vibration plates can realize following working method up and down:
(1) single-frequency frequency sweep mode: i.e. two identical following vibration plates of placement frequency at the bottom of the case, inject the material solution of certain depth, carry out the frequency sweep sonicated;
(2) multifrequency frequency sweep mode: i.e. placement frequency different two and with vibration plate up and down at the bottom of the case, inject the material solution of certain depth, carry out double frequency and multifrequency frequency sweep sonicated;
(3) double frequency is to the frequency sweep mode of shaking: i.e. two identical following vibration plates of placement frequency at the bottom of the case; Inject the material solution of certain depth; Be close to vibration plate on identical two of solution surface placement frequency, last vibration plate frequency is different from vibration plate down, carries out double frequency to the frequency sweep sonicated of shaking.
Wherein said alternative centre frequency (a) is 24kHz～68kHz, and the amplitude (δ) that changes up and down is 0～5kHz; When carrying out the sweep parameters setting, change of frequency amplitude (δ) can be set arbitrarily between 0～5kHz; Frequency sweep cycle is promptly accomplished a secondary frequencies and is increased to a+ δ kHz from a-δ kHz, reduces to the required time of a-δ kHz from a+ δ kHz again and can be between 1ms～1000ms changes arbitrarily; When δ=0, then be common fixed frequency mode of operation; The main setup parameter of pulsating ultrasound mode of operation comprises ultrasonic time and intermittent time, and ultrasonic time and intermittent time all can be adjusted between 1～1000s.
Circulating pump 4 is positioned at the outside of ultrasonic Treatment casing 3, links together through conveyance conduit and valve and ultrasonic Treatment cell body inside, from the lower right corner fluid of ultrasonic Treatment cell body 3, returns from the upper left corner of cell body 3.The lower right corner part of ultrasonic Treatment casing 3 is equipped with circulation valve 7 and discharging valve 8, when after hours, closes circulation valve 7, opens discharging valve 8, emits material solution.Circulating pump 4 is used to realize being processed circulating of material solution, strengthens convective mass transfer, improves the sonicated effect.
Described ultrasonic wave generation systems is exactly a supersonic generator group 13; Supersonic generator group 13 is made up of one group of different frequency supersonic generator; Its corresponding ultrasonic transducer of the supersonic generator of each frequency is corresponding, is used to control the excusing from death wave transducer and sends ultrasonic wave according to ultrasonic time and intermittent time, the whole ultrasonic processing time of the power that sets, frequency sweep cycle, frequency sweep fabric width (± δ kHz), impulse ultrasound; When carrying out the sweep parameters setting, be example with δ=2kHz, change of frequency amplitude (δ) can be set arbitrarily between 0～2kHz; Frequency sweep cycle (promptly accomplish a secondary frequencies and increase to a+ δ kHz from a-δ kHz, reduce to the required time of a-δ kHz from a+ δ kHz again) can change arbitrarily between 1ms～1000ms.When δ=0, then be common fixed frequency mode of operation.The main setup parameter of pulsating ultrasound mode of operation comprises ultrasonic time and intermittent time, and ultrasonic time and intermittent time all can be adjusted between 1～1000s.
Computer control system is by ultrasonic wave controller 10, and temperature controller 11 is formed with computer 12.Link together by cable; The ultrasonic time of power through ultrasonic wave controller 10 control each particular centre frequency (a kHz) supersonic generators of computer 12, frequency sweep cycle, frequency sweep fabric width (± δ kHz), impulse ultrasound and intermittent time, whole ultrasonic processing time.Computer 12 temperature are through the bath temperature of controller 14 control ultrasonic Treatment cell bodies 3.Can set through 12 pairs of above-mentioned all parameters of computer.
Application---the ultrasonic assisted extraction of Porphyra yezoensis albumen of the present invention in active ingredient of natural product extracts.The dried laver of 1kg (containing protein 41 .3%) is crushed to behind 60 orders with behind the 40L water mixing adds the ultrasonic Treatment groove.The transducer that at the bottom of treatment trough, to put into two frequencies be 28kHz is being close to the transducer (at a distance of about 20cm) of placing two 20kHz above the liquid level.Input power through every transducer of computer settings be 600W (4 general power 2400W), frequency sweep cycle be 20ms, frequency sweep fabric width for the ultrasonic time 100s of ± 2kHz, impulse ultrasound, idle hours 3s, the flow 5L/min of circulating pump, extract initial temperature be 45 ℃.Stop ultrasonic behind the ultrasonic assisted extraction 80min.The centrifugal 10min of extract 3000r/min gets supernatant and measures wherein protein content, and the recovery rate that calculates albumen is 90.2%.
Check experiment: check experiment comprise no ultrasonic routine extract and do not have a frequency sweep decide the ultrasonic assisted extraction of frequency.It is same as described above that no ultrasonic routine is extracted other parameter; The ultrasonic assisted extraction of frequency of deciding of no frequency sweep is meant that with the frequency sweep fabric width is whole be ± 0kHz that other parameter is same as described above.Conventional extraction and the fixed protein extracting ratio of ultrasonic assisted extraction frequently are respectively 78.1% and 81.3%.Sweep-frequency B multifrequency pulse ultrasonic Extraction has increased by 15.5% and 10.9% respectively than conventional extraction and the fixed protein extracting ratio of ultrasonic assisted extraction frequently.
Application---the ultrasonic assisted extraction of rapeseed protein of the present invention in active ingredient of natural product extracts.1.8kg rapeseed cake dregs crude protein (containing albumen 48%) is crushed to behind 90 orders with behind the 30L water mixing adds the ultrasonic Treatment groove, adjustment pH to 9.8.The transducer that at the bottom of treatment trough, to put into two frequencies be 28kHz is being close to the transducer (at a distance of about 15cm) of placing two 24kHz above the liquid level.Input power through every transducer of computer settings be 600W (4 general power 2400W), frequency sweep cycle be 18ms, frequency sweep fabric width for the ultrasonic time 50s of ± 2kHz, impulse ultrasound, idle hours 5s, the flow 16L/min of circulating pump, extract initial temperature be 30 ℃.Stop ultrasonic behind the ultrasonic assisted extraction 30min.The centrifugal 10min of extract 3200r/min gets supernatant and measures wherein protein content, and the recovery rate that calculates albumen is 24.2%.
Check experiment: check experiment comprise no ultrasonic routine extract and do not have a frequency sweep decide the ultrasonic assisted extraction of frequency.It is same as described above that no ultrasonic routine is extracted other parameter; The ultrasonic assisted extraction of frequency of deciding of no frequency sweep is meant that with the frequency sweep fabric width is whole be ± 0kHz that other parameter is same as described above.Conventional extraction and the fixed protein extracting ratio of ultrasonic assisted extraction frequently are respectively 12.8% and 19.8%.Sweep-frequency B multifrequency pulse ultrasonic Extraction has increased by 89.1% and 22.2% respectively than conventional extraction and the fixed protein extracting ratio of ultrasonic assisted extraction frequently.
The ultrasonic assist in synchronization of the application of the present invention in active ingredient of natural product extracts---rice bran protein and polysaccharide is extracted.With adding the ultrasonic Treatment groove behind 200g defatted rice bran and the 20L water mixing.The transducer that at the bottom of treatment trough, to put into two frequencies be 24kHz and 64kHz.Input power through every transducer of computer settings be 600W, frequency sweep cycle be 36ms, frequency sweep fabric width for the ultrasonic time 200s of ± 2kHz, impulse ultrasound, idle hours 5s, the flow 2L/min of circulating pump, extract initial temperature be 45 ℃.Stop ultrasonic behind the ultrasonic assisted extraction 80min.The centrifugal 10min of extract 3000r/min gets supernatant and measures wherein protein content, and the total recovery rate that calculates albumen and polysaccharide is 86.2%.
Check experiment: check experiment comprise no ultrasonic routine extract and do not have a frequency sweep decide the ultrasonic assisted extraction of frequency.It is same as described above that no ultrasonic routine is extracted other parameter; The ultrasonic assisted extraction of frequency of deciding of no frequency sweep is meant that with the frequency sweep fabric width is whole be ± 0kHz that other parameter is same as described above.Conventional extraction and the fixed albumen of ultrasonic assisted extraction frequently and total recovery rate of polysaccharide are respectively 62.1% and 71.3%.Sweep-frequency B multifrequency pulse ultrasonic Extraction has increased by 38.8% and 20.9% respectively than conventional extraction and the fixed protein extracting ratio of ultrasonic assisted extraction frequently.
Embodiment 4
The application of the present invention in the preparation of wheat germ Angiotensin-Converting (ACE) peptide for inhibiting enzyme process---to the preliminary treatment of raw material.Add the ultrasonic Treatment groove after taking by weighing wheat germ protein 300g and 30L water mixing.At the bottom of treatment trough, put into frequency and be two of the transducers of 68kHz, be close to two of transducers placing 24kHz above the liquid level.Input power through every transducer of computer settings be 600W (4 general power 2400W), frequency sweep cycle be 170ms, frequency sweep fabric width for the ultrasonic time 200s of ± 2kHz, impulse ultrasound, idle hours 4s, the flow 5L/min of circulating pump, feed liquid initial temperature be 35 ℃.Stop ultrasonicly behind the preliminary treatment 20min, emit feed liquid, add alkali protease according to the enzyme concentration (E/S) of 3500U/g; Regulate pH8.0, at 50 ℃ of following enzymolysis 90min, holding temperature and pH are constant in the enzymolysis process; Boil the enzyme that goes out behind the enzymolysis; The centrifugal 10min of enzymolysis liquid 3500r/min, supernatant obtains wheat germ protein range of hydrolysed peptides powder through after the freeze drying, measures its inhibition effect IC to Angiotensin-Converting (ACE)
50Value is 0.35mg/mL, the IC here
50Value is to instigate the active needed peptide concentration of 50% inhibiting rate that produces of ACE.
Check experiment: what check experiment comprised no ultrasonic pretreatment and no frequency sweep decides the frequency ultrasonic pretreatment.Fixed pattern frequently is meant that setting the frequency sweep fabric width is ± 0kHz.No ultrasonic pretreatment suppresses IC with the enzymolysis product that fixed ultrasonic pretreatment frequently obtains to ACE
50Value is respectively 0.42mg/mL and 0.39mg/mL, explains that the enzymolysis product that sweep-frequency B multifrequency pulse ultrasonic pretreatment of the present invention obtains suppresses IC to ACE
50Value has improved 20.0% and 11.4% respectively than no ultrasonic pretreatment and fixed ultrasonic pretreatment frequently.
The application of the present invention in the preparation of wheat germ ace inhibitory peptide enzyme process---to the preliminary treatment of protease.Protease A lcalase is diluted to 3000 times with the optimal pH buffer solution, is made into 0.033% enzyme solutions, places ice bath subsequent use.At the sonicated bottom land transducer that to put into two centre frequencies be 24kHz, the enzyme solutions of measuring 15L injects treatment trough.Input power through every transducer of computer settings is that to be 18ms, frequency sweep fabric width be the ultrasonic time 500s of ± 2kHz, impulse ultrasound, idle hours 35 ℃ of 5s, the flow 2L/min of circulating pump, enzyme liquid initial temperatures for 300W, frequency sweep cycle.Under frequency sweep mode and fixed pattern frequently, stop ultrasonicly respectively behind the sonicated 30min, measure the activity of protease A lcalase.Fixed pattern frequently is meant that setting the frequency sweep fabric width is ± 0kHz.The frequency sweep mode ultrasonic pretreatment make the Alcalase vigor than initial value improved 9.6%, fixed pattern ultrasonic pretreatment frequently makes the Alcalase vigor improve 7.1% than initial value.
The application of the present invention in the preparation of wheat germ ace inhibitory peptide enzyme process---under ultrasonic field, carry out enzymolysis.At the sonicated bottom land transducer that to put into two centre frequencies be 24kHz, measure concentration respectively and be 12 and inject treatment trough with 24g/L wheat germ protein aqueous solution 15L.Input power through every transducer of computer settings is that to be 18ms, frequency sweep fabric width be the ultrasonic time 500s of ± 2kHz, impulse ultrasound, idle hours 50 ℃ of 5s, the flow 2L/min of circulating pump, enzymolysis liquid initial temperatures for 600W, frequency sweep cycle.With 1mol/L NaOH enzymolysis liquid pH value is transferred to 8.0, according to enzyme concentration (E/S) the adding protease A lcalase of 3500U/g.When heating enzymolysis liquid temperature to 50 ℃, open ultrasonic generator, begin to carry out frequency sweep and the fixed ultrasonic assistance enzymolysis of two kinds of patterns frequently.Fixed pattern frequently is meant that setting the frequency sweep fabric width is ± 0kHz.Whole enzymolysis process is kept pH about 8.0 with demarcating good NaOH (0.539mol/L).Cessation reaction behind the enzymolysis 90min is boiled the enzyme that goes out, and the centrifugal 10min of enzymolysis liquid 3500r/min gets ACE inhibition activity and protein transform rate that supernatant is measured hydrolysate.
No ultrasonic field enzymolysis with traditional is contrast; When concentration of substrate is 12 when the 24g/L; The ultrasonic assistance enzymolysis of frequency sweep makes enzymolysis product ACE inhibition activity improve 41.2% and 35.6% than control group respectively, and fixed ultrasonic assistance enzymolysis frequently makes enzymolysis product ACE inhibition activity improve 24.0% and 19.6% than control group respectively; When concentration of substrate was 24g/L, the ultrasonic assistance enzymolysis of frequency sweep made the product yield of wheat germ protein improve 35.5% than control group, and fixed ultrasonic assistance enzymolysis frequently makes the product yield of wheat germ protein improve 30.1% than control group.
Claims (6)
1. the pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode is made up of ultrasonic Treatment system, ultrasonic wave generation systems and computer control system, and this three part is connected by cable successively; It is characterized in that described ultrasonic Treatment system by last ultrasonic transducer promptly go up vibration plate (1), ultrasonic transducer promptly descends vibration plate (2) down; Ultrasonic Treatment casing (3), circulating pump (4), circulation valve (7); Discharging valve (8); Water-bath heat-conductive water (5), electrothermal tube (6), transducer height adjustment screw rod (15) is formed; Wherein install electrothermal tube (6) in the ultrasonic Treatment casing (3); Following vibration plate (2) is positioned at the bottom of ultrasonic Treatment casing (3); Last vibration plate (1) and following vibration plate (2) symmetry are up and down placed, and last vibration plate (1) is fixed through transducer height adjustment screw rod (15), and carry out up and down when shaking; Distance between the upper and lower vibration plate (1,2) is adjusted through transducer height adjustment screw rod (15); Circulating pump (4) is positioned at the outside of ultrasonic Treatment casing (3), links together through conveyance conduit and circulation valve (7) inside with ultrasonic Treatment casing (3), in the lower right corner of ultrasonic Treatment casing (3) liquid outlet is set; From the upper left corner of casing (3) inlet is set, the lower right corner part of ultrasonic Treatment casing (3) is equipped with circulation valve (7) and discharging valve (8);
Wherein said vibration plate (1) and the following vibration plate (2) gone up, its centre frequency is 24kHz～68kHz, the amplitude δ that changes up and down is 0～5kHz, and δ is not 0; Carry out up and down when shaking, last vibration plate (1) is different with the centre frequency of following vibration plate (2).
2. the pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode according to claim 1; It is characterized in that described ultrasonic wave generation systems is exactly supersonic generator group (13); Supersonic generator group (13) is made up of one group of different frequency supersonic generator, and its corresponding ultrasonic transducer of the supersonic generator of each frequency is corresponding.
3. the pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode according to claim 1 is characterized in that described computer control system is made up of ultrasonic wave controller (10), temperature controller (11) and computer (12); Ultrasonic wave controller (10) and computer (12) are linked together by cable; Computer (12) is controlled each particular centre frequency supersonic generator through ultrasonic wave controller (10); Computer (12) is through the bath temperature of temperature controller (11) control ultrasonic Treatment casing (3).
4. the method for using of the pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode according to claim 1; It is characterized in that the combination of upper and lower four vibration plates realizes following working method according to the logarithm of the upper and lower vibration plate of different choice of the amount of production scale in the actual production and material handling:
(1) single-frequency frequency sweep mode: i.e. two identical following vibration plates of placement frequency at the bottom of the case, inject the material solution of certain depth, carry out the frequency sweep sonicated;
(2) multifrequency frequency sweep mode: i.e. the following vibration plate of different two or more than two of placement frequency at the bottom of the case, inject the material solution of certain depth, carry out double frequency or multifrequency frequency sweep sonicated;
(3) double frequency is to the frequency sweep mode of shaking: i.e. two identical following vibration plates of placement frequency at the bottom of the case; Inject the material solution of certain depth; Be close to vibration plate on identical two of solution surface placement frequency, last vibration plate frequency is different from vibration plate down, carries out double frequency to the frequency sweep sonicated of shaking.
5. the method for using of the pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode according to claim 4 is characterized in that wherein said alternative centre frequency a is 24kHz～68kHz, and the amplitude δ that changes up and down is 0～5kHz, and δ is not 0; When carrying out the sweep parameters setting, change of frequency amplitude δ sets arbitrarily between 0～5kHz, and δ is not 0; Frequency sweep cycle is promptly accomplished a secondary frequencies and is increased to a+ δ kHz from a-δ kHz, reduces to the variation arbitrarily between 1ms～1000ms of required time of a-δ kHz from a+ δ kHz again.
6. the method for using of the pulsed multi-frequency ultrasonic bioprocess apparatus of the pulsed multi-frequency ultrasonic bioprocess apparatus of frequency sweep mode according to claim 1 or the described frequency sweep mode of claim 4 is in the application of the ultrasonic assistance enzymolysis of the ultrasonic assisted extraction of active ingredient of natural product, large biological molecule, and the ultrasonic assistance enzymolysis of wherein said large biological molecule comprises preliminary treatment to raw material, carries out enzyme digestion reaction to the preliminary treatment of hydrolase with in ultrasound field.
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Application publication date: 20100526 Assignee: Jiangsu Ligong Fruit & Vegetable Machinery Co., Ltd. Assignor: Jiangsu University Contract record no.: 2015320000260 Denomination of invention: Device and method for biological processing with pulsed multi-frequency ultrasonic waves in sweep-frequency mode and use thereof Granted publication date: 20120829 License type: Exclusive License Record date: 20150420 |
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