CN101711969A - 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
扫频模式的脉冲多频超声波生物加工装置、方法及其应用，涉及农产品精深加工领域。由超声波处理系统、超声波发生系统和计算机控制系统组成，这三部分由线缆依次连接而成，其中所述的超声波处理系统由上超声波换能器、下超声波换能器、超声波处理槽体、循环泵和上振板高度调整螺杆组成；超声波发生器组由一组不同频率脉冲式超声波发生器组成，控制系统由超声波控制仪，温度控制仪和计算机组成；该设备可用于天然产物有效成分的超声辅助提取，提取效率提高10-90％；可用于生物大分子的超声辅助酶解，生物大分子的超声辅助酶解包括对原料的预处理、对水解酶的预处理和在超声波场中进行酶解反应，酶解效率提高25-65％。
The pulsed multi-frequency ultrasonic bioprocessing device, method and application thereof in frequency sweep mode relate to the field of intensive processing of agricultural products. It consists of an ultrasonic processing system, an ultrasonic generating system and a computer control system. These three parts are connected in sequence by cables. The ultrasonic processing system consists of an upper ultrasonic transducer, a lower ultrasonic transducer, an ultrasonic treatment tank, The circulating pump and the height adjustment screw of the upper vibration plate are composed; the ultrasonic generator group is composed of a group of pulsed ultrasonic generators with different frequencies, and the control system is composed of an ultrasonic controller, a temperature controller and a computer; this equipment can be used for the active ingredients of natural products Ultrasonic-assisted extraction, the extraction efficiency is increased by 10-90%; it can be used for ultrasonic-assisted enzymolysis of biomacromolecules. Enzymolysis reaction, enzymolysis efficiency increased by 25-65%.
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 the medium that chemical reaction is used always, produce a series of extreme conditions that approach, 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.Falling field such as poly-at extraction separation, the preparation of biologically active peptide, the polysaccharide of active ingredient of natural product has in recent years obtained using comparatively widely.
A large amount of studies 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 is subjected to 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 using method 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 be subjected to 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), here said " 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 gap 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 complexity, 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 by centre frequency a to the effect system, to 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 the difference according to the ultrasonication object at work, seek best frequency sweep cycle, purpose is to make system realize maximum sonic oscillation with the minimal power input.The present invention realizes by following dual mode the change of frequency sweep cycle: scanning constant speed changes the size of frequency change 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 (as 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.
Dynamic tracking, monitoring, adjustment and the control of major parameter 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 wave treatment system, ultrasonic wave generation systems and computer control system, this three part is connected in sequence by cable.
Described ultrasonic wave treatment system is promptly gone up vibration plate, is descended ultrasonic transducer promptly to descend vibration plate by last ultrasonic transducer, and ultrasonic wave is handled casing, circulating pump, and circulation valve, the discharging valve, the water-bath heat-conductive water, electrothermal tube, transducer Height Adjustment screw rod is formed.Wherein ultrasonic wave is handled in the casing and is installed electrothermal tube, can satisfy ultrasonic treatment fluid different temperatures and set; Following vibration plate is positioned at the bottom that ultrasonic wave is handled casing, last vibration plate and following vibration plate symmetry are up and down placed, last vibration plate is fixed by transducer Height Adjustment screw rod, and carry out up and down when shaking, distance between the vibration plate is adjusted by transducer Height Adjustment screw rod up and down, circulating pump is positioned at the outside that ultrasonic wave is handled casing, inside by conveyance conduit and valve and ultrasonic wave processing casing links together, the lower right corner of handling casing at ultrasonic wave is provided with liquid outlet, from the upper left corner of casing inlet is set, the lower right corner part that ultrasonic wave is handled 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 of Bian Huaing is 0～5kHz up and down; 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 that ultrasonic wave is handled casing, handling cell body inside by conveyance conduit and valve and ultrasonic wave links together, the lower right corner of handling casing at ultrasonic wave is provided with liquid outlet, from the upper left corner of casing inlet is set, from the lower right corner fluid of ultrasonic wave processing cell body, return from the upper left corner of cell body.The lower right corner part that ultrasonic wave is handled 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 circulating of material to be processed solution, strengthens convective mass transfer, improves ultrasonic treatment 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, the 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, frequency change amplitude (δ) can be set arbitrarily between 0～2kHz; Frequency sweep cycle (promptly finish 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; Computer is controlled the ultrasonic time of power, frequency sweep cycle, frequency sweep fabric width (± δ kHz), impulse ultrasound of each particular centre frequency (a kHz) supersonic generator and intermittent time, whole ultrasonic processing time by the ultrasonic wave controller.Computer is handled the bath temperature of cell body by temperature controller control ultrasonic wave.Can set above-mentioned all parameters by 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 ultrasonic processing of frequency sweep;
(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 the ultrasonic processing of multifrequency frequency sweep;
(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 ultrasonic processing of the frequency sweep of shaking.
Wherein said alternative centre frequency (a) is 24kHz～68kHz, and the amplitude of Bian Huaing (δ) is 0～5kHz up and down; When carrying out the sweep parameters setting, frequency change amplitude (δ) can be set arbitrarily between 0～5kHz; Frequency sweep cycle is promptly finished 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 by 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 as follows: put into vibration plate 2 down in the treatment trough bottom, inject the material solution of mixing behind certain depth to cell body 3; 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, adjust the distance that goes up between vibration plate 1 and the following vibration plate 2 by last vibration plate Height Adjustment screw rod 15; The ultrasonic time of the power by 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 by water-bath; After being heated to the operating temperature of setting, opening circulation valve 7 and ON cycle pump 4 and carry out the liquid circulation, open supersonic generator and begin to carry out ultrasonic processing.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), the 3-ultrasonic wave is handled casing, 4-circulating pump, 5-water-bath heat-conductive water, the 6-electrothermal tube, the 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 embodiments and the drawings 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 wave treatment system, ultrasonic wave generation systems and computer control system, this three part is connected in sequence by cable.Ultrasonic wave 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 by last ultrasonic transducer, and ultrasonic wave is handled 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 ultrasonic wave is handled installing electrothermal tube 6 in the casing 3, can satisfy ultrasonic treatment fluid different temperatures and set; Following vibration plate 2 is positioned at the bottom that ultrasonic wave is handled casing 3, last vibration plate and following vibration plate symmetry are up and down placed, last vibration plate 1 is fixed by transducer Height Adjustment screw rod 15, when carrying out up and down when shaking, distance between the vibration plate can be adjusted by 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 of Bian Huaing (δ) is 0～5kHz up and down; 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 ultrasonic processing of frequency sweep;
(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 the ultrasonic processing of multifrequency frequency sweep;
(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 ultrasonic processing of the frequency sweep of shaking.
Wherein said alternative centre frequency (a) is 24kHz～68kHz, and the amplitude of Bian Huaing (δ) is 0～5kHz up and down; When carrying out the sweep parameters setting, frequency change amplitude (δ) can be set arbitrarily between 0～5kHz; Frequency sweep cycle is promptly finished 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 that ultrasonic wave is handled casing 3, handles cell body inside by conveyance conduit and valve with ultrasonic wave and links together, and from the lower right corner fluid of ultrasonic wave processing cell body 3, returns from the upper left corner of cell body 3.The lower right corner part that ultrasonic wave is handled 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 circulating of material to be processed solution, strengthens convective mass transfer, improves ultrasonic treatment 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, the 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, frequency change amplitude (δ) can be set arbitrarily between 0～2kHz; Frequency sweep cycle (promptly finish 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 and computer 12 are formed.Link together by cable; The ultrasonic time of power by 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 handled the bath temperature of cell body 3 by controller 14 control ultrasonic waves.Can set by 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.Be crushed to the dried laver of 1kg (containing protein 41 .3%) behind 60 orders and 40L water mixing after add the ultrasonic wave treatment trough.The transducer that to put into two frequencies at the bottom of treatment trough be 28kHz is being close to the transducer (at a distance of about 20cm) of placing two 20kHz above the liquid level.Input power by 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%.The ultrasonic extraction of sweep-frequency B multifrequency pulse 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.Be crushed to 1.8kg rapeseed cake dregs crude protein (containing albumen 48%) behind 90 orders and 30L water mixing after add the ultrasonic wave treatment trough, adjust pH to 9.8.The transducer that to put into two frequencies at the bottom of treatment trough be 28kHz is being close to the transducer (at a distance of about 15cm) of placing two 24kHz above the liquid level.Input power by 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%.The ultrasonic extraction of sweep-frequency B multifrequency pulse has increased by 89.1% and 22.2% respectively than conventional extraction and the fixed protein extracting ratio of ultrasonic assisted extraction frequently.
Embodiment 3
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.The ultrasonic wave treatment trough will be added behind 200g defatted rice bran and the 20L water mixing.The transducer that to put into two frequencies at the bottom of treatment trough be 24kHz and 64kHz.Input power by 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%.The ultrasonic extraction of sweep-frequency B multifrequency pulse 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 wave treatment trough 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 by 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 ultrasonic behind the preliminary treatment 20min, emit feed liquid, enzyme concentration (E/S) according to 3500U/g adds alkali protease, 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 preliminary treatment and no frequency sweep decides the ultrasonic preliminary treatment of frequency.Fixed pattern frequently is meant that setting the frequency sweep fabric width is ± 0kHz.Do not have ultrasonic preliminary treatment and ACE is suppressed IC with the enzymolysis product that fixed ultrasonic preliminary treatment frequently obtains
50Value is respectively 0.42mg/mL and 0.39mg/mL, illustrates that the enzymolysis product that the ultrasonic preliminary treatment of sweep-frequency B multifrequency pulse of the present invention obtains suppresses IC to ACE
50Value has improved 20.0% and 11.4% respectively than no ultrasonic preliminary treatment and fixed ultrasonic preliminary treatment frequently.
Embodiment 5
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 standby.The transducer that to put into two centre frequencies at the bottom of ultrasonic treatment trough be 24kHz, the enzyme solutions of measuring 15L injects treatment trough.Input power by 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.Behind ultrasonic processings 30min under frequency sweep mode and the fixed pattern frequently, stop respectively ultrasonic, the activity of mensuration protease A lcalase.Fixed pattern frequently is meant that setting the frequency sweep fabric width is ± 0kHz.The ultrasonic preliminary treatment of frequency sweep mode make the Alcalase vigor than initial value improved 9.6%, the fixed ultrasonic preliminary treatment of pattern frequently makes the Alcalase vigor improve 7.1% than initial value.
Embodiment 6
The application of the present invention in the preparation of wheat germ ace inhibitory peptide enzyme process---under ultrasonic field, carry out enzymolysis.The transducer that to put into two centre frequencies at the bottom of ultrasonic treatment trough be 24kHz, measure concentration respectively and be 12 and 24g/L wheat germ protein aqueous solution 15L inject treatment trough.Input power by 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.
With traditional no ultrasonic field enzymolysis is contrast, when concentration of substrate is 12 and during 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 (7)
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