阅读说明：本技术 一种脉冲电场强化超声辅助提取红曲色素工艺方法 (A kind of impulse electric field reinforcing ultrasound assisted extraction monascorubin process ) 是由 何毅 刘志伟 李明 杨宁 何丽丽 于 2019-09-06 设计创作，主要内容包括：本文发明采用脉冲电场强化超声辅助法对液态发酵的红曲霉菌体中的红曲色素进行提取,并提供一种有效提取红曲霉色素提供最佳工艺条件,其中乙醇与菌丝体的料液比为3：1(g/L),脉冲电场的电场强度为500 V/cm,脉冲温度为60℃下脉冲3次处理,然后在超声功率为390 W,超声温度为60℃,超声时间为30 min的条件下处理,测得红曲色素的黄色素色价为439.8 U/g,橙色素色价为229.3 U/g,红色素色价为350.4 U/g,总色价为519.5 U/g。与传统的乙醇溶液浸提法相比,脉冲强化超声辅助提取红曲色素的效果提高了85.2%。(Invention herein is strengthened ultrasonic wave added method using impulse electric field and is extracted to the monascorubin in the monascus thallus of liquid state fermentation, and provide a kind of effectively extraction Monascouruarin offer optimum process condition, wherein ethyl alcohol and mycelial solid-liquid ratio are 3:1(g/L), the electric field strength of impulse electric field is 500 V/cm, pulse temperature is pulse 3 times processing at 60 DEG C, it then is 390 W in ultrasonic power, ultrasonic temperature is 60 DEG C, ultrasonic time is handled under conditions of being 30 min, the uranidin color value for measuring monascorubin is 439.8 U/g, citraurin color value is 229.3 U/g, haematochrome color value is 350.4 U/g, total color value is 519.5 U/g.Compared with traditional ethanol solution extraction, the effect that ultrasound assisted extraction monascorubin is strengthened in pulse improves 85.2%.)

1. a kind of impulse electric field strengthens ultrasound assisted extraction monascorubin process, it is characterized in that, strengthened using impulse electric field Ultrasonic wave added method extracts the monascorubin in the monascus thallus of liquid state fermentation.

2. a kind of impulse electric field according to claim 1 strengthens ultrasound assisted extraction monascorubin process, feature For ethyl alcohol and mycelial solid-liquid ratio are 1-5:1-5(g/L in extraction process), the electric field strength of impulse electric field is 300-700 V/cm, pulse temperature are pulse 1-5 times processing at 40-80 DEG C, are then 250-500 W in ultrasonic power, ultrasonic temperature is 40-80 DEG C, ultrasonic time is handled under conditions of being 20-40 min.

3. a kind of impulse electric field according to claim 2 strengthens ultrasound assisted extraction monascorubin process, feature For ethyl alcohol and mycelial solid-liquid ratio are 3:1(g/L), the electric field strength of impulse electric field is 500 V/cm, pulse temperature 60 Then pulse 3 times processing at DEG C are 390 W in ultrasonic power, ultrasonic temperature is 60 DEG C, and ultrasonic time is the condition of 30 min Lower processing.

Technical field

The present invention relates to a kind of method for extracting pigment, and in particular to a kind of impulse electric field reinforcing ultrasound assisted extraction red yeast rice color Plain process.

Background technique

Monascus ruber is a kind of filamentous fungi, and mycelia has tabula and multicore, and there are many thallus branch, in its growth course In, color can become aubergine from colourless.

The people in China begin to produce red yeast rice using monascus ruber before thousands of years.Monascouruarin can both work as Food can also work as drug, and Monascouruarin can substitute some traditional food additives, be similar to nitrite or rouge Rouge is red etc.Before for a long time, when soy sauce is studied not yet, red yeast rice is then used to make pork braised in brown sauce etc by ancients Color is red food.Monascouruarin is a kind of natural pigment produced by microbial fermentation, is synthesized with industrial chemistry Pigment is compared, it is not limited by stringent safety, has extensive be applicable in^]。

Monascus ruber can provide very big edible or medical value to the mankind.Scientists have been found that until today About 200 kinds of Monascus Strains, wherein there is the bacterial strain of a quarter that can be used to ferment.Because what monascus ruber generated Pigment be it is pure natural and edible, people are subconscious to think that it is more safer than chemical staining agent, so more people Such a natural pigment can be selected among food processing.Since the economic value of monascus ruber increases constantly, such as What can develop and use it also into the scientific research project of domestic and international many researchers to greatest extent.

Monascorubin is that monascus ruber obtains after fermentation after a period of time as a kind of natural colouring substance, it It is the hybrid pigment of red color tone and yellow hue, nutritive value and bioactivity are all very high, and are edible natural pigments, science Family gradually study monascorubin and can replace Nitrates in meat products.Monascorubin belongs to polyketides, according to Understand, the mankind understand and analyze about 90 kinds of monascus pigment component of structure, and monascorubin main component is Monascus color Pigment, red yeast rice citraurin, monascus yellow pigment etc..Monascorubin meets safety, nutrition, multi-functional hair in international food pigment method Open up direction, and be widely used range and long-range development prospect.

Solvent extraction method, this method long, solvent acquisition amount there are the time are mainly used to the extraction of monascorubin at present Greatly, the very low disadvantage of recovery rate, and product is red yeast rice hybrid pigment.Most of solvent extraction method is to study how to improve pigment Overall recovery rate and color value, if the extraction condition used is different, the extraction effect of different component also can in monascorubin As a result difference causes the monascus pigment component extracted to have very big difference.Therefore, in order to reach the quality mark of monascorubin product The method of new extraction monascorubin is further studied and explored to standard, the monascorubin of separation and exploitation heterogeneity and tone It is necessary.

Summary of the invention

Goal of the invention: invention herein strengthens ultrasonic wave added method in the monascus thallus of liquid state fermentation using impulse electric field Monascorubin extracts, and has studied the influence for the monascorubin that impulse electric field is strengthened in ultrasound assisted extraction monascus thallus Factor and Extraction technique provide optimum process condition effectively to extract Monascouruarin.Mainly a kind of pulse is electric by the present invention Field strength ultrasound assisted extraction monascorubin process mainly strengthens ultrasonic wave added method to liquid state fermentation using impulse electric field Monascorubin in monascus thallus extracts.Wherein ethyl alcohol and mycelial solid-liquid ratio are 1-5:1-5(g/L), pulse electricity The electric field strength of field is 300-700 V/cm, and pulse temperature is pulse 1-5 times processing at 40-80 DEG C, is then in ultrasonic power 250-500 W, ultrasonic temperature are 40-80 DEG C, and ultrasonic time is handled under conditions of being 20-40 min.Preferably, ethyl alcohol and bacterium The solid-liquid ratio of filament is 3:1(g/L), the electric field strength of impulse electric field is 500 V/cm, and pulse temperature is pulse 3 times at 60 DEG C Then processing is 390 W in ultrasonic power, ultrasonic temperature is 60 DEG C, and ultrasonic time is handled under conditions of being 30 min.This hair It is bright the utility model has the advantages that being 3:1(g/L in ethyl alcohol and mycelial solid-liquid ratio), the electric field strength of impulse electric field is 500 V/cm, pulse Temperature is pulse 3 times processing at 60 DEG C, is then 390 W in ultrasonic power, ultrasonic temperature is 60 DEG C, ultrasonic time 30 It is handled under conditions of min, the uranidin color value for measuring monascorubin is 439.8 U/g, and citraurin color value is 229.3 U/g, red Pigment color value is 350.4 U/g, and total color value is 519.5 U/g.Compared with traditional ethanol solution extraction, ultrasound is strengthened in pulse The effect of assisted extraction monascorubin improves 85.2%.

Figure of description

Influence of Fig. 1 solid-liquid ratio to monascorubin extraction effect.Fig. 2 pulse pre-treatment temperature is to monascorubin extraction effect Influence.Influence of Fig. 3 pulse number to monascorubin extraction effect.

Influence of Fig. 4 electric field strength to monascorubin extraction effect.Fig. 5 ultrasonic power, which extracts monascorubin, imitates The influence of fruit.Influence of Fig. 6 ultrasonic time to monascorubin extraction effect.Fig. 7 ultrasonic temperature extracts monascorubin The influence of effect.

Specific embodiment

Select the Monascus ruber mould separated in rice.

Agents useful for same

1 agents useful for same list of table

Reagent	Purity grade	Production unit
			DEXTROSE ANHYDROUS	Analyze pure AR	Sinopharm Chemical Reagent Co., Ltd.
Beef extract	Biochemical reagents BR	Sinopharm Chemical Reagent Co., Ltd.
			Pancreas peptone soybean broth	Analyze pure AR	OXOID company, Britain
Yeast extract	Biochemical reagents BR	Beijing extensive and profound in meaning star biotechnology Co., Ltd
			Sodium nitrate	Analyze pure AR	Sinopharm Chemical Reagent Co., Ltd.
Magnesium sulfate	Analyze pure AR	Sinopharm Chemical Reagent Co., Ltd.
			Agar Agar	Analyze pure AR	German BIOFROXX company
Dehydrated alcohol	Analyze pure AR	Sinopharm Chemical Reagent Co., Ltd.

Instrument is shown in Table 2 table, 2 instrument list

Laboratory apparatus title	Production unit
		High-temp steam sterilizing pot GI36DWS	ZEALWAY INSTRUMENT INC.
Electronic analytical balance AL204	Mei Teletuo multiple instruments Co., Ltd
		Centrifuge S718R	German OHAUS company
Aseptic operating platform DL-CJ-1NDII	Beijing Dong Lianhaer instrument manufacturing Co., Ltd
		Constant-temperature shaking incubator HZ300L	Wuhan Rui Hua instrument and equipment Co., Ltd
Mold incubator MJP400	Wuhan Rui Hua instrument and equipment Co., Ltd
		Sweep type ultraviolet specrophotometer Evolution 220	Power & light company, the U.S.
II N of ultrasonic extractor JY92-	NingBo XinZhi Biology Science Co., Ltd
		High-pressure pulse electric generator THU-PEF4	Wuhan Xin Tianpu laboratory equipment Co., Ltd
Biomicroscope CX40	Ningbo ShunYu Instruments Co., Ltd
		Thermostat water bath HH-2	Guo Hua Electrical Appliances Co., Ltd
Laboratory pH meter ST2100	German OHAUS company
		Electric heating constant-temperature blowing drying box DHG-9243BS- III	Shanghai new talent medical instrument Manufacturing Co., Ltd

The preparation of culture medium

The preparation of dextrose solid medium is shown in Table 3

3 dextrose solid medium of table

Component	Quality	Component	Quality
				Glucose	40.0 g	Yeast extract	5.0g(summer 3.0g)
Beef extract	3.0 g	Magnesium sulfate	1.0 g
				Peptone	8.0 g	Sodium nitrate	2.0 g
Agar	20.0 g	Distilled water	1000 mL

Mentioned reagent is added in the beaker of 1000 mL, heating is dissolved and stirred evenly.The usage amount when culture medium is uncolled The solid medium that cylinder measures 200 mL is transferred in the vial with blue cap.Height is put into after the lid of vial is unscrewed Warm steam sterilization pan, it is 121 DEG C that temperature, which is arranged, in autoclave, and setting time is 20 min.It is taken out after having sterilized, by vial Indigo plant lid is tightened, and is stored after cooling spare.

The preparation of dextrose broth is shown in Table 4

4 dextrose broth of table

Component	Quality	Component	Quality
				Glucose	40.0 g	Yeast extract	5.0g(summer 3.0g)
Beef extract	3.0 g	Magnesium sulfate	1.0 g
				Peptone	8.0 g	Sodium nitrate	2.0 g

Mentioned reagent is added in the beaker of 1000 mL, the distilled water of 1000 mL is added, heating makes reagent on Muffle furnace It all dissolves and is stirred continuously using glass bar, trained when culture medium is uncolled using the liquid of glucose that graduated cylinder measures 200 ml Group-transfer is supported into the conical flask of 250 mL, bottleneck is stoppered using rubber stopper and wraps bottleneck with brown paper.It is equipped with all The conical flask of fluid nutrient medium is put into high-temp steam sterilizing pot, and it is 121 DEG C that temperature, which is arranged, in autoclave, and setting time is 20 min. Conical flask is taken out after having sterilized, being cooled to after 40 DEG C can be used.

The preparation of slant medium: above-mentioned sterilized dextrose solid medium is heated into 5-6 in micro-wave oven Min, it is therefore an objective to the culture medium for being cooled to solid be made to be melted into liquid.Aseptic operating platform shifts to an earlier date 30 min and opens ultraviolet-sterilization Lamp is put into aseptic operating platform from culture medium is taken out in micro-wave oven, when the temperature of culture medium being waited to drop to 40 DEG C -50 DEG C, The production of test tube slant culture medium is completed in aseptic operating platform.It needs to be put into before hand is put in aseptic operating platform sterile Thing and both hands in station spray alcohol, light alcolhol burner, sterilized empty test tube are taken out, by rubber stopper and test tube Mouth is placed on calcination 5 seconds on the flame of alcolhol burner, and the dextrose solid medium of about 5 cm is then poured into test tube, is continued rubber Rubber plug and test tube mouth are placed on the flame of alcolhol burner stoppered test tube after calcination 5 seconds.Test tube is placed on and is laid flat in aseptic operating platform On a piece glass bar, slightly it is tilted a certain angle test tube and leans against on glass bar, when the culture medium in test tubes being waited to be cooled to solid-state The production of test tube slant culture medium just completes.Operation above step needs to carry out in alcolhol burner flame periphery, the reason is that The temperature of alcolhol burner flame periphery is higher, it is believed that is a sterile environment, can guarantee the solid medium of production not It can be contaminated.Cooling test tube slant culture medium is wrapped with brown paper, is placed in 4 DEG C of refrigerator.It is needed after the completion of experiment whole Aseptic operating platform is managed, laboratory is cleaned up.

The inoculated and cultured of Monascus: the inoculated and cultured of Monascus is to need to be operated in aseptic operating platform, sterile behaviour As platform need in advance 30 min open ultraviolet lamp sterilize.By the calcination on alcolhol burner flame of the transfer needle in aseptic operating platform 1 min, it is therefore an objective to kill the bacterium on transfer needle, prevent Monascus contaminated.Furthermore the rod of transfer needle is also required in flame Upper calcination 2-3 times, waiting the syringe needle of transfer needles to burn rubicundity can be used.The test tube slant for having culture medium is taken first And one cultivated 14 days or so Monascus test tube slants, its test tube mouth and rubber stopper are all placed on alcolhol burner flame Several seconds of upper calcination, then protrudes into transfer needle on the test tube wall of Monascus, waits the temperature of transfer needle to decline, it is therefore an objective to keep away The temperature for exempting from transfer needle excessively high makes Monascus lose activity.One piece of Portugal for having Monascus is marked from test tube slant using transfer needle Grape sugar solid medium, is then transferred on the test tube slant of new dextrose solid medium.That face with Monascus It is in contact, is scratched red yeast rice fritter to upper end from test tube slant lower end using transfer needle, then shift test tube bottom into slant medium End operates the 2-3 rear red yeast rice culture medium for taking out fritter back and forth.By test tube mouth and rubber stopper on flame in several seconds rear plugs of calcination Rubber stopper.Often connect a test tube will calcination transfer needle and transfer needle again rod, after the rubber stopper calcination of test tube is complete Handstand is placed on beside alcolhol burner.It is inoculated with carrying out for need of work 3-4 days once for red yeast rice, it is sterilized to be inoculated with 15-20 branch every time Dextrose solid medium test tube slant.The Monascus slant tube being inoculated with need to be put into 28 DEG C of constant incubator into Row culture.

The collection of Monascus spore: the red yeast rice examination that 18-20 branch cultivated at least two weeks is taken out in constant incubator Pipe inclined-plane determines that the monascus ruber of every test tube grows fine.Aseptic operating platform needs 30 min opening ultraviolet lamp progress in advance 8 mL sterile waters are added to red yeast rice test tube slant using the pipette tips of 1 mL in aseptic operating platform in sterilizing, will be in aseptic operating platform Loop-carrier on alcolhol burner flame 1 min of calcination back and forth, it is therefore an objective to kill the bacterium on loop-carrier, prevent Monascus dirty Dye.Furthermore it is i.e. usable to be also required on flame calcination 2-3 time for inoculation rod.Loop-carrier after calcination is protruded into test tube, In The temperature decline of stick to be seeded such as on test tube wall, it is therefore an objective to avoid the temperature of loop-carrier is excessively high Monascus is made to lose activity.Then It puts forth one's strength and scratches back and forth on the inclined-plane for have Monascus slightly, it is therefore an objective to by the spore of red yeast rice mycelia disengaging culture medium as much as possible It is suspended in sterile water.Test tube is laid flat slightly, the spore liquid of test tube mouth is drawn using the pipette tips of 1 mL, is injected into height The sterilized funnel with sterile lens wiping paper is filtered operation in warm steam sterilization pan, is connected below funnel equipped with about 10 In the conical flask of grain bead.After having filtered by conical flask and plug on flame several seconds of calcination, plug is stoppered into conical flask simultaneously It is wrapped with brown paper.Conical flask is placed in constant-temperature shaking incubator, and temperature is 28 DEG C, and shaking speed is 180 r/min, when oscillation Between be 20 min.The purpose of shaking table is to open the cleistothecium of red yeast rice spore, to obtain more spores.Wait the shaking table time After complete, take out conical flask, spore suspension is dispensed into the compact centrifuge for being put into 8000 r/min in the centrifuge tube of 2 mL from 15 min of the heart.The supernatant in centrifuge tube is outwelled after being centrifuged, remaining spore liquid in centrifuge tube is sucked out with pipette tips, is collected into In one centrifuge tube, spore liquid is concentrated in this.

The concentration spore liquid that about 100 μ L are taken out from centrifuge tube uses blood cell plate meter after being diluted multiple appropriate Number calculates spore count according to formula, determines the spore liquid volume being injected into fluid nutrient medium.

It is directly counted under the microscope using blood counting chamber, is a kind of common microbial enumeration method.The method tool Have the advantages that direct and quick.Blood counting chamber glass slide and lid will be placed in by appropriate diluted bacteria suspension (or spore suspension) In counting chamber between slide, counted under the microscope.The blood counting chamber in laboratory is 25 × 16 types, blood count The counting chamber of plate is 5 × 5 grid.Blood counting chamber is cleaned with sterile water and is dried with lens wiping paper, opens 20 mm with rear cover one The coverslip of × 20 mm, the dilution spore liquid for selecting suitable liquid-transfering gun to draw 20 μ L or so are slowly squeezed under coverslip, are covered There cannot be bubble otherwise to influence to observe in slide.80 sub-boxes are remembered when counting altogether.The spore liquid of same concentrations needs to count Number 3 times, the average value of spore count under the concentration is calculated, calculates red yeast rice spore contained in every 1 mL concentration spore liquid as follows Sub- number.

Wherein c: every milliliter spore count

The endosporous number of the small lattice of n:80

F: extension rate

According to calculating so that the spore liquid inoculum concentration in each conical flask is identical and should beA/ml, since spore can be concentrated It is slowly sunk in spore liquid, will suitably rock conical flask when so drawing spore liquid with pipette tips every time, make as far as possible each Dextrose broth miospore concentration is identical.The conical flask being inoculated with is wrapped with brown paper, is put into 28 DEG C of perseverance In warm shaking table, the revolving speed of shaking table is 160 r/ min, is cultivated 10-14 days.

The collection of fermentation liquid: to fermentation ends, conical flask is taken out in constant-temperature shaking incubator, the mycelium in conical flask It is put into the funnel with filter paper and filters, rinsed 2-3 times with the wash bottle equipped with distilled water, gently squeeze mycelial water with hand Point.Mycelium is placed in glass dish, is placed in 70 DEG C of electric heating constant-temperature blowing drying boxes, the time for drying moisture is not at least low In 48 h.Glass dish is taken out, is ground using mortar, then leads to and sieves with 100 mesh sieve to get to dry Monascus anka Nakazawa et sato filament powder, be collected into It is spare in centrifuge tube.

Monascorubin determination of yield: para chrome (pigment in mycelium) measuring method: 0.3000 g Monascus anka Nakazawa et sato filament is taken 10 mL, 70% ethyl alcohol is added in powder, and 1 h of water-bath at 60 DEG C takes filtrate after filtering, after diluting multiple appropriate with 70% ethyl alcohol, Its light absorption value is scanned from 300 nm to 600 nm with ultraviolet-visible spectrophotometer, while using 70% ethyl alcohol as negative control. Monascus Strains, which are calculated separately, divided by the concentration of sample with the light absorption value measured under 390 nm, 470 nm and 520 nm produces born of the same parents The yield of interior uranidin, citraurin and haematochrome, unit are U/g.

Test one: influence of the impulse electric field conditions to monascorubin extraction effect

Influence of the solid-liquid ratio to monascorubin extraction effect: respectively with 1: 1,2: 1,3: 1,4: 1,5: 1(g/L) solid-liquid ratio it is molten In five part of 100 mL, 70% ethyl alcohol, 1 h of water-bath at 60 DEG C, is 500 V/cm, flow velocity 1.5 in the electric field strength of impulse electric field ML/s is handled sample under conditions of pulse 3 times.The sample handled is fitted into the centrifuge tube of 10 mL and is put into centrifuge Middle centrifugation.The revolving speed of centrifuge is set as 5 000 r/min, and centrifugation time is 20 min.Take the supernatant that has been centrifuged with 70% After ethyl alcohol dilutes multiple appropriate, its light absorption value is scanned from 300 nm to 600 nm with sweep type ultraviolet specrophotometer, simultaneously Use 70% ethyl alcohol as negative control.With the light absorption value measured under 390nm, 470 nm and 520 nm divided by the concentration of sample Calculate separately the yield that Monascus Strains produce uranidin intracellular, citraurin and haematochrome, unit is U/g.

Influence of the pre-treatment temperature to monascorubin extraction effect: it accurately weighs the dry thallus powder of five part of 0.3000 g and is dissolved in 100 mL, 70% ethanol solution, 1 h of water-bath at being respectively 30,40,50,60,70 DEG C in temperature, in the electric-field strength of impulse electric field Degree is 500 V/cm, and flow velocity is 1.5 mL/s, is handled under conditions of pulse 3 times sample.The sample handled uses 2.2.3.1 method is post-processed.With the light absorption value measured under 390 nm, 470 nm and 520 nm divided by the dense of sample It spends to calculate separately the yield that Monascus Strains produce uranidin intracellular, citraurin and haematochrome, unit is U/g.

Influence of the pulse number to monascorubin extraction effect: it accurately weighs the dry thallus powder of five part of 0.3000 g and is dissolved in 100 mL, 70% ethanol solution, temperature are 1 h of water-bath at 60 DEG C, are 500 V/cm in the electric field strength of impulse electric field, flow velocity is To sample difference pulse 1 time, 2 times, 3 times, 4 times, 5 processing under conditions of 1.5 mL/s.The sample handled uses 2.2.3.1 Method post-processed.Divided with the light absorption value measured under 390 nm, 470 nm and 520 nm divided by the concentration of sample Not Ji Suan Monascus Strains produce the yield of uranidin intracellular, citraurin and haematochrome, unit is U/g.

Influence of the electric field strength to monascorubin extraction effect: it accurately weighs the dry thallus powder of five part of 0.3000 g and is dissolved in 100 mL, 70% ethanol solution, temperature be 60 DEG C at 1 h of water-bath, respectively with the electric field strength of impulse electric field be 200,300, 400,500,600 V/cm, flow velocity are 1.5 mL/s, are handled under conditions of pulse 3 times sample.The sample handled makes It is post-processed with the method for 2.2.3.1.With the light absorption value measured under 390 nm, 470 nm and 520 nm divided by sample Concentration calculates separately the yield that Monascus Strains produce uranidin intracellular, citraurin and haematochrome, and unit is U/g.

Impulse electric field strengthens influence of the ultrasonic wave added to monascorubin extraction effect: it is dry to accurately weigh five part of 0.3000 g Dry thallus powder is dissolved in 100 mL, 70% ethanol solution, and temperature is 1 h of water-bath at 60 DEG C, is 500 in the electric field strength of impulse electric field V/cm, flow velocity rush 3 processing to sample sectors under conditions of being 1.5 mL/s.

Influence of the ultrasonic power to monascorubin extraction effect: will be by impulse electric field treated sample respectively super Acoustic power is 130,195,260,325,390 W, extracts 30 min at 60 DEG C of temperature.The sample handled is packed into 10 mL's Centrifugation is put into a centrifuge in centrifuge tube.The revolving speed of centrifuge is set as 5 000 r/min, and centrifugation time is 20 min.Take from After the complete supernatant of the heart dilutes multiple appropriate with 70% ethyl alcohol, with sweep type ultraviolet specrophotometer from 300 nm to 600 Nm scans its light absorption value, while using 70% ethyl alcohol as negative control.With the extinction measured under 390 nm, 470 nm and 520 nm Value calculates separately the yield that Monascus Strains produce uranidin intracellular, citraurin and haematochrome, unit divided by the concentration of sample It is U/g.

Influence of the ultrasonic time to monascorubin extraction effect: will be by impulse electric field treated sample in ultrasonic wave function Rate is 390 W, is handled respectively with the sonication times of 10,20,30,40,50,60,70 min at 60 DEG C of temperature.It has handled Sample is post-processed using the method for 2.2.4.1.With the light absorption value measured under 390 nm, 470 nm and 520 nm divided by tested The concentration of sample calculates separately the yield that Monascus Strains produce uranidin intracellular, citraurin and haematochrome, and unit is U/g.

Influence of the ultrasonic temperature to monascorubin extraction effect: will be by impulse electric field treated sample respectively with temperature Degree is 30 DEG C, and 40 DEG C, 50 DEG C, 60 DEG C, 70 DEG C of ultrasonic temperature, ultrasonic power is to extract 30 min under 390 W.Place The sample managed is post-processed using the method for 2.2.4.1.It is removed with the light absorption value measured under 390 nm, 470 nm and 520 nm The yield that Monascus Strains produce uranidin intracellular, citraurin and haematochrome is calculated separately with the concentration of sample, and unit is U/ g.Experiment two: the single factor experiment of impulse electric field extraction monascorubin

The determination of liquid-to-solid ratio: as shown in Figure 1, since solid-liquid ratio is 5:1, since ethanol solution usage amount is less, ethyl alcohol with it is red Aspergillus filament, which cannot mix well, causes monascorubin extraction efficiency low, with the increase of ethanol consumption, red yeast rice color in extracting solution Total color value of element gradually increases, when solid-liquid ratio reaches 3:1, Monascus color in monascorubin, and red yeast rice orange, the yellow three kinds of components of red yeast rice Color value value reaches peak, and total color value also reaches maximum value.When ethanol consumption further increases, the color value of monascorubin is instead It decreases, this may be since during using impulse electric field, ethanol consumption is excessive and red yeast rice thallus does not dissolve in Ethanol solution, so ethyl alcohol is first inhaled into impulse electric field processor, the thallus to precipitate is not uniformly mixed with ethyl alcohol, So the extraction efficiency of monascorubin can reduce instead.

The determination of pulse pre-treatment temperature: as shown in Figure 2, pulse pre-treatment temperature rises to 60 DEG C from 30 DEG C, extracts Ascendant trend is presented in the color value of monascorubin in liquid, the Monascus color in monascorubin when temperature reaches 60 DEG C, red yeast rice orange, red yeast rice The color value value of yellow three kinds of components reaches peak, and total color value also reaches maximum value.Impulse electric field can generate fuel factor, and heat production Can make the temperature of processing solution can increase, since monascorubin is to decompose slightly at high temperature, so in order to improve red yeast rice color The Yield selection impulse electric field treatment temperature of element is 60 DEG C.

The determination of pulse number: from the figure 3, it may be seen that sample is handled 1-5 times under impulse electric field, due to just starting pulse number Low, there is no all rupture, monascorubins not to flow completely out for the cell of red yeast rice thallus, but increases monascorubin with pulse number Color value increase, after pulse number reaches 3 times, Monascus color in monascorubin, red yeast rice orange, red yeast rice Huang three kinds of components color Value reaches peak, and total color value also reaches maximum value, more than 3 times after the color value of monascorubin have a declining tendency, sample Temperature is increased significantly after having handled, and be might have partial pigment and is decomposed, so as to cause the decline of color value.Therefore pulse time Number should select 3 times.The determination of electric field strength: as shown in Figure 4, under conditions of electric field strength is 200-600 V/cm, due to rigid Beginning electric field strength is low, and the cell of red yeast rice thallus is there is no all ruptures, and monascorubin does not flow completely out, with electric field strength Raising, the color value of monascorubin also gradually rises.When electric field strength is 500 V/cm, cell membrane, the cell of red yeast rice thallus For wall by irreversible breakdown, the content of material of exudation reaches constant, so when electric field strength is further added by, the color value of monascorubin It is almost unchanged.Therefore it is more suitable to select 500 V/cm for electric field strength.

Test three: the single factor experiment of impulse electric field reinforcing ultrasound assisted extraction monascorubin

The determination of ultrasonic power: as shown in Figure 5, since the maximum power of instrument can only use 60% to instrument quota power, When ultrasonic power is gradually increased by 130 W-390 W, the color value of monascorubin is consequently increased.When ultrasonic power is 390 W When, the color value value of Monascus color in monascorubin, red yeast rice orange, red yeast rice three kinds of components of Huang reaches peak, and total color value also reaches most Big value.It is thus determined that 390 W of ultrasonic power is more suitable for.

The determination of ultrasonic wave action time: it will be appreciated from fig. 6 that experimental selection ultrasonic time is 10 min-30 min.Work as ultrasound When time is 30 min, the color value value of Monascus color in monascorubin, red yeast rice orange, red yeast rice three kinds of components of Huang reaches peak, always Color value also reaches maximum value.But when ultrasonic time continue extend when, monascorubin color value is but declined, this may be due to The too long fuel factor that produces of ultrasonic time causes the temperature of solution to increase, since monascorubin is to decompose slightly at high temperature , this will lead to the color value decline of monascorubin.The sonication treatment time known to analyzing above is that 30 min are most suitable.

The determination of ultrasonic temperature: as shown in Figure 7, red as ultrasonic temperature increases when ultrasonic temperature is at 40-60 DEG C The color value of bent pigment rises with it, when ultrasonic treatment temperature reaches 60 DEG C, Monascus color in monascorubin, and red yeast rice orange, red yeast rice Huang The color value value of three kinds of components reaches peak, and total color value also reaches maximum value.But ultrasonic temperature is when being more than 60 DEG C, red yeast rice The color value of pigment but reduces, this is because the fuel factor of ultrasonic wave causes the temperature of solution to increase, monascorubin is for a long time in height Unstable easy decomposition under temperature, therefore ultrasonic treatment temperature is advisable with 60 DEG C.

The verification test of optimum condition: the optimal conditions of each single factor test has finally been determined by single factor experiment.Herein Under the conditions of carry out verification test, as the result is shown: using monascorubin extract optimum condition when, i.e., in the electric field strength of impulse electric field For 500 V/cm, pulse number is 3 times, and pulse pre-treatment temperature is 60 DEG C, solid-liquid ratio 3:1(g/L) under conditions of handle, It then is 390 W in ultrasonic power, ultrasonic temperature is 60 DEG C, and sonication treatment time is handled under conditions of being 30 min, is measured red The uranidin color value of bent pigment is 239.8 U/g, and citraurin color value is 129.3 U/g, and haematochrome color value is 150.4 U/g, always Color value is 519.5 U/g.The ethanol solution with optimal conditions same volume is added in sample, 60 DEG C of 1 h of water-bath take supernatant Carry out the measurement of color value.The uranidin color value that ethanol solution extraction measures monascorubin is 439.8 U/g, and citraurin color value is 229.3 U/g, haematochrome color value are 350.4 U/g, and total color value is 519.5 U/g.Compared with traditional ethanol solution extraction, The effect that ultrasound assisted extraction monascorubin is strengthened in pulse improves 85.2%.The above only expresses preferred reality of the invention Mode is applied, the description thereof is more specific and detailed, and but it cannot be understood as limitations on the scope of the patent of the present invention.It should refer to Out, for those of ordinary skill in the art, without departing from the inventive concept of the premise, can also make several Deformation is improved and is substituted, and these are all within the scope of protection of the present invention.Therefore, the scope of protection of the patent of the present invention should be with appended Subject to claim.