CN102461642A - Natural refrigerated fresh meat film antistaling agent and application - Google Patents
Natural refrigerated fresh meat film antistaling agent and application Download PDFInfo
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- CN102461642A CN102461642A CN2010105408150A CN201010540815A CN102461642A CN 102461642 A CN102461642 A CN 102461642A CN 2010105408150 A CN2010105408150 A CN 2010105408150A CN 201010540815 A CN201010540815 A CN 201010540815A CN 102461642 A CN102461642 A CN 102461642A
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Abstract
The invention belongs to the field of natural product preparation, and particularly relates to a polymethoxylated flavonoids extract which is prepared from orange peels and application thereof. An antistaling agent provided by the invention is prepared from the following steps of: drying, grinding, screening and placing the orange peels into ethanol solution, adding cellulose, using a water bath for digestion and enzymolysis, and obtaining flavonoids ethanol extracting solution; then concentrating and removing the ethanol of the extracting solution, extracting with diethyl ether, merging a ether layer, and washing the ether layer until a water phase part is colorless; evaporating the diethyl ether, refrigerating, drying, and obtaining a polymethoxylated flavonoids crude; dissolving the polymethoxylated flavonoids crude in methyl alcohol, letting the solution to pass through a Sephadex LH-20 gel column, using the methyl alcohol-water to carry out gradient elution, adopting an ultraviolet detector to detect, collecting the maximum absorption peak, concentrating, refrigerating, drying and obtaining the polymethoxylated flavonoids extract; using absolute ethyl alcohol to redissolve, adding activated carbon to decolorize, concentrating, refrigerating, drying, and then obtaining polymethoxylated flavonoids extract powder; and compounding the polymethoxylated flavonoids extract with polysorbate and edible alcohol, then obtaining the antistaling agent. The antistaling agent can be used for keeping refrigerated fresh meat fresh.
Description
Technical field
The invention belongs to food processing technology field, be specifically related to a kind of preparation and application of natural cold fresh meat coating antistaling agent.
Background technology
Living standards of the people increase substantially in recent years, the cold fresh meat that an urgent demand eats clean hygiene, is of high nutritive value.Cold fresh meat is the main flow of fresh from now on meat consumption, and how prolonging its shelf-life is problem anxious to be solved at present.Cold fresh meat is rich in nutriment, and water activity is higher, in common processing, transportation, storage and process of consumption, is easy to receive contamination by micro and other Effect of Environmental, makes its generation putrid and deteriorated.Cold fresh meat from begin to produce today states such as America and Europe, Japan popularize and the developing rapidly of developing country, its shelf storage periods is the central issue (Hu Yanfei, Wang Changhai, 2005) that the production process of circulation is paid close attention to all the time.Cold fresh meat shelf life length depends primarily on initial bacterium number and Preservation Treatment mode (Lu Zhi etc., 2004 of raw meat; LI XUEMEI etc., 2005), reducing the initial bacterium number of cold fresh meat is the key issue that prolongs the cold fresh meat shelf-life with suppressing residual microbial growth breeding.At present, the cold fresh meat preservation method that generally adopts has both at home and abroad been obtained good result such as methods such as controlled atmospheric packing, microwave treatment, radiation treatment, HIGH PRESSURE TREATMENT, and the application of antisepsis antistaling agent also has more report.In recent years, the researcher is applied to the fresh-keeping of cold fresh meat with edible film-coating, wherein adopt single or composite fresh-keeping liquid such as shitosan, Nisin sprays or the cold fresh meat research of impregnation process report at most (Duan Jing rues etc., 2001; Sun Xiangjun etc., 2002; Jiang builds equality, and 2005).Add antistaling agent and have effective, easy and simple to handle, low cost and other advantages, become the normal method of using in actual production.But use chemical preservative to surpass certain quantity, can produce some adverse influences, therefore develop focus and the trend that natural antisepsis fresh-keeping agent becomes research from now on the health of human body.
Polymethoxyflavone is the characteristic component of citrus fruit, enjoys domestic and international concern owing to it has good biologically active.China's oranges and tangerines cultivated area and output have accounted for the No. 1 in the world at present; The brocade orange is that the sweet orange kind is used in the most important processing of China, contains abundant polymethoxyflavone in its pericarp, and a large amount of skin slags that processing produces do not obtain good use; Caused the significant wastage (single poplar, 2008) of resource.Therefore, from orange peel residue, separate, extract various functional components, utilize its distinctive physiological function to develop new food additives or medicine, China's Citrus resource deep development utilization is had important economic value and vast market prospect.According to research, the solubility of flavone compound has very big-difference because of structure and existence (aglycon, monoglycosides, disaccharide glycosides or three glucosides) difference.General free aglycon indissoluble or water insoluble is soluble in organic solvent such as methyl alcohol, ethanol, ethyl acetate, ether and the dilute alkaline aqueous solution, and wherein flavones, flavonols, chalcone are insoluble in water, and flavanone and flavanonol etc. are more soluble in water.In the intensive polar solvents such as flavonoid glycoside is generally soluble in water, methyl alcohol and ethanol.Sugar chain is long more, and then water solubility is big more.Therefore, adopt water extraction can not make that flavone compound effectively leaches in the oranges and tangerines, then can improve recovery rate greatly with ethanol.Exist with the bigger glucosides form of polarity because flavones is many in the sweet orange peel; Adopt at present both at home and abroad extensively the organic solvent that has than high polarity carry out lixiviate (Li et al., 2006a), bibliographical information has heat reflow method (Zhao Xuemei etc.; 2,003 2004), microwave auxiliary extraction method (Zhang Xionglu, 2005; Zhang Haide etc., 2003), ultrasonic wave auxiliary extraction method (Wang Wanneng etc., 2006; Jiang Zhiguo, 2006) etc., solvent is mainly methyl alcohol, ethanol etc.
It is on the basis of traditional solvent extraction process that enzymatic isolation method extracts; According to the formation of plant cell wall, utilize enzyme digestion reaction to have characteristics such as highly single-minded, select corresponding enzyme with hydrolysis of cell membrane constituent or degraded; Destroy cell wall structure; Active ingredient is fully come out, dissolve or be suspended in and extract in the solvent, thereby reach the purpose of extracting active ingredient in the cell.Since in the extracts active ingredients process the natural cover for defense---cell membrane is destroyed, thereby enzymatic isolation method helps improving the leaching rate of active ingredient, becomes the research focus of modern method for distilling (Toebes et al., 2005 in recent years; Chung et al., 2005).Existing report adopts enzymatic isolation method to extract aldehydes matter in the oranges and tangerines skin; Kind, extraction temperature and enzyme liquid concentration etc. with enzyme are measurement index; Discover that enzymatic isolation method is higher than water extraction efficient; Its essence be under the lower temperature enzyme can through degradation of cell wall polysaccharide reach preferably extraction effect (Li et al., 2006b).
This patent adopts cellulase that bright and beautiful orange peel is carried out enzymolysis processing, is the solvent lixiviate then with ethanol, and purpose is to inquire into the optimised process that Flavonoid substances extracts in the bright and beautiful orange peel.Do not see at present the report of relevant polymethoxyflavone as yet to the fresh-keeping application study of cooling meat; So this patent is the examination material with the chilled pork; Polymethoxyflavone is processed antistaling agent solution; To the fresh pork processing of filming, and adopt PE (polyethylene) aerobic packaged form, with the index that is changed to of color and luster, pH value, VBN value (TVB-N value), total number of bacteria and pseudomonad number; Estimate the fresh-keeping effect of polymethoxyflavone antistaling agent, for the application of polymethoxyflavone in food preservation provides foundation to cooling meat.
Summary of the invention
The object of the present invention is to provide a kind of natural cold fresh meat coating antistaling agent,, prolong the shelf life of cold fresh meat to improve the freshness of cold fresh meat.Main points of the present invention are to be a kind of polymethoxyflavone compounds that is applicable to that cold fresh meat is fresh-keeping of separation and Extraction the orange peel pericarp from orange peel; Utilize this antistaling agent to improve cold fresh meat permeability of cell membrane; Suppress the respiration of microorganism, thereby suppress the growth and the breeding of the spoilage organisms in the cold fresh meat.
The present invention realizes through following technical proposal:
A kind of natural cold fresh meat coating antistaling agent, the proportioning of each component is as follows:
To prepare the 1000ml antistaling agent: polymethoxyflavone extract powder 5g; Tween 10mL; The 900mL95% edible alcohol; Tween is added in the edible alcohol, and making the polymethoxyflavone final concentration in the solution is 5.0mg/mL.Described natural cold fresh meat coating antistaling agent prepares through following steps:
1) orange peel (for example sweet orange kind brocade is orange, but is not limited thereto) is dried to constant weight under 40 ℃, pulverize the back and cross 40 mesh sieves, obtain the orange peel powder;
2) it is 1: 20 by mass volume ratio; 100g orange peel powder is placed the ethanolic solution of 80% concentration, in described ethanolic solution, add the cellulase (analyze pure, available from Shanghai east wind biochemical technology Co., Ltd product) of 5% mass fraction again; Transfer pH to 4; With 60 ℃ of water-bath lixiviates, enzymolysis time is 2h, obtains the flavones ethanol extract;
3) the flavones ethanol extract is concentrated removing alcohol, is to add extracted with diethyl ether 3 time at 1: 30 by mass volume ratio, makes the whole strippings of active ingredient, merges three times the ether layer;
4) the NaOH weak caustic solution with 0.4% concentration washs described ether layer, to aqueous portion colourless till, the evaporative removal ether;
5) extract of step 4) is put in the freeze drier (Germany produces the christ freeze drier, and model BETR 2-8LD plus is according to the product description operation), be 28-32h drying time, gets polymethoxyflavone crude extract (about 564mg);
6) the polymethoxyflavone crude extract is redissolved in methanol solution; (2.5cm * 30cm), adopt the methanol-water gradient elution detects in 340nm with Ultraviolet Detector to cross Sephadex LH-20 gel column; Collect the product of maximum absorption band; Concentrated frozen dry the higher polymethoxyflavone extract of purity, the polymethoxyflavone extract of gained is carried out qualitative and quantitative analysis with high performance liquid chromatograph (HPLC), and employing LC-MS analytical method is identified to its structure.
7) the polymethoxyflavone extract with step (6) gained redissolves with absolute ethyl alcohol, is to add activated carbon decolorizing at 1: 7 by mass ratio, concentrates, and freeze drying gets the polymethoxyflavone extract powder;
8) with the polymethoxyflavone extract powder 5g of step 7) gained; Use the edible alcohol dissolving of 95% concentration of the 900mL that is added with the 10mL tween; Making the final concentration of the polymethoxyflavone extract in the solution is 5.0mg/mL, is natural cold fresh meat coating antistaling agent.
The application process of natural cold fresh meat coating antistaling agent in cold fresh meat is fresh-keeping of the present invention's preparation is; Cold fresh meat is cut into strip or bulk; The amount of smearing 10mL according to the heavy 10g of meat spreads upon described antistaling agent on the surface of cube meat or cutlet; Dry the back and pack with polyethylene film, put 4 ℃ ± 1 ℃ refrigerated condition storage down, the shelf-life can reach 8 days.
Of the present invention and existing cold fresh-keeping agent for meat is compared and is had the following advantages:
1, the present invention make full use of oranges and tangerines leftover bits and pieces pericarp extract as crude antistaling agent, improved the value of orange peel, practiced thrift resource.
2, antistaling agent of the present invention has the broad-spectrum antibacterial activity, has prolonged the shelf-life and the shelf life of cold fresh meat.
3, antistaling agent of the present invention has no adverse effects to cold fresh meat quality and sense organ.
Description of drawings
Fig. 1: be in the embodiments of the invention enzyme liquid concentration to the impact analysis of general flavone yield in the orange peel.
Fig. 2: be in the embodiments of the invention hydrolysis temperature to the impact analysis of general flavone yield in the orange peel.
Fig. 3: be in the embodiments of the invention enzymolysis time to the impact analysis of general flavone yield in the orange peel.
Fig. 4: be in the embodiments of the invention pH value to the impact analysis of general flavone yield in the orange peel.
Fig. 5: be in the embodiments of the invention solid-liquid ratio to the impact analysis of general flavone yield in the orange peel.
Fig. 6: be in the embodiments of the invention concentration of alcohol to the impact analysis of general flavone yield in the orange peel.
Fig. 7: the HPLC that is polymethoxyflavone extract in the embodiments of the invention analyzes.
Fig. 8: the chemical structure analysis that is polymethoxyflavone in the embodiments of the invention.
Fig. 9: the cation that is polymethoxyflavone extract in the embodiments of the invention detects ESI-MS total ion current figure under the pattern.
Figure 10: be in the embodiments of the invention antistaling agent of the present invention to the impact analysis of cooling meat brightness.
Figure 11: be in the embodiments of the invention antistaling agent of the present invention to cooling off the impact analysis of flesh-coloured degree.
Figure 12: be in the embodiments of the invention antistaling agent of the present invention to the impact analysis of cooling meat pH value.
Figure 13: be in the embodiments of the invention antistaling agent of the present invention to the impact analysis of VBN in the cooling meat.
Figure 14: be in the embodiments of the invention antistaling agent of the present invention to the impact analysis of total number of bacteria in the cooling meat.
Figure 15: be in the embodiments of the invention crude antistaling agent of the present invention to the impact analysis of pseudomonad in the cooling meat.
The specific embodiment
Embodiment 1 (preparing embodiment basically)
The oranges and tangerines kind of present embodiment is sweet orange kind " a brocade orange ", and this kind is a sweet orange kind in the citrus fruit, and this kind originates in Jiangjin, Sichuan (at present belonging to Chongqing City's Jiangjin District).The sixties are introduced Hubei, for the master in Hubei plants the oranges and tangerines kind.Peel the present embodiment Jincheng collected in Hubei songzi Wei water citrus farm, the data were collected Jincheng peel preserved frozen at -18 ℃.
Test method is following:
1) orange peel (the sweet orange kind: brocade is orange, but is not limited thereto kind) is dried to constant weight under 40 ℃, pulverize the back and cross 40 mesh sieves, obtain the orange peel powder;
2) it is 1: 20 by mass volume ratio; 100g orange peel powder is placed the ethanolic solution of 80% concentration, in described ethanolic solution, add the cellulase (analyze pure, available from Shanghai east wind biochemical technology Co., Ltd product) of 5% mass parts again; Transfer pH to 4; With 60 ℃ of water-bath lixiviates, enzymolysis time is 2h, obtains the flavones ethanol extract;
3) the flavones ethanol extract is concentrated removing alcohol, is to add extracted with diethyl ether 3 time at 1: 30 by mass volume ratio, makes the whole strippings of active ingredient, merges three times the ether layer;
4) the NaOH weak caustic solution with 0.4% concentration washs described ether layer, to aqueous portion colourless till, the evaporative removal ether;
5) extract of step 4) is put into freeze drier, be 28-32h drying time, gets the polymethoxyflavone crude extract;
6) the polymethoxyflavone crude extract is redissolved in methanol solution; Cross the Sephadex LH-20 gel column of 2.5cm * 30cm, adopt the methanol-water gradient elution, with Ultraviolet Detector (HID-5 computer Ultraviolet Detector; Nanjing University's product; The method operation of by specification) detects in the 340nm place, collect maximum absorption band, the dry polymethoxyflavone extract that gets of concentrated frozen;
7) the polymethoxyflavone extract with step (6) gained redissolves with absolute ethyl alcohol, is to add activated carbon decolorizing at 1: 7 by mass ratio, concentrates, and freeze drying gets the polymethoxyflavone extract powder;
8) with the polymethoxyflavone extract powder 5g of step 7) gained; Use the edible alcohol dissolving of 95% concentration of the 900mL that is added with the 10mL tween; Making the final concentration of the polymethoxyflavone extract in the solution is 5.0mg/mL, is natural cold fresh meat coating antistaling agent.
Embodiment 2: (method for optimizing embodiment)
The material source of present embodiment is with embodiment 1.Experimental technique is with reference to the basic step according to embodiment 1.The applicant is in the enzymolysis and extraction process of test comparing embodiment; Investigated the influence of enzyme dosage, reaction pH value, hydrolysis temperature, enzymolysis time and concentration of alcohol and solid-liquid ratio respectively to hydrolysis result; And enzymatic hydrolysis condition optimized, to confirm the optimum process condition of enzymolysis.
(1) cellulose enzyme liquid concentration is to the influence of general flavone leaching rate
Cellulase in the present embodiment is pure for analyzing, available from Shanghai east wind biochemical technology Co., Ltd product.The enzyme liquid concentration gradient of the cellulase of present embodiment is set at 2%, 3%, 4%, 5%, 6% respectively, and solid-liquid ratio (in mass volume ratio) is 1: 10, and concentration of alcohol is 80%, and the pH value is 5, and bath temperature is 50 ℃, and the water-bath time is 3h.The general flavone leaching rate is as shown in Figure 1 in the extract of measuring.The general flavone leaching rate is the highest when cellulase concentration is 4%, reaches 2.23 ± 0.04%, continues the concentration of the plain enzyme of increased fiber, and the general flavone leaching rate then descends to some extent.
(2) the cellulase degradation temperature is to the influence of general flavone leaching rate in the orange peel
Be provided with 30 ℃, 40 ℃, 50 ℃, 60 ℃, 70 ℃ five in the present embodiment and extract temperature levels; The enzyme liquid concentration of cellulase is 4%; All the other conditions are constant, investigated the influence of cellulase degradation temperature to the general flavone leaching rate, and result of the test is as shown in Figure 2.Can be known that by Fig. 2 extracting temperature is that the general flavone leaching rate in the orange peel below 50 ℃ increases with the temperature rising, the general flavone leaching rate when temperature reaches 50 ℃ in the orange peel is maximum.
(3) the cellulase degradation time is to the influence of general flavone leaching rate in the orange peel
The enzyme liquid concentration of design cellulase is 4%, and hydrolysis temperature is 50 ℃, and other conditions are constant.The cellulase hydrolysis time is as shown in Figure 3 to the result of general flavone leaching rate in the orange peel.Along with the increase of enzymolysis time, the general flavone recovery rate in the orange peel improves gradually, and the general flavone leaching rate is maximum during to 2.5h, continue to prolong enzymolysis time flavones recovery rate leaching rate and descend on the contrary, so the enzymolysis time of cellulase is advisable with 2.5h.
(4) the pH value is to the influence of general flavone leaching rate in the orange peel
In view of the activated centre of cellulase is the slant acidity condition, to work as enzyme liquid concentration be 4% so investigated, 50 ℃ of hydrolysis temperatures, and enzymolysis time 2.5h when other conditions are constant, has compared the influence of different pH values to general flavone lixiviate from orange peel.The result is as shown in Figure 4, and cellulase activity is higher in sour environment, this be since enzyme activity influenced by pH bigger; When pH is 4, help complex enzyme and bring into play maximum vigor; Destroy cell membrane, reduce resistance to mass tranfer, thereby help the extraction of general flavone in the bright and beautiful orange peel; Too high or too lowly all be unfavorable for enzymolysis, so extracting efficiency is lower.
(5) solid-liquid ratio is to the influence of general flavone leaching rate in the orange peel
Embodiment adds ethanol with 1: 5,1: 10,1: 15,1: 20,1: 25 material (orange peel) liquor ratio respectively; The enzyme liquid concentration of cellulase is 4%, 50 ℃ of hydrolysis temperatures, enzymolysis time 2.5h; The pH value of solution value is 4 o'clock, has investigated the influence of different feed liquid comparison lixiviate ancestor flavones.Can be found out that by Fig. 5 the general flavone leaching rate reaches maximum when the feed liquid mass volume ratio is 1: 20, solid-liquid ratio is that 1: 25 o'clock general flavone leaching rate and 1: 20 o'clock leaching rate difference are little.Consider reasons such as financial cost, the suggestion solid-liquid ratio is selected for use and is got final product at 1: 20.
(6) influence of general flavone leaching rate in the concentration of alcohol orange peel
The enzyme liquid concentration of design cellulase is decided to be 4%, and concentration of alcohol is a variable, and hydrolysis temperature is decided to be 50 ℃; Enzymolysis time is decided to be 2.5h; The pH value of solution value is decided to be 4, and solid-liquid ratio is decided to be at 1: 20 o'clock and investigates the influence of different ethanol concentration to general flavone leaching rate in the orange peel, and test effect is as shown in Figure 6.When concentration of alcohol is 80%; The general flavone leaching rate is the highest in the orange peel, continues to increase the concentration leaching rate and descends on the contrary, and this is because some low pole composition stripping quantities increase; Slowed down of the diffusion of orange peel flavones, thereby caused the orange peel flavones to extract the reduction of yield to the main body solvent.Therefore select 80% alcohol extract best results for use.
(7) orthogonal experiment of cellulose enzyme solution
According to the experiment of single factor result; Be that (cellulase of present embodiment is pure for analyzing for the microorganism of enzymolysis with the cellulase; Available from Shanghai east wind biochemical technology Co., Ltd product), selecting enzyme liquid concentration, hydrolysis temperature, enzymolysis time, reaction pH value for use is influence factor, has designed the orthogonal experiment of 4 factors, 3 levels; See table 1, table 2 is seen in result of the test and data analysis.Can be known that by table 2 best of breed of four factors is A2B3C1D2, promptly when enzyme liquid concentration 5%, 60 ℃ of hydrolysis temperatures, enzymolysis time 2h, pH value 4, the general flavone leaching rate is the highest.The secondary factors that influences hydrolysis result is B＞D＞A＞C successively, and promptly hydrolysis temperature is the most remarkable to the influence of hydrolysis result, and next is followed successively by pH value, enzyme concentration, enzymolysis time.After optimizing extraction process, the general flavone leaching rate reaches 2.67 ± 0.06% in the sweet orange peel.
The L9 (3 of table 1 cellulase degradation condition
4) orthogonal array
Table 2 Orthogonal experiment results
Data are mean SD. in a table
Embodiment 3: the isolation identification of polymethoxyflavone extract in the bright and beautiful orange peel
The preparation of the polymethoxyflavone extract of present embodiment obtains with reference to the method for embodiment 1.
High performance liquid chromatography (HPLC) analytical method:
The polymethoxyflavone extract powder of getting 1mg the present invention preparation is dissolved in the 10mL methyl alcohol, excessively behind the 0.45 μ m filter membrane, with WatersHPLC and PDAD (PDA) quantitative analysis.Chromatographic condition: chromatographic column Hypersil ODS C18 post (250mm * 4.6mm, Yi Lite); Detect wavelength: 330nm; Flowing phase: the 1.5% glacial acetic acid aqueous solution-acetonitrile, with 50% acetonitrile solution wash-out 25min; Flow velocity: 0.6mL/min; Sample size: 20 μ L; Column temperature: 30 ℃.UV scanning scope: 200-400nm.Chromatographic column needs balance 15min before the last appearance.Nobiletin and hesperetin content adopt external standard method to confirm.All the other constituent contents are that interior mark conduct is with reference to sxemiquantitative through adding the Radix Astragali plain (maximum absorption band is 330nm).
Fig. 7 is the HPLC analysis chart of the polymethoxyflavone extract of the present invention's preparation under the 330nm, contains 7 kinds of components altogether in the polymethoxyflavone extract of demonstration the present invention preparation among the figure, and main component is 1,3,4,5,7 peaks.Wherein retention time is the 3# peak-to-peak value maximum of 11.454min; Relative amount reaches 32.7%; Secondly be the 4# peak of 12.103min for retention time; Relative amount is about 20.1%, and retention time is the 1# peak of 9.574min, the 7# peak of 14.803min and the 5# peak of 12.859min, and its relative amount is respectively 17.6%, 15.6% and 8.3%.
The structure of polymethoxyflavone is identified in the brocade orange peel:
Sample is with Agilent HPLC-MS/ESI network analysis.Consider compatibility; Employing ZORBAX Eclipse XDB-C18 post (2.1mm * 150mm I.D., Agilent), flowing phase is methyl alcohol (A)-1.5% glacial acetic acid aqueous solution (B); Elution requirement is changed into the interior B of 65min by 35-71%, and B is by 71% to 35% in the 65-70min.Sample introduction is before with 35%B balance 10min once more.Flow velocity: 0.25mL/min, sample size: 10 μ L.
The mass spectrum condition: the ESI ion gun, injection electric: 4500V, expulsion pressure: 30psi, dry gas flow velocity: 8L/min, the dry gas temperature: 250 ℃, quality of scanning scope: 100-1000m/z, positive ion mode.
Table 3 and Fig. 8 show that LC-MS analyzes the structure of the polymethoxyflavone one-component that obtains, and through identifying with reporting its spectroscopic data of document comparison.Data and document are very identical, and data in literature derives from the analysis of tangerine oil, fruit juice and pericarp (Wang et al., 2007; Zhou et, al., 2007; Lu et al., 2006; Mata Bilbao, 2007; Raman et al., 2005; Dugo et al., 2000).Fig. 9 detects the total ion current figure under the pattern for polymethoxyflavone at cation, considers compatibility, and flowing phase is adjusted, and delay has taken place appearance time, but peak sequence does not change.Table 3 analytic induction the molecular weight and the structural analysis of each component of polymethoxyflavone.
The molecular weight and the structural analysis of table 3 polymethoxyflavone one-component
Annotate: "～" expression acromion
Embodiment 4: (applicating example of antistaling agent of the present invention in the cold fresh meat of pig)
The source of the cold fresh meat of pig: the used cold fresh meat bacterium of pig of present embodiment and other embodiment is available from hundred storage supermarkets in the Lopa Nationality lion road, Hongshan District, Wuhan City.
The reagent source of microbe culture: pseudomonas isolation agar (analyze pure, available from the extensive and profound in meaning star biotechnology in Beijing Co., Ltd); Nutrient agar (analyze pure, available from Qingdao Hai Bo Bioisystech Co., Ltd).
The preparation of polymethoxyflavone extract powder and the preparation of antistaling agent are with reference to the method for embodiment 1.
The processing of cooling meat: select the cooling meat of pig longissimus dorsi muscle, at first evenly spray 75% alcohol and carry out surface disinfection, be cut into the square little cube meat about 10g, be divided into two groups at random, be placed on respectively in the polyester pallet of having sterilized on the pork surface.Processed group is evenly smeared 10mL polymethoxyflavone solution with aseptic hairbrush at meat surface; Control group is evenly smeared 95% (in percentage concentration) edible alcohol that 10mL contains 1% tween at meat surface with aseptic hairbrush; Handle all at desinfection chamber normal temperature for two groups and place and dry the surface; Pack with the polyethylene preservative film; Put into 4 ± 1 ℃ refrigerator immediately and deposit, respectively at measuring physics and chemistry and microbiological indicator on the the 0th, 2,4,6 and 8 day.
Physical and chemical index is estimated:
(1) mensuration (L of color and luster
*Value, a
*Value): get meat appearance and in cuvette, measure with chromascope (available from U.S. HunterLab Company products).From Figure 10-11, can find out, the color and luster variation tendency basically identical of processed group and control group cooling meat in storage process, the brightness value of cooling meat and red scale value all present elder generation's rising and reduce variation tendency again.But on the whole, control group is than the brightness value height of processed group cooling meat and red scale value is low, explain that smearing antistaling agent of the present invention can reduce and cool off meat brightness and improve red degree.
(2) pH pH-value determination pH: adopt pH meter (410A type PH meter; The thick product of U.S. Thermo orion company); According to the standard GB/T9695.5-2008 of intelligent planning people's republic " meat and meat products pH measure ", China Standard Press, the method for 2009 editions introductions detects.10g meat appearance is rubbed the back add 40mL distilled water, filtrate filtered is measured its pH value with above-mentioned pH meter.As can beappreciated from fig. 12, the pH value variation tendency basically identical of control group and processed group cooling meat in storage process, the pH value of the cooling meat of contrast raises gradually, but the cooling meat pH value of the ratio control group of processed group is low.Explain that antistaling agent of the present invention brought into play bacteriostatic activity in cooling meat storage process, effectively reduce the growth of microorganism breeding, reduced the decomposition rate of microorganism, thereby reached certain fresh-keeping effect protein.
(3) VBN (TVB-N) detects: according to State Standard of the People's Republic of China GB/T5009.44-2003 " analytical method of meat and meat products sanitary standard "; China Standard Press; The method of 2003 editions introductions is carried out, and adopts the semimicro nitriding.Concrete steps are: get 10g meat appearance in 100mL distilled water, filter behind the jolting 30min, distill titration.The content of VBN calculates by following formula in the sample:
In the formula:
X---the content of TVB-N in the sample, unit is milligram every hectogram (mg/100g);
V
1---measure with appearance liquid and consume the hydrochloric acid standard solution volume, unit is a milliliter (mL);
V
2---reagent blank consumes the hydrochloric acid standard solution volume, and unit is a milliliter (mL);
The actual concentrations of c---hydrochloric acid standard solution, unit are every liter (mol/L) of mole;
14---the quality of the nitrogen suitable with 1.00mL hydrochloric acid standard titration solution [c (HCL)=1.000mol/L], unit is a milligram (mg);
M---sample mass, unit is gram (g).
As can be seen from Figure 13; The TVB-N value is 13.2mg/100g when the 6th day of storage; Reaching country-level fresh meat sanitary standard, was 16.6mg/100g in the time of the 8th day, reached national secondary fresh meat sanitary standard; And control group in the time of the 6th day the TVB-N value greater than 20.0mg/100g, near putrid and deteriorated.
2.4 microbiological indicator detects
Adopt flat board to pour into counting method (reference: Niu Tiangui chief editor, Food Microbiology experimental technique, China Agricultyre University Press) total number of bacteria and detect the use nutrient agar, cultivate 24h for 37 ℃.The pseudomonad number detects and adopts the pseudomonas isolation agar culture medium, cultivates 48h for 27 ℃.Aforesaid operations all carries out in gnotobasis.
Clearly stipulate one-level fresh meat total number of bacteria≤10 among the State Standard of the People's Republic of China GB/T5009.44-2003 " analytical method of meat and meat products sanitary standard "
6Individual/g; Secondary fresh meat 10
6Individual/g＜total number of bacteria≤10
7Individual/g.Corrupt total amount of meat bacteria＞10
8Individual/g.Shown in Figure 14-15; The cold fresh meat that uses crude antistaling agent of the present invention to film to handle is in reduced levels 0-4 days total number of bacteria, is to increase rapidly after 3.54, the 4 days to the 4th day total number of bacteria logarithm value; Total number of bacteria was 4.93 in the 6th day; Belong to the one-level fresh meat, the total number of bacteria logarithm value is 6.79 when 8d, belongs to the secondary fresh meat.And total number of bacteria increase in control group 0-4 days is very fast, and the total number of bacteria logarithm value has reached 6.00 in the time of the 4th day, in the time of the 8th day the total number of bacteria logarithm value up to 8.48, putrid and deteriorated fully.Pseudomonad is similar with the total number of bacteria variation tendency, and it is very fast that a control group 0-4 days pseudomonad is counted increase, and processed group pseudomonad number was in reduced levels at 0-4 days and crude antistaling agent is filmed.The pseudomonad number increases comparatively fast after the 4th day, but compares with control group, and processed group pseudomonad number obviously is in reduced levels.The control group pseudomonad is counted logarithm value and reaches 4.48 during to the 6th day, and processed group is merely 1.90.The control group pseudomonad is counted logarithm value and reaches 4.83 when storing the 8th day, and processed group is 2.85.Show that crude antistaling agent of the present invention can kill or suppress the growth and the breeding of cold fresh meat spoilage organisms, thereby total number of bacteria is obviously reduced.
The main reference document
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Claims (2)
1. natural cold fresh meat coating antistaling agent is characterized in that each set of dispense is such as shown in down:
To prepare the 1000ml antistaling agent:
Polymethoxyflavone extract powder 5g;
Tween 10mL
The edible alcohol of 900mL95% concentration;
It prepares through following steps:
1) orange peel is dried to constant weight under 40 ℃, pulverize the back and cross 40 mesh sieves, obtain the orange peel powder;
2) be 1: 20 by mass volume ratio, described orange peel powder is placed the ethanolic solution of 80% concentration, in described ethanolic solution, add the cellulase of 5% mass parts again; Transfer pH to 4; With 60 ℃ of water-bath lixiviates, enzymolysis time is 2h, obtains the flavones ethanol extract;
3) the flavones ethanol extract is concentrated removing alcohol, is to add extracted with diethyl ether 3 time at 1: 30 by mass volume ratio, makes the whole strippings of active ingredient, merges three times the ether layer;
4) the NaOH weak caustic solution with 0.4% concentration washs described ether layer, to aqueous portion colourless till, the evaporative removal ether;
5) extract of step 4) is put into freeze drier, be 28-32h drying time, gets the polymethoxyflavone crude extract;
6) the polymethoxyflavone crude extract is redissolved in methanol solution, cross Sephadex LH-20 gel column, adopt the methanol-water gradient elution, detect in the 340nm place, collect maximum absorption band, the dry polymethoxyflavone extract that gets of concentrated frozen with Ultraviolet Detector;
7) the polymethoxyflavone extract with step (6) gained redissolves with absolute ethyl alcohol, is to add activated carbon decolorizing at 1: 7 by mass ratio, concentrates, and freeze drying gets the polymethoxyflavone extract powder;
8) with the polymethoxyflavone extract powder 5g of step 7) gained; Use the edible alcohol dissolving of 95% concentration of the 900mL that is added with the 10mL tween; Making the final concentration of the polymethoxyflavone extract in the solution is 5.0mg/mL, is natural cold fresh meat coating antistaling agent.
2. the application of the described natural cold fresh meat coating antistaling agent of claim 1 in cold fresh meat is fresh-keeping; It is characterized in that; Cold fresh meat is cut into strip or bulk; The amount of smearing 10mL according to the heavy 10g of meat spreads upon the surface of cube meat or cutlet with described antistaling agent, dries the back and pack with polyethylene film, puts 4 ℃ ± 1 ℃ refrigerated condition and preserves down.
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| CN102823919A (en) * | 2012-09-19 | 2012-12-19 | 昆明理工大学 | Method for extracting insoluble polyphenol in plant dross |
| CN102960423A (en) * | 2012-12-11 | 2013-03-13 | 浙江海洋学院 | Special compound preservative for fish |
| CN103385283A (en) * | 2013-05-08 | 2013-11-13 | 南京年吉冷冻食品有限公司 | Preparation method for antiseptic insect-proof antistaling agent used for bacon |
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| CN114645069A (en) * | 2022-03-28 | 2022-06-21 | 三峡大学 | Polymethoxylated flavone, and full-aqueous phase preparation method and application thereof |
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| CN102823919A (en) * | 2012-09-19 | 2012-12-19 | 昆明理工大学 | Method for extracting insoluble polyphenol in plant dross |
| CN102960423A (en) * | 2012-12-11 | 2013-03-13 | 浙江海洋学院 | Special compound preservative for fish |
| CN102960423B (en) * | 2012-12-11 | 2014-08-20 | 浙江海洋学院 | Special compound preservative for fish |
| CN103385283A (en) * | 2013-05-08 | 2013-11-13 | 南京年吉冷冻食品有限公司 | Preparation method for antiseptic insect-proof antistaling agent used for bacon |
| CN103385283B (en) * | 2013-05-08 | 2014-06-25 | 南京年吉冷冻食品有限公司 | Preparation method for antiseptic insect-proof antistaling agent used for bacon |
| CN104222255B (en) * | 2014-09-11 | 2016-08-17 | 江苏省农业科学院 | A kind of biological fresh-keeping method extending the aquatic products shelf-life |
| CN104222255A (en) * | 2014-09-11 | 2014-12-24 | 江苏省农业科学院 | Biological preservation method capable of prolonging shelf life of aquatic products |
| CN107087792A (en) * | 2017-04-26 | 2017-08-25 | 湘西华方制药有限公司 | A kind of polymethoxyflavone and preparation method thereof |
| CN108739974A (en) * | 2018-05-08 | 2018-11-06 | 浙江海洋大学 | One kind compounding natural sandy gravel for raw fish |
| CN108739974B (en) * | 2018-05-08 | 2022-04-05 | 浙江海洋大学 | Natural biological preservative compounded for sashimi |
| CN109090208A (en) * | 2018-09-06 | 2018-12-28 | 辽宁石油化工大学 | A kind of preparation method of the natural coating antistaling agent of cold fresh meat |
| CN114645069A (en) * | 2022-03-28 | 2022-06-21 | 三峡大学 | Polymethoxylated flavone, and full-aqueous phase preparation method and application thereof |
| CN114645069B (en) * | 2022-03-28 | 2023-08-22 | 三峡大学 | Polymethoxy flavone and its full water phase preparation method and application |
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