CN106957889A - When prediction ultrasonic assistant based on Mathematical Modeling is pickled in pickling liquid content of microorganisms method - Google Patents

When prediction ultrasonic assistant based on Mathematical Modeling is pickled in pickling liquid content of microorganisms method Download PDF

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CN106957889A
CN106957889A CN201710173823.8A CN201710173823A CN106957889A CN 106957889 A CN106957889 A CN 106957889A CN 201710173823 A CN201710173823 A CN 201710173823A CN 106957889 A CN106957889 A CN 106957889A
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pickling liquid
ultrasonic
pickled
microorganisms
mathematical modeling
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张万刚
康大成
蒋雨鹤
葛庆丰
周光宏
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Nanjing Agricultural University
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
    • C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
    • C12Q1/06—Quantitative determination
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
    • C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
    • C12Q1/10—Enterobacteria
    • G—PHYSICS
    • G06—COMPUTING; CALCULATING OR COUNTING
    • G06F—ELECTRIC DIGITAL DATA PROCESSING
    • G06F30/00—Computer-aided design [CAD]
    • G06F30/20—Design optimisation, verification or simulation

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Abstract

When prediction ultrasonic assistant based on Mathematical Modeling is pickled in pickling liquid content of microorganisms method.When being pickled there is provided a kind of prediction ultrasonic assistant based on Mathematical Modeling in pickling liquid content of microorganisms method, different ultrasonic treatment times are substituted into Mathematical Modeling to the calculated value for calculating viable count in pickling liquid, theoretical reference is provided applied to pickling liquid control of microorganisms for enterprise.Comprise the following steps：By the initial number N of tested microorganism0, the quantity N that processing substitutes into tested microorganism after calculating ultrasonication t minutes in formula for t minute is carried out to pickling liquid using ultrasonic wave：The present invention first determines time δ and profile shape parameter p of a certain strain in pickling liquid under the conditions of the ultrasonication needed for one logarithm value of micro organism quantity decline under a certain ultrasonic intensity, content of microorganisms in pickling liquid after determination is handled in the initial content and the treatment time substitution cold bactericidal formulas of Weibull before then ultrasonic microbial ripple auxiliary is pickled.

Description

Content of microorganisms in pickling liquid when prediction ultrasonic assistant based on Mathematical Modeling is pickled Method
Technical field
The present invention relates to food processing field, and in particular to when a kind of prediction ultrasonic assistant based on Mathematical Modeling is pickled The method of content of microorganisms in pickling liquid, sets up the sterilized model of ultrasonic wave cold under the conditions of the different ultrasonic intensities of a certain strain, is Ultrasonic sterilization time offer theoretical reference is provided in actual production.
Background technology
Meat products pickle be CARCASS QUALITY an important step, current meat pickles antiseptic preservation simple from the past It is changed into improve local flavor and color, improves the manufacturing process of meat.But not only time-consuming for conventional dead curing process, and And be also easy to produce and pickle the phenomenons such as uneven, microorganism pollution, it is difficult to adapt to the requirement of modernization industry production.Research shows, surpasses Cavitation effect when sound wave auxiliary is pickled can improve meat quality, improve the tenderness and water-retaining property of product, while when reduction is pickled Between, the usage amount of salt in pickling liquid is reduced, in terms of ultrasonic sterilization, ultrasonication is produced to microorganism wall and cell membrane Raw destruction, while the free radical produced can further crack somatic cells film, so as to reach the effect for reducing micro organism quantity Really.Therefore pickling liquid is pre-processed using ultrasonic technology in meat curing process, reduces the wind of contamination of products microorganism Danger, expands application of the ultrasonic wave in meat is pickled significant.Applicant submitted on the 17th in September in 2015 The application for a patent for invention of " a kind of to be used to quickly determine the model of beef salt content and its application when ultrasonic assistant is pickled ", specially Profit number is " 201510594474.8 ", by pickling dynamic analysis to ultrasonic intensity and ultrasonic assistant, can quickly be determined Salt content in beef when ultrasonic assistant is pickled, being conducive to enterprise to be carried out when producing to salinity in beef quickly, calculating and technique change Enter.
Intensity, frequency, treatment time of ultrasonication etc. are to influence the principal element of ultrasonic sterilization effect.Study table It is bright, high intensity (>4kW), low frequency (20~40kHz) and for a long time (>60min) ultrasonication can significantly reduce microbe quantity Amount.Most researchs achieve the technology connected applications such as ultrasonic wave and super-pressure, heat treatment, organic acid and sodium hypochlorite at present Preferable bactericidal effect (Alarcon-Rojo, Janacua, Rodriguez, Paniwnyk, ＆Mason, 2015；Cichoski, et al.,2015；Evelyn&Silva,2016).But aspect is pickled in ultrasonic assistant, Smith (2011) researchs are found, Ultrasonic assistant pickles processing and salmonella and Escherichia coli quantity is not made significant difference.In addition, at present typically with traditional meter Number methods determine the content of each quasi-microorganism after ultrasonication, and cumbersome, workload is big, it is impossible to easily determine ultrasonic wave The time of optimization process when auxiliary is pickled.
The content of the invention
The pickling liquid when present invention is pickled for problem above there is provided a kind of prediction ultrasonic assistant based on Mathematical Modeling The method of middle content of microorganisms, different ultrasonic treatment times is substituted into Mathematical Modeling the calculating for calculating viable count in pickling liquid Value, theoretical reference is provided for enterprise applied to pickling liquid control of microorganisms.
The technical scheme is that：Comprise the following steps：
1), green meat is cleaned up and removed after fat, manadesma, in immersion pickling liquid, is determined using colony counting method The initial number N of tested microorganism0；
2) processing t minutes, is carried out to pickling liquid using ultrasonic wave；
3), by N0The quantity N that calculates ultrasonication t minute after tested microorganism is substituted into formula (1) with t：
Wherein：N0Micro organism quantity in pickling liquid before finger ultrasonication, t refers to ultrasonic treatment time (min), and δ refers to super Micro organism quantity declines the time (min) needed for a logarithm value in pickling liquid under sonicated, and p refers to the form parameter of curve, p >1 is convex curve, and p=1 is straight line, p<1 is concave curve.
δ and p measuring and calculating step is in the formula (1)：
1) tested microorganism sample, is taken, its growth curve is determined and determines optimal incubation time；
2) viable bacteria, is prepared under the conditions of optimal incubation time and connects bacterium solution, its initial microbial population N is determined0’。
3), fresh beef is cleaned up and removed after fat, manadesma, in immersion pickling liquid, bacterium solution is connect by what is prepared Add in pickling liquid, handled using ultrasonic wave；
4) two plants of bacterium in pickling liquid when, colony counting method calculates different ultrasonic intensities, difference ultrasonic treatment time t ' Viable count N '；
5), by N0', t ' and N ' substitute into formula (1), solve equation calculating using MATLAB softwares, to obtain ultrasonic wave strong δ and p value under degree, are programmed as follows in MATLAB softwares：
[x y]=solve (' logN0’-logN1'=(t1'/x) ^y', ' logN0’-logN2'=(t2/x)^y','x',' Y'),
Wherein：X refers to δ, and y refers to p.
The step 1) in pickling liquid include 60g/L sodium chloride, 15g/L sucrose, 4g/L composite phosphate.
The step 2) in using ultrasonic probe is submerged in pickling liquid at from meat surface 2cm during ultrasonication.
The step 2) in ultrasonication intensity be more than or equal to 6.23Wcm-2。
The beneficial effects of the invention are as follows：First determine that a certain strain is under the conditions of the ultrasonication under a certain ultrasonic intensity Micro organism quantity declines the time δ and profile shape parameter p needed for a logarithm value in pickling liquid, then by the ultrasonic microbial Ripple aids in the initial content before pickling and treatment time to substitute into micro- life in pickling liquid after determination processing in the cold bactericidal formulas of Weibull Thing content, this method practical simplicity is workable, is that manufacturing enterprise carries out meat products product control of microorganisms and really when pickling Fixed sterilization parameter provides theoretical reference, while being conducive to ultrasonic assistant to pickle the popularization and application of new technology.
Brief description of the drawings
Fig. 1 is Escherichia coli O 157：H7 growth curve,
Fig. 2 is the growth curve of bacillus cereus,
Fig. 3 is different ultrasonic intensities and treatment time to Escherichia coli O 157：The influence of H7 viable counts, curve (a)~ (e) it is the cold sterilized models fitting curves of Weibull,
Fig. 4 is the influence of different ultrasonic intensities and treatment time to bacillus cereus viable count, curve (f)~(h) For the cold sterilized models fitting curves of Weibull.
Embodiment
The present invention is illustrated with reference to embodiment.
Strain is with Gram-negative bacteria Escherichia coli O 157 in the present embodiment：H7(Escherichia coli O157:H7) Exemplified by gram-positive bacteria bacillus cereus (Bacillus cereus), meat is by taking beef as an example, and the ultrasonic wave of use is strong Degree is respectively 2.39Wcm-2、6.23Wcm-2、11.32W cm-2With 20.96W cm-2.Concrete operation step is as follows：
(1) Escherichia coli O 157：H7(Escherichia coli O157:) and bacillus cereus (Bacillus H7 Cereus) growth curve determines and connect bacterium solution preparation
- 80 DEG C of Escherichia coli O 157 will be stored in：H7(Escherichia coli O157:) and waxy gemma bar H7 Bacterium (Bacilluscereus) and 20% glycerine preserve liquid by volume 1:1000 are inoculated in LB culture mediums respectively, 37 DEG C 180rpm incubated overnights carry out rejuvenation, then take the bacterium solution of 1mL incubated overnights to be inoculated in 100mL LB culture mediums to carry out secondary work Change, take the 1mL bacterium solutions of re-activation to be inoculated in 100mL LB culture mediums, determine and once inhaled per 30min under 600nm wavelength Light value is while colony counting method determines the viable count in bacterium solution, and using growth time as abscissa, light absorption value or clump count are vertical seat The growth curve of two plants of bacterium of system is marked and drawed, as shown in Figure 1-2.From Fig. 1-2, after culture 10h, two plants of bacterium viable counts enter Plateau, in addition light absorption value be also basically unchanged, therefore using re-activation culture 10h two plants of bacteria suspensions as connecing bacterium solution.
(2) pickling liquid and sample treatment：
Prepare pickling liquid：60g/L sodium chloride, 15g/L sucrose, 4g/L composite phosphates, the pickling liquid prepared is in 10 DEG C Refrigeration is standby.Take fresh ox back eye muscle position, clean surface watery blood, remove fat and manadesma, be cut into 5 × 5 × 1cm (it is long × Width x thickness) size cube meat it is stand-by；
Ultrasonic assistant pickles processing：By the meat sample handled well and pickling liquid by 1:20 (w/v, g/mL) ratios are mixed, and are taken 5mL cultures 10h bacteria suspension by volume 1:100 are inoculated in pickling liquid, while beaker, which is placed in ice-water bath, prevents ultrasound Temperature is raised in processing procedure, and ultrasonic probe is submerged into pickling liquid from (having been surveyed at the 2cm of beef surface in above-mentioned (1) Viable count, calculates N in proportion0)。
(3) setting ultrasonic intensity is respectively 2.39Wcm-2、6.23Wcm-2、11.32W cm-2With 20.96W cm-2, it is each super Control pickling liquid temperature at 10 DEG C under the conditions of intensity of acoustic wave, take pickling liquid 5mL to carry out plate count after processing 30min, measure salts down Viable count in liquid processed.According to same sample treatment and assay method to ultrasonication under the conditions of each ultrasonic intensity 60,90 And Escherichia coli O 157 in 120min pickling liquids：H7(Escherichia coli O157:) and bacillus cereus H7 (Bacilluscereus) viable count carries out plate count, the measured value N of viable count in pickling liquid is obtained, as shown in table 1-2.No During with ultrasonic intensity and treatment time, Escherichia coli O 157：H7 and bacillus cereus viable count as shown in Figures 3 and 4, by scheming 3-4 understands that varying strength ultrasonication can significantly reduce Escherichia coli O 157：H7 viable counts, but for waxy gemma bar Bacterium, ultrasonic intensity 2.39Wcm-2When can slow down its growth rate, ultrasonic intensity be more than or equal to 6.23Wcm-2When, surpass Sonicated can reduce viable count in pickling liquid, therefore for bacillus cereus, ultrasonic intensity is 6.23Wcm-2、11.32W cm-2With 20.96W cm-2Set up model.
Escherichia coli O 157 after the different ultrasonic intensity processing of table 1:H7 measured value and calculated value
The measured value and calculated value of bacillus cereus after the different ultrasonic intensity processing of table 2
(4) N known to0, N and t, substitute into formula (1)：
Wherein：N0Micro organism quantity in pickling liquid before finger ultrasonication, t refers to ultrasonic treatment time (min), and δ, which refers to, to salt down Micro organism quantity declines the time (min) needed for a logarithm value in liquid processed, and p refers to the form parameter of curve, p>1 is that convex is bent Line, p=1 is straight line, p<1 is concave curve；
Calculating is solved equation using the progress of MATLAB softwares and obtains δ and p value.In known bacteria suspension initial concentration logN0, time During t under conditions of bacterial content number logN, programmed in MATLAB softwares as follows：
[x y]=solve (' logN0-logN1=(t1/x) ^y', ' logN0-logN2=(t2/x) ^y', ' x', ' y')
Wherein：X refers to δ, and y refers to p, can be solved and be set in the δ and p value under one pair of which treatment time, the present embodiment by above formula Four treatment times are determined, wherein each two treatment time simultaneous will solve equation, as a result as shown in table 3-4.
Escherichia coli O 157 under the different ultrasonic intensities of table 3:H7 parameters are calculated
Bacillus cereus parameter is calculated under the different ultrasonic intensities of table 4
From upper table 3 and table 4, for Escherichia coli O 157:H7, ultrasonication early stage can significantly reduce pickling liquid Middle viable count, curve is in " recessed " shape, and the time δ needed for one logarithm value of reduction is reduced with the increase of ultrasonic intensity；For Bacillus cereus, because it has thicker cell membrane, viable count declines relatively slow, song in ultrasonication early stage pickling liquid Line is assumed a " convex " shape, and the time δ needed for one logarithm value of reduction is reduced with the increase of ultrasonic intensity.
(5) will calculate obtained parameter δ andpSubstitute into formula (1) and obtain under the conditions of different ultrasonic intensities killing for two plants of bacterium Bacterium model, then ultrasonic treatment time is substituted into the viable count for calculating two plants of bacterium under the conditions of varying strength in formula (1) respectively, obtain To series of values be pickling liquid in viable count calculated value, as a result as shown in table 1-2.
(6) calculated value and measured value by more than substitute into formula (2) and calculate accurate coefficients, thus judge two plants of bacterial contents The degree of agreement of calculated value and measured value, determines the degree of accuracy of the Mathematical Modeling, while the root-mean-square error of computation model (RMSE)：
Wherein μpredictedRepresent the microorganism viable bacteria number that model is calculated；μobservedRefer to the microorganism viable bacteria number of measuring. As a result it is as shown in table 5.
The Escherichia coli O 157 of table 5:H7 and the calculated value of bacillus cereus content accurate coefficients
AfRepresent the accurate factor；RMSE represents root-mean-square error；- represent not calculate
As shown in Table 5, accurate factor values are all higher than 1 under the conditions of each ultrasonic intensity, while RMSE value is respectively less than 0.15, table When bright use formula (1) is as Mathematical Modeling, micro organism quantity in pickling liquid can be calculated when ultrasonic assistant is pickled exactly, It is closer to practical measurement result, available for the prediction of micro organism quantity in actual production and when determining that ultrasonic assistant is sterilized Between.
Handled in actual ultrasonic assistant pickles application frequently with fixed ultrasonic power, therefore the present embodiment Influence of the different ultrasonic intensity conditions to micro organism quantity is analyzed, so that more fully analysis ultrasonic wave is to microorganism killing Effect.In addition the present embodiment is pickled in production process two plants of bacterial strains with Typical Representative for beef and analyzed, can be with See that different Glan Salmonellas are different to ultrasonic wave sensitivity, bacillus is common spoilage organisms in spiced beef, rear Continuous digestion process is also more difficult to be killed, therefore it is considered that ultrasonic intensity should be not less than 6.23Wcm in actual applications-2, it is right Micro organism quantity can play significant killing effect., also can be before pickling to pickling in order to reduce microorganism pollution in pickling liquid Liquid is pre-processed.

Claims (5)

  1. When 1. the prediction ultrasonic assistant based on Mathematical Modeling is pickled in pickling liquid content of microorganisms method, it is characterised in that Comprise the following steps：
    1), green meat is cleaned up and removed after fat, manadesma, in immersion pickling liquid, determines to be measured using colony counting method The initial number N of microorganism0；
    2) processing t minutes, is carried out to pickling liquid using ultrasonic wave；
    3), by N0The quantity N that calculates ultrasonication t minute after tested microorganism is substituted into formula (1) with t：
    log N = log N 0 - ( t &delta; ) p - - - ( 1 )
    Wherein：N0Micro organism quantity in pickling liquid before finger ultrasonication, t refers to ultrasonic treatment time (min), and δ refers at ultrasonic wave Micro organism quantity declines the time (min) needed for a logarithm value in the lower pickling liquid of reason, and p refers to the form parameter of curve, p>1 is convex Type curve, p=1 is straight line, p<1 is concave curve.
  2. Content of microorganisms in pickling liquid when 2. the prediction ultrasonic assistant according to claim 1 based on Mathematical Modeling is pickled Method, it is characterised in that δ and p measuring and calculating step is in the formula (1)：
    1) tested microorganism sample, is taken, its growth curve is determined and determines optimal incubation time；
    2) viable bacteria, is prepared under the conditions of optimal incubation time and connects bacterium solution, its initial microbial population N is determined0’。
    3), fresh beef is cleaned up and removed after fat, manadesma, in immersion pickling liquid, the bacterium solution that connects prepared is added In pickling liquid, handled using ultrasonic wave；
    4) when, colony counting method calculates different ultrasonic intensities, difference ultrasonic treatment time t ' in pickling liquid two plants of bacterium work Bacterium number N '；
    5), by N0', t ' and N ' substitute into formula (1), carry out solving equation calculating obtaining under ultrasonic intensity using MATLAB softwares δ and p value, in MATLAB softwares program it is as follows：
    [x y]=solve (' logN0’-logN1'=(t1'/x) ^y', ' logN0’-logN2'=(t2/ x) ^y', ' x', ' y'),
    Wherein：X refers to δ, and y refers to p.
  3. Content of microorganisms in pickling liquid when 3. the prediction ultrasonic assistant according to claim 1 based on Mathematical Modeling is pickled Method, it is characterised in that the step 1) in pickling liquid include 60g/L sodium chloride, 15g/L sucrose, 4g/L Composite phosphate.
  4. Content of microorganisms in pickling liquid when 4. the prediction ultrasonic assistant according to claim 1 based on Mathematical Modeling is pickled Method, it is characterised in that the step 2) in using ultrasonication when ultrasonic probe is submerged in pickling liquid from meat table At the 2cm of face.
  5. Content of microorganisms in pickling liquid when 5. the prediction ultrasonic assistant according to claim 1 based on Mathematical Modeling is pickled Method, it is characterised in that the step 2) in ultrasonication intensity be more than or equal to 6.23Wcm-2。
CN201710173823.8A 2017-03-22 2017-03-22 When prediction ultrasonic assistant based on Mathematical Modeling is pickled in pickling liquid content of microorganisms method Pending CN106957889A (en)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107960607A (en) * 2017-12-11 2018-04-27 徐州汉戌堂食品有限公司 A kind of ultrasonic wave meat curing method
CN112488561A (en) * 2020-12-10 2021-03-12 广州市来利洪饼业有限公司 Moon cake safety quality guarantee method and system based on short-time heat drying and ultraviolet rays

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CN105277611A (en) * 2015-09-17 2016-01-27 南京农业大学 Model for rapid determination of salt content of ultrasonic assisted pickled beef and application

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CN105277611A (en) * 2015-09-17 2016-01-27 南京农业大学 Model for rapid determination of salt content of ultrasonic assisted pickled beef and application

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107960607A (en) * 2017-12-11 2018-04-27 徐州汉戌堂食品有限公司 A kind of ultrasonic wave meat curing method
CN112488561A (en) * 2020-12-10 2021-03-12 广州市来利洪饼业有限公司 Moon cake safety quality guarantee method and system based on short-time heat drying and ultraviolet rays
CN112488561B (en) * 2020-12-10 2024-05-14 广州市来利洪饼业有限公司 Moon cake safety quality guaranteeing method and system based on short-time hot baking and ultraviolet rays

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Application publication date: 20170718