CN108872293A - A kind of metabonomic analysis of Alveolar echinococcosis and the construction method of preliminary screening model - Google Patents
A kind of metabonomic analysis of Alveolar echinococcosis and the construction method of preliminary screening model Download PDFInfo
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Abstract
The invention discloses a kind of metabonomic analysis of Alveolar echinococcosis and the construction methods of preliminary screening model, including：Collect Healthy People and Alveolar echinococcosis patients serum and urine sample；The one-dimensional nuclear magnetic resonance hydrogen spectrum for acquiring each sample obtains the metabolic profile spectrum of sample；Metabolic profile is composed and carries out data prediction, obtains two-dimensional data matrix；Two-dimensional data matrix is subjected to orthogonal partial least squares discriminant analysis, obtains OPLS-DA model；Model is verified, if model does not have over-fitting, gained OPLS-DA model is the metabonomic analysis and preliminary screening model of Alveolar echinococcosis.Metabonomic technology is applied in the analysis and preliminary screening of Alveolar echinococcosis by the present invention for the first time, it can noninvasive, quickly and conveniently filter out Alveolar echinococcosis people at highest risk, effectively improve the screening efficiency of the full crowd in Alveolar echinococcosis district occurred frequently, screening cost is substantially reduced, convenient for promoting the use of.
Description
Technical field
The invention belongs to analysis technical fields, and in particular to a kind of metabonomic analysis and preliminary sieve of Alveolar echinococcosis
The construction method for looking into model can be convenient using the model that construction method of the invention obtains, quickly to Alveolar echinococcosis
Carry out preliminary screening.
Background technique
Alveolar echinococcosis (echinococcosis multilocularis) is the larva alveolitoid by Echinococcus multilocularis
A kind of chronic liver parasitic disease of infecting both domestic animals and human caused by echinococcus (Echinococcus multilocularis) infected liver.Alveolar echinococcosis protopathy
Positioned at liver, growth pattern can be sent out by approach such as blood fortune to devices such as lung, brains similar to the same infiltration type proliferation growth of tumour
Official generates secondary or metastatic lesion, is clinically known as the title of " worm cancer " or " parasitic liver cancer ".The disease disease incidence is low, but
It is that will cause fatefulue injury if treatment is not in time.In China, which is distributed mainly on the Inner Mongol, Tibet, Xinjiang, Qinghai
Equal northwests farming and pastoral area.
Surgical resection therapy is usually taken in Alveolar echinococcosis, but disease early stage is substantially asymptomatic, and incubation period is
20 years, general patient was difficult to find by itself, when symptom medical treatment occur in Most patients, missed often in middle and advanced stage
Golden hour, cannot perform the operation excision, even if carrying out, liver divides or the excision of half leaf, Postoperative recurrent rate are also very high.Clinically answer
It is albendazole (mebendazole) with more drug, continuous treatment for a long time is needed, but report according to foreign countries, even if treatment phase
Limit for length checked, recurrence rate 37% for 1~6 year after the treatment up to 43 years, and effect is dissatisfied.Therefore, it aims at prevention, prevention and treatment knot
The comprehensive preventive health measures of conjunction is particularly critical.Carry out Mass screening in Alveolar echinococcosis morbidity Prevalent district, is treatment liver alveolitoid
The most effective shortcut of echinococcosis, passes through Mass screening, it is possible to find patient suspected makes a definite diagnosis patient, realizes that Alveolar echinococcosis is high-incidence
The area masses of farmers and herdsmen early have found, early diagnose, early treatment, reduce the death rate of Alveolar echinococcosis patient, mitigate patient's early stage inspection
Look into and treat the financial burden of Alveolar echinococcosis.
Metabolism group is the emerging technology research field that rear era gene occurs, and is biological endogenous about quantitative description
Property metabolite entirety and its to internal cause and external cause variation response rule science.It utilizes high-throughput, highly sensitive and height
The modern analytical technique of accuracy carries out dynamically track analysis to the small molecule metabolites of vital movement terminal, and by polynary
The information processing means such as statistical analysis technique, mode identification technology, from molecular level body in disease etc. it is extraneous because
The response pattern of element interference, can be used for disclosing the pathomechanism of disease or the early screening of disease.
High resolution nuclear magnetic resonance wave spectrum (1H NMR) it is one of most common analytical technology in metabolism group, with chromatography-
Mass spectrometric hyphenated technique is compared, and major advantage includes：The structure and property of sample are not destroyed, it can not only be to intact tissue, life
Object liquid carries out Metabolite analysis, and can carry out the Methanogenesis in situ of living body；To the nothing of all compounds
Skewed popularity measurement, spectrum peak position and intensity can be qualitative and quantitative, convenient for the quantitative analysis of metabolite；Can study Chemical Exchange,
The dynamic processes such as diffusion and internal motion, provide multidate information extremely abundant；A variety of edits, experimental method can be designed
It is versatile and flexible, meet the purpose that metabolism group detects compound as much as possible.Therefore, based on the generation of nuclear magnetic resonance
It thanks to a group method and has attracted the numerous studies in the fields such as basic life science and biomedicine, and had been widely used toxicity
The research in the fields such as relationship, the disorder in screening of research, food and health.But it is based on1The generation of the Alveolar echinococcosis of H NMR technology
It thanks to group credit analysis and tentatively at home and abroad there is not been reported so far for screening model.
Summary of the invention
In place of making up the deficiencies in the prior art, a kind of metabolism group of Alveolar echinococcosis is provided
The construction method of credit analysis and preliminary screening model, method is simple, is caused using metabonomic technology detection Alveolar echinococcosis
Organism metabolism production concentration variation, provide serum and urine sample metabonomic analysis model, can before imageological examination first
Alveolar echinococcosis Risk Population is carried out early stage and preliminary screening, it is economical and practical, convenient and efficient, it is easy to promote and utilize.
The technical solution adopted by the present invention to solve the technical problems is：
A kind of metabonomic analysis of Alveolar echinococcosis and the construction method of preliminary screening model, including：
1) Healthy People and Alveolar echinococcosis patients serum and urine sample are collected respectively, as analysis sample；
2) using nuclear magnetic resonance spectrometer detection each analysis sample one-dimensional nuclear magnetic resonance hydrogen spectrum (1H-NMR), health is obtained
The metabolic profile of people and Alveolar echinococcosis patients serum and urine spectrum；
3) metabolic profile of Healthy People and Alveolar echinococcosis patients serum and urine spectrum is subjected to data prediction respectively,
Every behavioural analysis sample is obtained, the two-dimensional data matrix of metabolite profile peak information is often classified as, for further statistical modeling point
Analysis；
4) two-dimensional data matrix of step 3) arrived is subjected to orthogonal partial least squares discriminant analysis (OPLS-DA), obtained
OPLS-DA model simultaneously verifies model, if model does not have over-fitting, gained OPLS-DA model is liver
The metabonomic analysis of alveolar echinococcosis and preliminary screening model.
In one embodiment：In the step 2), metabolic profile spectrum acquisition is in the Bruker for being equipped with CryoProbeTM
It is carried out on 600MHz nuclear magnetic resonance spectrometer, temperature is room temperature (298K)；Wherein, blood serum sample uses NOSESPRR pulse train simultaneously
Increase a CPMG spin echo segment on this basis to be detected, inhibits the shadow of the macromoleculars signals such as Proteins in Serum
It rings；Urine sample is detected using NOESYPR 1D pulse train；Acquire Healthy People and Alveolar echinococcosis patients serum and
The high-resolution one-dimensional nuclear magnetic resonance hydrogen of urine is composed, and the one-dimensional nuclear magnetic resonance hydrogen modal data of the serum collected and urine uses
V.8.1.2 software carries out Fourier transformation, phasing, baseline correction and calibration to MestReNova, obtains sample1H NMR
Metabolic profile spectrum.
In one embodiment：In the step 3), the data prediction includes：A) spectral peak alignment is carried out, to reduce spectral peak
The influence of drift；B) remaining water peak and b extent are removed, and subsection integral is carried out to modal data by spectral peak, reduces the dimension of data
Number；C) using PQN method carry out data normalization (Dieterle, F., Ross, A., Schlotterbeck, G., Senn,
H.Probabilistic quotient normalization as robust method to account for
dilution of complex biological mixtures.Application in 1H NMR
Metabonomics.Anal Chem, 2006,78,4281-4290), reduce the influence of dilution effect；Finally obtain every behavior point
Sample is analysed, the two-dimensional data matrix of metabolite profile peak information is often classified as；Spectral peak is carried out 2-D data square after metabolin ownership
Battle array imports 14.0 software of SIMCA and carries out orthogonal partial least squares discriminant analysis (OPLS-DA) described in step 4) and model.
In one embodiment：The serum and urine sample of the Alveolar echinococcosis patient is selected from：In conjunction with medical history, clinical table
Existing, imageological examination etc. is diagnosed as the patient of Alveolar echinococcosis, excludes complication with diabetes, nephrosis, autoimmunity disease etc. to urine
The influential patient of metabolite in liquid, and exclude be associated with malignant tumor of liver and organ transplant it is postoperative, fever, serious liver kidney function
Can not congruent metabolic disorder patient；The serum and urine sample of the Healthy People voluntarily participate in same period family numbers of patients
The Healthy People of research.
In one embodiment：The serum and urine sample of the Healthy People and the serum and urine of Alveolar echinococcosis patient
Liquid sample is all from 30~60 years old crowd.
In one embodiment：The blood and urine sample of the health and Alveolar echinococcosis patient are collected in the morning 8:00
~12:Between 00, acquire on an empty stomach.
In one embodiment：The serum and urine sample are through centrifuging and taking supernatant, after liquid nitrogen flash freezer, it is stored in -85~-
75 DEG C to be measured, and the holding time is no more than 6 months.
In one embodiment：The urine sample is derived from 15~25mL of urina sanguinis middle section, at 0~5 DEG C 8000~12000rpm from
8~12min of the heart, takes supernatant, after liquid nitrogen flash freezer, is placed in -85~-75 DEG C and saves for use；The blood serum sample take whole blood 1~
3mL is placed in the EP pipe that anti-coagulants is not added, and 6000~12000rpm is centrifuged 5~12min at 0~5 DEG C, supernatant is taken, through liquid nitrogen
After quick-frozen, it is placed in -85~-75 DEG C and saves for use.
In one embodiment：By serum and urine sample after -85~-75 DEG C of taking-ups defrostings, according to 1.5~2.5:1 volume ratio
Take serum be added to heavy water configure 85~95mM, pH 7.3~7.5 phosphate buffer, be centrifuged, take after mixing
Clear liquid is fitted into nuclear magnetic tube, to be measured in 0~5 DEG C of preservation, as analysis sample；According to 0.8~1.2:1 volume ratio takes urine sample
1.4~1.6M, the pH 7.3~7.5 configured to heavy water is added in product, containing 0.4~0.6mM TSP as interior target phosphate-buffered
Liquid is centrifuged after mixing, and supernatant is taken to be fitted into nuclear magnetic tube, to be measured in 0~5 DEG C of preservation, as analysis sample.
In one embodiment：In the shot chart of serum and the OPLS-DA model of urine sample, Alveolar echinococcosis patient and Healthy People
There is obvious differentiation, the R of serum OPLS-DA model between sample sets2Y is 80% or more, Q2It is 65% or more；Urine sample OPLS-DA
The R of model2For 90% or more, Q2It is 70% or more.
In above-mentioned serum and the metabonomic analysis model of urine sample, Healthy Human Serum, urine sample and liver alveolitoid Echinococcus hydatid cyst
Patient's serum, urine sample metabolin information be located at the different location of model, classification trend is obvious, remembers Healthy People blood
Clearly, urine sample group be located region be negative areas, Alveolar echinococcosis patients serum, urine sample group the region that is located be
Positive region.
Using the metabonomic analysis model of above-mentioned serum and urine sample, it can be used for Healthy People and Alveolar echinococcosis patient
The metabonomic analysis of serum and urine sample.
Using a kind of metabonomic analysis and screening model of Alveolar echinococcosis of the present invention, including following step
Suddenly：It acquires serum or urine sample to be checked to be pre-processed, reaches nuclear magnetic resonance experiment requirement；By pretreated serum to be checked
Or urine sample carries out nuclear magnetic resonance spectroscopy, obtains the original metabolic profile spectrum of serum or urine sample to be checked；By these originals
Beginning metabolic profile spectrum is pre-processed, and the two-dimensional data matrix comprising metabolin information is obtained；This data matrix is imported changeable
It measures in analysis model, for differentiating the classification of serum or urine sample to be checked.In practical applications, if serum to be checked or urine
Sample is seated that the Alveolar echinococcosis patients serum of model, (i.e. positive region, i.e. prediction probability are greater than in urine sample group region
50%), then it represents that risk of the individual with Alveolar echinococcosis is high, needs to carry out further clinical examination；If to be checked
Serum and urine sample are seated that the Healthy Human Serum of model, (i.e. negative areas, i.e. prediction probability are less than in urine sample group region
50%), then it represents that risk of the individual with Alveolar echinococcosis is low.
Compared with the background art, it has the following advantages that the technical program：
It is the screening of Alveolar echinococcosis that the present invention utilizes the metabolism group method based on NMR spectrum technology for the first time
Technical support is provided.The present invention is based on the metabolism group methods of NMR spectrum technology to suffer from Healthy People and Alveolar echinococcosis
Person's is detected to obtain corresponding metabolic profile map with urine sample, and constructs OPLS-DA model, these models can distinguish
The metabolin situation of serum and urine sample, and can tentatively be judged whether by the metabolic profile of serum and urine sample with liver alveolitoid packet
Parasitosis illness is dangerous.Full crowd's early screening can be carried out to the high region of disease of Alveolar echinococcosis using the method for the present invention, screened out
There is no illness dangerous or the crowd of low-risk, then carry out other further clinical medicine inspections for high-risk individual, passes through
Whether further other clinical medicine inspections diagnosis suffers from Alveolar echinococcosis, and the range of clinical examination is greatly reduced, has
There is important economic and social benefit.
Biological sample of the invention is Healthy People and the serum and urine sample of Alveolar echinococcosis patient, serum and urine sample
It collects and treatment process is simple and convenient, cost is low, belongs to Non-invaive examination, effectively increases Alveolar echinococcosis Risk Population
The efficiency of screening, reduces screening cost, has important economic and social benefit, easy to promote and utilize.In addition, by being based on
The metabolism group method of NMR spectrum technology can be quickly obtained liver alveolitoid without serum and urine sample is intrusively detected
Echinococcosis causes the metabolic profile of variation, reflects global information, solves the organ or vesica of Alveolar echinococcosis patient
The problem being difficult to obtain has biggish meaning to the preliminary screening of Alveolar echinococcosis.
Detailed description of the invention
Present invention will be further explained below with reference to the attached drawings and examples.
Fig. 1 is Healthy People and the representative one-dimensional nuclear magnetic resonance spectrogram of Alveolar echinococcosis patient.(a) serum；(b) it urinates
Sample.
The O-PLSDA shot chart (left side) and arrangement accordingly that Fig. 2 is Healthy People and Alveolar echinococcosis patient's spectral data
Permutation test figure (n=200) (right side).(on) serum；(under) urine sample.Wherein Control group is that screening is negative, indicates health
The serum or urine sample of people, HAE group are that screening is positive, indicate the serum or urine sample of Alveolar echinococcosis patient.
Fig. 3 is the ROC curve of screening model.(a) serum；(b) urine sample.
Fig. 4 is the difference metabolin general view that enhanced volcano figure filters out.(a) serum；(b) urine sample.
Specific embodiment
The contents of the present invention are illustrated below by embodiment：
1. research object
The present invention select in July, 2016 to September in affiliated hospital of Qinghai University liver and gallbladder pancreas surgery through clinical diagnosis (in conjunction with disease
History, clinical manifestation, imageological examination result) it is studied for 30~60 years old Tibetan patient, 18 people of Alveolar echinococcosis, wherein
Complication with diabetes, nephrosis, autoimmunity disease etc. is excluded to have an impact to metabolite in urine, be associated with malignant tumor of liver and organ
The patient of the metabolic disorders such as post-transplantation, fever, serious hepatic and kidney function obstacle voluntarily participates in the 30 of research in same period family numbers of patients
~60 years old 18 people of Healthy People are as control.
2. reagent and instrument and equipment
Experiment reagent includes heavy water, chloroform, methanol, sodium dihydrogen phosphate, dipotassium hydrogen phosphate.
Testing consumptive material includes 5mm nuclear magnetic tube, pipettor, 1000 μ L and 200 μ L pipette tips, 5mL EP pipe, marking pen, latex hand
Set, mask.
Experimental facilities includes pipettor, low-temperature and high-speed centrifuge, Brooker 600MHz nuclear magnetic resonance spectrometer.
3. the collection and pretreatment of sample
The collection of the blood and urine of patient and its family members is in the morning 8:00~12:It is completed in 00.It is required when acquisition empty
Abdomen.It takes cleaning middle section urina sanguinis 20mL or so to be placed in when A urine sample collection device to indicate in the urine cup of information in advance, and centrifugation (10000rpm, 4
DEG C, 10min), it then pipettes 1mL urine using pipettor and is placed in corresponding 5mL EP pipe, when liquid relief avoids bacterium infection simultaneously
Whole process is completed on ice.Whole blood about 1mL is taken to be placed in the EP pipe that anti-coagulants is not added, after centrifugation (10000rpm, 4 DEG C, 10min),
Supernatant is pipetted with pipettor be placed in corresponding 5mL EP pipe obtain blood serum sample.All serum and urine sample liquid nitrogen speed
After jelly, it is placed in -80 DEG C of low temperature refrigerators and saves.
Serum and urine sample are taken out after thawing, takes the serum of 400 μ L that the phosphate buffer of 200 μ L heavy water configuration is added
(90mM, pH 7.4) is centrifuged (10000rpm, 4 DEG C, 10min) after mixing, the supernatant liquor of 550 μ L is pipetted with pipettor
Be fitted into 5mm nuclear magnetic tube, be stored in it is to be measured in 4 DEG C of refrigerators, as analysis sample；Take the urine sample of 300 μ L that 300 μ L weight is added
The phosphate buffer (1.5M, pH 7.4, TSP containing 0.5mM is as internal standard) of water configuration, is centrifuged after mixing
(10000rpm, 4 DEG C, 10min), is fitted into 5mm nuclear magnetic tube with the supernatant liquor that pipettor pipettes 550 μ L, is stored in 4 DEG C of refrigerators
In it is to be measured, as analysis sample.
4. nuclear magnetic resonance spectroscopy
Nuclear magnetic resonance experiment is on the nuclear magnetic resonance spectrometer of Brooker (Bruker) 600MHz equipped with CryoProbeTM
It completes, experimental temperature is set as 298K, and blood serum sample uses NOSESPRR pulse train and increases a CPMG on this basis
Spin echo segment is detected, and the influence of the macromoleculars signals such as Proteins in Serum is inhibited；Urine sample uses NOESYPR
1D pulse train is detected；The one-dimensional nuclear-magnetism of high-resolution for acquiring Healthy People and Alveolar echinococcosis patients serum and urine is total
Vibration wave spectrum (1H-NMR)。
5. the pretreatment of nmr spectrum
V.8.1.2 software carries out using MestReNova for the serum collected and the one-dimensional nuclear magnetic resonance wave spectrum of urine
The pretreatments such as Fourier transformation, phasing, baseline correction, spectral peak calibration, remaining water peak removal, to treated, spectrogram is carried out
Integral and PQN obtain every behavioural analysis sample after composing normalization entirely, are often classified as the two-dimensional data matrix of metabolin information, spectral peak
The documents and materials that metabolin ownership refers to the database of KEGG, HMDB and delivered, the metabolin information belonged to out are shown in Fig. 1.
6. multivariable and univariate statistics analysis
Two-dimensional data matrix is imported into 14.0 software of SIMCA and carries out orthogonal partial least squares discriminant analysis (OPLS-DA).
Using the data Standard graduation conversion mode of adaptive conversion.OPLS-DA analyzes result as shown in Fig. 2, the screening positive (i.e. liver alveolitoid Echinococcus hydatid cyst
Patient's serum or urine sample) and screening negative (Healthy Human Serum or urine sample) between there is apparent classification trend, secretly
Show that Alveolar echinococcosis causes patient's body metabolic patterns to change.The present invention examines the matter to model using permutation
Amount and validity do further inspection.From permutation inspection result Fig. 2 of model as it can be seen that serum and urine sample OPLS-DA model
Q2It is higher, illustrate that the predictive ability of model is preferable.
7. the OPLS-DA model of Alveolar echinococcosis early screening
The present invention does further inspection to the predictive ability of model respectively using Monte Carlo Cross-Validation.In Monte Carlo
In cross validation, 13 samples (about the 70% of gross sample) is selected at random every time as training set and establishes OPLS-DA model, is remained
Test set of lower 5 samples (about the 20% of gross sample) as model, Healthy People and Alveolar echinococcosis patients serum or urine
The training set of liquid sample composition shows apparent separation, and test set is introduced into model, and experiment repetition (200) is secondary, as a result such as
Shown in table 1.Table 1 shows that the judgment accuracy of serum and urine sample model is up to 100% and 97.2% respectively, illustrates in the present invention
The metabonomic analysis model of Alveolar echinococcosis patients serum and urine sample has differentiates predictive ability well.
The wrong classification chart of table 1.OPLS-DA model
8. the Difference of Metabolism of Alveolar echinococcosis and healthy population is analyzed
Multivariable and univariate statistics analysis result are visualized using enhanced volcano figure, synthesization expression, is sought
Look for characteristic metabolic object relevant to Alveolar echinococcosis.Each dot represents a kind of metabolin, the size and face of dot in the figure
Color respectively represents from O-PLSDA model derived variable importance scoring (VIP) and related coefficient absolute value (| r |), X-axis table
Show Alveolar echinococcosis patient for the log of certain metabolin relative concentration folding value of normal person2, Y-axis expression liver alveolitoid Echinococcus hydatid cyst
Patient and normal person carry out the-log10 of the p value obtained after t inspection, and the screening criteria of difference metabolin is：VIP>top
30%, | r |>0.5 ,-log10(p value)>2 (i.e. p<And log 0.01)2(folding value)>0.25, therefore in the figure of volcano, difference
Metabolin is mainly reflected in be located at the warm colour large circle point on upper left side and upper right side region, as shown in Figure 4.
The difference metabolin list of 2. Alveolar echinococcosis of table and healthy population
aCONC：The relative concentration values of each metabolin indicate that wherein mean is that the integral of every kind of metabolin is flat with mean ± SE
Mean value, SE are standard errors.
bp-value：T examines the significance,statistical obtained horizontal.
cr：Related coefficient, positive number (or negative) indicate that metabolite concentration is positively correlated between comparative group (or negatively correlated).
dVIP：Variable importance scoring.
9. conclusion
A kind of metabonomic analysis and screening mould of Alveolar echinococcosis of the present invention it can be seen from verifying above
Type, constructed OPLS-DA model have good prediction effect, and no over-fitting is dangerous, and classify to become between having apparent group
Gesture, can be very well by Healthy People or the serum and urine of doubtful Alveolar echinococcosis patient by serum and the metabolic condition of urine sample
Sample distinguishes, and can be used for Alveolar echinococcosis early screening.
Using a kind of metabonomic analysis and screening model of Alveolar echinococcosis of the present invention, including following step
Suddenly：
(1) serum or urine sample to be checked are acquired according to above-mentioned steps 3 to be pre-processed, reaches nuclear magnetic resonance experiment and wants
It asks；
(2) pretreated serum to be checked or urine sample are subjected to nuclear magnetic resonance spectroscopy according to above-mentioned steps 4, obtained
The original metabolic profile map of serum or urine sample to be checked；
(3) these original metabolic profile maps are subjected to spectrogram pretreatment according to above-mentioned steps 5, obtain believing comprising metabolin
The two-dimensional data matrix of breath；
(4) this data matrix is imported in serum and the metabonomic analysis model of urine sample according to above-mentioned steps 6, for dividing
Analyse the metabolic condition of serum or urine sample to be checked.In practical applications, if serum to be checked or urine sample are seated model
Alveolar echinococcosis patients serum, urine sample group region (i.e. positive region, i.e. prediction probability be greater than 50%), then it represents that suffer from
There is the risk of Alveolar echinococcosis higher, needs to carry out further clinical examination；If serum to be checked or urine sample are located
In the Healthy Human Serum of model, urine sample group region, (i.e. negative areas, i.e., prediction probability is less than 50%), then it represents that suffers from liver
The risk of alveolar echinococcosis is smaller.
The above is only the preferred embodiment of the present invention, the range implemented of the present invention that therefore, it cannot be limited according to, i.e., according to
Equivalent changes and modifications made by the invention patent range and description, should still be within the scope of the present invention.
Claims (10)
1. the construction method of a kind of metabonomic analysis of Alveolar echinococcosis and preliminary screening model, it is characterised in that：Including：
1) Healthy People and Alveolar echinococcosis patients serum and urine sample are collected respectively, as analysis sample；
2) using the one-dimensional nuclear magnetic resonance hydrogen spectrum of nuclear magnetic resonance spectrometer detection each analysis sample, Healthy People and liver alveolitoid packet are obtained
The metabolic profile of parasitosis patients serum and urine spectrum；
3) metabolic profile of Healthy People and Alveolar echinococcosis patients serum and urine spectrum is subjected to data prediction respectively, is obtained
Every behavioural analysis sample, is often classified as the two-dimensional data matrix of metabolite profile peak information, analyzes for further statistical modeling；
4) two-dimensional data matrix of step 3) arrived is subjected to orthogonal partial least squares discriminant analysis, obtains OPLS-DA model simultaneously
Model is verified, if model does not have over-fitting, gained OPLS-DA model is the generation of Alveolar echinococcosis
Thank group credit analysis and a preliminary screening model.
2. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：In the step 2), metabolic profile spectrum acquisition carries out on Bruker 600MHz nuclear magnetic resonance spectrometer, temperature
For room temperature；Wherein, blood serum sample uses NOSESPRR pulse train and increases a CPMG spin echo segment on this basis
It is detected；Urine sample is detected using NOESYPR 1D pulse train；Acquire Healthy People and Alveolar echinococcosis patient
The one-dimensional nuclear magnetic resonance hydrogen of serum and the high-resolution one-dimensional nuclear magnetic resonance hydrogen of urine spectrum, the serum collected and urine composes number
According to Fourier transformation, phasing, baseline correction and calibration is carried out, sample is obtained1H NMR metabolic profile spectrum.
3. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：In the step 3), the data prediction includes：A) spectral peak alignment is carried out, to reduce the shadow of spectral peak drift
It rings；B) remaining water peak and b extent are removed, and subsection integral is carried out to modal data by spectral peak, reduces the dimension of data；C) sharp
Data normalization is carried out with PQN method, reduces the influence of dilution effect；Every behavioural analysis sample is finally obtained, metabolism is often classified as
The two-dimensional data matrix of object spectral peak information；It will be described in two-dimensional data matrix progress step 4) after carrying out metabolin ownership to spectral peak
Orthogonal partial least squares discriminant analysis simultaneously models.
4. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：The serum and urine sample of the Alveolar echinococcosis patient is selected from：In conjunction with medical history, clinical manifestation, image
It learns and checks etc. and to be diagnosed as the patient of Alveolar echinococcosis, exclude complication with diabetes, nephrosis, autoimmunity disease etc. to being metabolized in urine
The influential patient of product, and exclude to be associated with malignant tumor of liver and organ transplant is postoperative, fever, serious hepatic and kidney function obstacle etc.
The patient of metabolic disorder；The serum and urine sample of the Healthy People voluntarily participate in the strong of research in same period family numbers of patients
Health people.
5. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：The serum and urine sample of the Healthy People and the serum and urine sample of Alveolar echinococcosis patient are equal
From 30~60 years old crowd.
6. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：The blood and urine sample of the health and Alveolar echinococcosis patient are collected in the morning 8:00~12:00
Between, it acquires on an empty stomach.
7. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：The serum and urine sample are through centrifuging and taking supernatant, after liquid nitrogen flash freezer, be stored in -85~-75 DEG C to
It surveys, the holding time is no more than 6 months.
8. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：The urine sample is derived from 15~25mL of urina sanguinis middle section, at 0~5 DEG C 8000~12000rpm centrifugation 8~
12min takes supernatant, after liquid nitrogen flash freezer, is placed in -85~-75 DEG C and saves for use；The blood serum sample takes 1~3mL of whole blood to set
In the EP pipe that anti-coagulants is not added, 6000~12000rpm is centrifuged 5~12min at 0~5 DEG C, supernatant is taken, through liquid nitrogen flash freezer
Afterwards, -85~-75 DEG C are placed in save for use.
9. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：By serum and urine sample after -85~-75 DEG C of taking-ups defrostings, blood serum sample is according to 1.5~2.5:1 volume ratio
Blood sampling be added to heavy water configure 85~95mM, pH 7.3~7.5 phosphate buffer, be centrifuged, take after mixing
Clear liquid is fitted into nuclear magnetic tube, to be measured in 0~5 DEG C of preservation, as analysis sample；Urine sample is according to 0.8~1.2:1 volume ratio
Take urine sample that 1.4~1.6M, the pH 7.3~7.5 configured to heavy water is added, containing 0.4~0.6mM TSP as interior target phosphate
Buffer is centrifuged after mixing, and supernatant is taken to be fitted into nuclear magnetic tube, to be measured in 0~5 DEG C of preservation, as analysis sample.
10. the construction method of the metabonomic analysis of Alveolar echinococcosis according to claim 1 and preliminary screening model,
It is characterized in that：In the shot chart of serum and the OPLS-DA model of urine sample, Alveolar echinococcosis patient and Healthy People sample sets it
Between exist it is obvious distinguish, the R of serum OPLS-DA model2Y is 80% or more, Q2It is 65% or more；The R of urine sample OPLS-DA model2
For 90% or more, Q2It is 70% or more.
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