CN116646023B - Method for evaluating similarity between sample and breast milk in multiple dimensions - Google Patents
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- G16C—COMPUTATIONAL CHEMISTRY; CHEMOINFORMATICS; COMPUTATIONAL MATERIALS SCIENCE
- G16C20/00—Chemoinformatics, i.e. ICT specially adapted for the handling of physicochemical or structural data of chemical particles, elements, compounds or mixtures
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Abstract
The invention relates to the technical field of dairy product evaluation, and discloses a method for evaluating similarity between a sample and breast milk in multiple dimensions. The method comprises the following steps: (1) establishing a local breast milk ganglioside database; (2) collecting a sample to be evaluated and performing pretreatment; (3) Measuring the content of characteristic ganglioside subclasses and the content of characteristic ganglioside molecules in the pretreated sample to be evaluated, and extracting peak areas, basal peak heights and peak numbers from a total ion flow diagram; (4) calculating the score G of the sample to be evaluated according to a formula. The invention starts from three aspects of characteristic ganglioside subclass composition, characteristic ganglioside molecular composition and overall distribution condition evaluation, and the similarity of infant formula milk powder, raw auxiliary materials and breast milk substitution fat and breast milk ganglioside is comprehensively evaluated in a multi-dimensional way, so that the simulation quality of the infant formula milk powder and the raw auxiliary materials thereof on the breast milk ganglioside can be better evaluated.
Description
Technical Field
The invention relates to the technical field of dairy product evaluation, in particular to a method for evaluating similarity between a sample and breast milk in a multi-dimensional manner.
Background
Lipids are one of the important nutrients required by the human body and also one of the important components of breast milk. For infants 45-55% of the daily energy required is provided by lipids. Studies have shown that the composition of the milk fat affects infant growth. Thus, breast milk is considered a gold standard for infant formula development.
Gangliosides are acidic glycosphingolipids, which consist of three parts, namely sphingosine, fatty acid and sialic acid-containing sugar chains, and are widely distributed in tissues and body fluids of the human body, wherein the distribution in the nervous system is most abundant. Breast milk is the main source of gangliosides, the most predominant ganglioside species being bissialylglycoside GD3 (neu5acα2-8neu5acα2-3galβ1-4Glc βcer) and monosialylglycoside GM3 (neu5acα2-3galβ1-4Glc βcer). Gangliosides in breast milk are mainly positioned on the outer surface of the milk fat globule membrane, which is helpful for the formation and stabilization of milk fat globules. Gangliosides are one of key nutrients for early growth and development of infant brain, play an important role in neural development, memory formation and synaptic signal transduction, can adsorb and take away pathogens in infant intestinal tracts, exert the effect of directly resisting pathogen adhesion, participate in regulating immune system and supporting the maturation of infant intestinal tracts, and promote bifidobacterium proliferation in infant intestinal tracts. The composition and content of ganglioside in breast milk are affected by lactation period, living habit, region, etc., while the composition and content of ganglioside in infant formula milk powder are mainly affected by raw and auxiliary materials and processing technology.
Throughout the history of infant formula lipid development, a tighter match was achieved mainly around the use of vegetable oils, the simulation of structural lipids and the supplementation of functional fatty acids, i.e. in terms of composition, content and structure of glyceric acid, when the mother milk lipid was simulated. With the verification of the important efficacy value of milk gangliosides, the simulation of milk gangliosides is increasingly emphasized. However, methods for assessing ganglioside analog similarity in infant formulas have been reported only rarely. In China, most infant formulas are based on fresh milk or milk powder, while the lipid fraction is made up of milk or milk powder in which fat, vegetable oil and milk substitute fat combine to simulate milk fat. The raw materials and the auxiliary materials used for processing the formula powder directly determine the composition and the content of gangliosides in the formula powder, however, the similarity evaluation method for the raw materials and the auxiliary materials and the breast milk is not reported.
At present, most of the evaluation of infant formula powder and raw and auxiliary material lipid thereof adopts chromatographic instruments, mass spectrometry instruments and the like to detect data, and comparison analysis is carried out, so that the difference between infant formula powder, raw and auxiliary materials and breast milk substituted lipid and mother milk lipid is illustrated. The prior evaluation technology is mainly used for evaluation and comparison at the level of fatty acid or lipid subclass or at the level of certain ganglioside, the influence of the subclass and molecular composition, content and overall distribution of ganglioside on infant formula powder is not considered, and the weight problem of gangliosides of different types is not considered.
In view of this, the present invention has been made.
Disclosure of Invention
The invention aims to provide a method for comprehensively refining and evaluating the similarity of infant formula powder, raw materials and auxiliary materials thereof, breast milk substitution fat and breast milk ganglioside in a multi-dimension way by taking a breast milk ganglioside database as a gold standard.
In order to achieve the above object, the present invention provides a method for evaluating similarity between a sample and breast milk in multiple dimensions, the method comprising the steps of:
(1) Establishing a local breast milk ganglioside database;
(2) Collecting a sample to be evaluated, and preprocessing;
(3) Measuring the content of the characteristic ganglioside subclasses and the content of the characteristic ganglioside molecules in the pretreated sample to be evaluated, and extracting peak areas, basal peak heights and peak numbers from a total ion flow diagram of the sample to be evaluated;
(4) The score G of the sample to be evaluated is calculated according to the following formula:
G = G s + G m + G t ；
wherein ,；
in the formula ,；H i is the content of characteristic ganglioside subclass i in breast milk, mg/L; h t Is the total ganglioside content in breast milk, mg/L; a is that i mg/L for the content of the characteristic ganglioside subclass i in the sample;
；
in the formula ,；R i is the correction coefficient of ganglioside, namely the sum of the content ratio of characteristic ganglioside molecules of the same kind in the sample and breast milk; h n Is the content of characteristic ganglioside molecules n in breast milk, mg/L; h t Is the total ganglioside content in breast milk, mg/L; a is that n mg/L for the content of characteristic ganglioside molecule n in the sample;
G t = (S p + S h + S n )/3；
wherein ,；
in the formula ,P n peak area of peak n of ganglioside total ion flow graph in sample;P i is thatPeak area of peak i of ganglioside total ion flow graph in breast milk;
；
in the formula ,I n the height of basal peak of ganglioside total ion flow diagram in the sample;I i the height of basal peak of the ganglioside total ion flow diagram in breast milk;
；
wherein ,N n peak number of ganglioside total ion flow diagram in sample;N i peak number of ganglioside total ion flow diagram in breast milk.
Through the technical scheme, the beneficial technical effects obtained by the invention are as follows:
(1) The invention establishes a database of healthy breast milk gangliosides, takes the healthy breast milk gangliosides as gold standard, and evaluates the similarity of infant formula powder, raw materials and auxiliary materials thereof, breast milk substitution fat and the breast milk gangliosides in detail on ganglioside subclasses and molecular level.
(2) The invention starts from three aspects of characteristic ganglioside subclass composition, characteristic ganglioside molecular composition and overall distribution condition evaluation, and the similarity of infant formula milk powder, raw and auxiliary materials and breast milk substitution fat and breast milk ganglioside is comprehensively evaluated in a multi-dimensional way; the composition, content and weight of each ganglioside are considered during evaluation, so that the simulation quality of infant formula milk powder and raw and auxiliary materials thereof on breast milk gangliosides can be better evaluated.
(3) From the aspect of nutrition, the evaluation method is objectively verified according to the clinical experimental result of the actual sample and the score comparison of the clinical experimental result.
Drawings
FIG. 1 is a total ion flow diagram of the ganglioside of the human milk standard of example 1;
FIG. 2 is a total ion flow diagram of gangliosides of the infant formula with MFGM added thereto in example 2;
FIG. 3 is a total ion flow diagram of gangliosides of the plain infant formula of example 2;
FIG. 4 is a graph of serum immune factors and hormone levels of 31 day-old piglets fed differently in example 3;
FIG. 5 is a hierarchical clustering heat map of piglet serum metabolites for different feeding modes in example 3;
FIG. 6 is a SPLS-DA plot of serum metabolites of piglets fed differently in example 3;
FIG. 7 is a graph showing the body length and body weight of infants 1 month old, 4 months old, and 6 months old with different feeding modes in example 3;
FIG. 8 is a graph of ASQ results for infants of 4 and 6 months of age fed by the different feeding regimen of example 3.
Detailed Description
The endpoints and any values of the ranges disclosed herein are not limited to the precise range or value, and are understood to encompass values approaching those ranges or values. For numerical ranges, one or more new numerical ranges may be found between the endpoints of each range, between the endpoint of each range and the individual point value, and between the individual point value, in combination with each other, and are to be considered as specifically disclosed herein.
The first aspect of the invention provides a method for evaluating the similarity between a sample and breast milk in a multi-dimensional manner, which comprises the following steps:
(1) Establishing a local breast milk ganglioside database;
(2) Collecting a sample to be evaluated, and preprocessing;
(3) Measuring the content of the characteristic ganglioside subclasses and the content of the characteristic ganglioside molecules in the pretreated sample to be evaluated, and extracting peak areas, basal peak heights and peak numbers from a total ion flow diagram of the sample to be evaluated;
(4) The score G of the sample to be evaluated is calculated according to the following formula:
G = G s + G m + G t ；
wherein ,；
in the formula ,；H i is the content of characteristic ganglioside subclass i in breast milk, mg/L; h t Is the total ganglioside content in breast milk, mg/L; a is that i mg/L for the content of the characteristic ganglioside subclass i in the sample;
；
in the formula ,；R i is the correction coefficient of ganglioside, namely the sum of the content ratio of characteristic ganglioside molecules of the same kind in the sample and breast milk; h n Is the content of characteristic ganglioside molecules n in breast milk, mg/L; h t Is the total ganglioside content in breast milk, mg/L; a is that n mg/L for the content of characteristic ganglioside molecule n in the sample;
G t = (S p + S h + S n )/3；
wherein ,；
in the formula ,P n peak area of peak n of ganglioside total ion flow graph in sample;P i is thatPeak area of peak i of ganglioside total ion flow graph in breast milk;
；
in the formula ,I n the height of basal peak of ganglioside total ion flow diagram in the sample;I i the height of basal peak of the ganglioside total ion flow diagram in breast milk;
；
wherein ,N n peak number of ganglioside total ion flow diagram in sample;N i peak number of ganglioside total ion flow diagram in breast milk.
In the present invention, R i The correction coefficient is the sum of the ratio of the content of the characteristic ganglioside molecules of the common species in the sample and the breast milk, namely the sum of the ratio of the content of the characteristic ganglioside molecules of the common species in the sample (all) and the breast milk (the ratio of the content of the total ganglioside in the sample) (the ratio of the total ganglioside in the sample) and the total ganglioside in the sample).
The invention starts from three aspects of ganglioside composition, content and total ion flow graph anastomosis degree, and evaluates the similarity of infant formula milk powder, raw and auxiliary materials, breast milk substitution fat and breast milk ganglioside in a multidimensional way. Firstly, integrally judging infant formula powder and raw and auxiliary materials through ganglioside subclasses; then, the infant formula powder and raw materials and auxiliary materials of the infant formula powder are compared with the composition and content similarity of the breast milk ganglioside by taking the composition and content of the breast milk ganglioside as the standard; finally, evaluating the simulation situation of the ganglioside integral distribution situation of the infant formula powder by comparing the ganglioside integral distribution situation in breast milk with the ganglioside integral distribution situation in the infant formula powder. The invention can comprehensively evaluate the simulation quality of infant formula milk powder and ganglioside which is the raw material and auxiliary material thereof.
The total ion flow graph is a plot of total ion flow intensity of a mass spectrum detector over time, with the ordinate representing the total intensity of current collecting stored ions and the abscissa representing the time of generation of ions or the number of scans of a continuous scan. As can be seen by comparing the total ion flow patterns of the samples, when the response intensities and retention times of the chromatographic peaks between the respective patterns substantially overlap, a smaller difference between the corresponding samples is indicated.
Because the total ganglioside content in breast milk is low and only accounts for 0.005% -0.12%, the qualitative and quantitative result cannot fully show the whole distribution of gangliosides in breast milk, and part of gangliosides with low content are undetected because the content is lower than the detection limit of an instrument. Since each time point of the total ion flowsheet contains the concentration of each ion, ganglioside information below the detection limit is also contained in the total ion flowsheet, ganglioside overall composition similarity can be assessed by comparing the similarity of the total ion flowsheets.
The invention researches the integral composition condition of gangliosides in breast milk fat globules through a total ion flow graph (TIC), simultaneously detects the integral distribution condition of gangliosides in infant formula powder, and evaluates the influence of a processing technology and the like on the integral distribution condition of gangliosides by comparing the total ion flow graph.
According to some embodiments of the invention, the pretreatment in step (2) comprises the steps of:
diluting the internal standard with a compound solution until the concentration of the internal standard is consistent with that of breast milk, adding the diluted compound solution into a sample to be evaluated, adding ultrapure water, methanol and dichloromethane to mix, adding the ultrapure water and the dichloromethane to mix, performing first centrifugation, and separating to obtain a first organic phase and a first aqueous phase; adding ultrapure water, methanol and dichloromethane into the first organic phase, mixing, performing secondary centrifugation, and separating to obtain a second organic phase and a second aqueous phase; mixing the first aqueous phase with dichloromethane, performing third centrifugation, and separating to obtain a third organic phase and a third aqueous phase; and combining the second water phase and the third water phase, drying with nitrogen, and re-dissolving with a complex solution to obtain a pretreated sample to be evaluated.
According to some embodiments of the invention, the multiple solution is a solution having a volume ratio of methanol to water of 1-9:1 or 4:1.
According to some embodiments of the invention, the internal standard is a standard comprising C18:0GM3-d 5 and C18 Ganglioside GD3-d 3.
According to some embodiments of the invention, the conditions of the first centrifugation include: centrifuging for 10-20min, or 15min; the centrifugation speed is 3000-8000r/min, or 6000r/min.
According to some embodiments of the invention, the conditions of the second centrifugation include: centrifuging for 5-15min, or 10min; the centrifugation rate is 2000-4000 Xg, or 3000 Xg.
According to some embodiments of the invention, the conditions of the third centrifugation comprise: centrifuging for 10-20min, or 15min; the centrifugation speed is 3000-8000r/min, or 6000r/min.
According to some embodiments of the invention, in step (3), the characteristic ganglioside subclass and the characteristic ganglioside molecule content are determined using ultra-high performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry. In the invention, ultra-high performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) is adopted, an ESI electrospray ion source and a PeakView2.2 and MultiQuant3.0.2 data processing system are equipped, ganglioside composition in a sample is measured, and information such as peak area, basal peak height, peak number and the like is extracted from a total ion flow diagram.
According to some embodiments of the invention, the method of ultra high performance liquid chromatography is: a Waters BEH C18 chromatographic column with the specification of 100mm multiplied by 2.1mm and 1.7 mu m, the sample injection amount of 10 mu L, the flow rate of 0.3mL/min and the column temperature of 50 ℃; mobile phase A is 1mM ammonium acetate in methanol/water solution with the volume ratio of 1:9, and mobile phase B is 1mM ammonium acetate in methanol; the elution gradient is:。
according to some embodiments of the invention, the conditions of the quadrupole time-of-flight mass spectrum are: detecting in negative ion mode, mass spectrum detection parameters: the accumulation time is 0.25secs, the scanning range is 100-2000Da, the elution time is 14min, the heating air pressure is 40psi, the auxiliary heating air pressure is 35psi, the air curtain air pressure is 15psi, the ion source temperature is 400 ℃, the ion spray voltage is-4500V, the cluster removing voltage is-40V, and the collision energy is-40V.
According to some embodiments of the invention, the characteristic ganglioside subclass in step (3) comprises GM3 and GD3.
According to some embodiments of the present invention, the characteristic ganglioside molecules in step (3) include GM3 40:1;2, GM3 42:1;3, GD 3:1; 2, GM3 34:1;2, GD 3:40:2; 3, GM3 36:1;2, GM3 38:1;2, GM3 34:2;3, GD 3:1; 2, GM 3:2; 2, GD 3:0; 2, GD 3:42:1; 3, GM 3:0; 3, GD 3:2; 2, GM 3:1; 2, GD 3:42:0; 3, GM 3:0; 2, GD 3:40:2, GD 3:40:1; 2, GM 3:42; 3, GD3 36:1; 2) GM3 34:1, GM3 36:0, 2, GM3 34:0, 2, GM3 38:0, 2, GM3 40:2, 2, GD3 34:1, 2, GD3 40:0, 2, GM3 32:1, 2, GM3 38:2, 2, GM3 34:0, 3, GM3 40:0, 3, GM3 42:0, 2, GM3 36:2, 2, GM3 38:0, 3, GM3 36:1, 3, GM3 36:0, 3, GM 40:1, 3, GM 44:1, 2, GM3 32:0, 2, GM3 34:2, GM3 38:1, 3, GM3 32:1, 3, and GM3 36:2, 3.
The present invention will be described in detail by examples.
The following examples were conducted under conventional conditions or conditions recommended by the manufacturer, without specifying the specific conditions. The reagents or apparatus used were conventional products available commercially without the manufacturer's knowledge.
Example 1
The embodiment provides a method for establishing a local breast milk database.
61 healthy parturients from four areas of Beijing, tangshan, liuyang and Luoyang were selected. All volunteers had normal physical signs and infants were produced at term.
And (3) tracking and collecting 233 total breast milk samples of 0-6 months from 61 healthy puerpera, and recording the development condition of each breast milk sample corresponding to the infant.
The breast milk collecting method comprises the following steps: all the primary volunteers had normal physical signs and had term delivery (38-42 weeks gestation) without congenital or genetic disease. All volunteers were required to empty one breast at 6:00-7:00 a.m., followed by collection of whole milk from one breast (previously empty) at 9:00-11:00 a.m.. Mixing whole milk, packaging into 1mL sterile freezing tube, and storing in-80deg.C ultra-low temperature refrigerator.
The sample needs to be pretreated before entering the chromatograph and the mass spectrum, and the specific method for pretreatment is as follows: the standard was diluted to a concentration similar to that of breast milk using a reconstituted solution of methanol in water (4:1, v/v). mu.L of 100mg/L C18:0 GM3-d5 and 25 mu.L of 100mg/L C Ganglioside GD3-d3 standard were added to 200. Mu.L of sample to be evaluated, 200. Mu.L of ultrapure water, 2mL of methanol, 900. Mu.L of dichloromethane were added, then 200. Mu.L of ultrapure water and 900. Mu.L of dichloromethane were added, and the mixture was centrifuged at 6000r/min for 15min to separate the organic phase and the aqueous phase. The organic phase was added with 1mL of ultrapure water, 2.2mL of methanol and 600. Mu.L of methylene chloride, and the mixture was centrifuged at 3000 Xg for 10min to separate the organic phase and the aqueous phase. The aqueous phase was mixed with 1.8mL of dichloromethane, centrifuged at 6000r/min for 15min, the aqueous phases were separated and combined. The aqueous phase was dried with nitrogen and redissolved with 200 μl of the redissolved solution to obtain the pretreated sample to be evaluated.
And (3) adopting ultra-high performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS), and providing an ESI electrospray ion source and a PeakView2.2 and MultiQuant3.0.2 data processing system to measure ganglioside composition in the sample and extract information such as peak area, basal peak height, peak number and the like from the total ion flow diagram.
The detection conditions are as follows: waters BEH C18 column (100 mm. Times.2.1 mm,1.7 μm) chromatography column for ganglioside separation; the sample injection amount is 10 mu L, the flow rate is 0.3mL/min, and the column temperature is 50 ℃; mobile phase a was a 1mM solution of ammonium acetate in methanol/water (1:9, v/v) and mobile phase B was a 1mM solution of ammonium acetate in methanol; the elution gradient is shown in table 1 below.
Table 1 gradient elution conditions for gangliosides:
the quadrupole time-of-flight mass spectrometry conditions are: detecting in negative ion mode, mass spectrum detection parameters: the accumulation time (accumulation time) is 0.25secs, the scanning range (tof maps) is 100-2000Da, the elution time (duration) is 14min, the heating air pressure (GS 1) is 40psi, the auxiliary heating air pressure (GS 2) is 35psi, the air curtain air pressure (CUR) is 15psi, the ion source Temperature (TEM) is 400 ℃, the Ion Spray Voltage (ISVF) is 4500V, the declustering voltage (DP) is 40V, and the Collision Energy (CE) is 40V.
The sample prepared by mixing the above 233 breast milk samples in equal volume was used as a breast milk standard, and analyzed by the above chromatography and mass spectrometry method to obtain 2 ganglioside subclasses and 43 characteristic ganglioside molecules, and the results are shown in tables 2 and 3.
Table 2 example 1 table of weight results for characteristic ganglioside subclasses in breast milk standard:
。
table 3 example 1 table of molecular weight results for characteristic gangliosides in breast milk standard:
。
it should be noted that the other items in table 3 are the sum of the contents of other ganglioside molecules except the 43 characteristic ganglioside molecules, such as GM3 30:1;2,GM3 44:2;2,GD3 32:1;2,GD3 34:0;2,GD3 34:1;3,GD3 38:0;2,GD3 38:0;3,GD3 40:0;3,GD3 40:2;2, etc., which are not fully listed in this example.
The subsequent analysis of other milk samples uses the characteristic ganglioside subclasses and the characteristic ganglioside molecular content of the breast milk standard as evaluation references.
However, it should be noted that, due to the difference between eating habits and living habits of various regions, the ganglioside components and contents in the breast milk of healthy women in different regions are different, and thus, for those skilled in the art, the source of the breast milk sample may be selected according to actual needs, and accordingly, the measurement results of the characteristic ganglioside subclasses and the characteristic ganglioside molecules of the breast milk standard prepared with the breast milk sample will also vary.
The total ion flow diagram of breast milk is shown in figure 1. And obtaining information such as peak area, basal peak intensity, peak number and the like of the total ion flow graph by using Data and Peak Table functions in PeakView software.
Example 2
The score of each sample was evaluated using the characteristic ganglioside subclass content, characteristic ganglioside molecular content, and total ion flow graph fitness of the breast milk standard described in example 1 as evaluation criteria.
Fig. 2 shows a total ion flow diagram of gangliosides of infant formulas with MFGM added. Fig. 3 shows a general infant formula ganglioside total ion flow diagram.
The score G is calculated according to the following formula:
G = G s + G m + G t ；
wherein ,；
in the formula ,；H i is the content of characteristic ganglioside subclass i in breast milk, mg/L; h t Is the total ganglioside content in breast milk, mg/L; a is that i mg/L for the content of the characteristic ganglioside subclass i in the sample;
；
in the formula ,；R i is the correction coefficient of ganglioside, namely the sum of the content ratio of characteristic ganglioside molecules of the same kind in the sample and breast milk; h n Is the content of characteristic ganglioside molecules n in breast milk, mg/L; h t Is the total ganglioside content in breast milk, mg/L; a is that n mg/L for the content of characteristic ganglioside molecule n in the sample;
G t = (S p + S h + S n )/3；
wherein ,；
in the formula ,P n peak area of peak n of ganglioside total ion flow graph in sample;P i is thatPeak area of peak i of ganglioside total ion flow graph in breast milk;
；
in the formula ,I n the height of basal peak of ganglioside total ion flow diagram in the sample;I i the height of basal peak of the ganglioside total ion flow diagram in breast milk;
；
wherein ,N n peak number of ganglioside total ion flow diagram in sample;N i peak number of ganglioside total ion flow diagram in breast milk.
wherein ,A i 、A n 、P n 、I n AndN n the measurement was carried out in the same manner as in the breast milk sample in example 1.
The scores of the infant formulas with MFGM added and the normal infant formulas calculated as examples according to the above methods are shown in table 4.
The infant formula powder added with the MFGM is derived from 1-stage milk powder added with the MFGM in a certain brand on the market, and the common infant formula powder is derived from 1-stage milk powder not added with the MFGM in a certain brand on the market.
Table 4 sample evaluation score table:
。
as can be seen from Table 4, the two formulations G s Score the same but due to G m and G t The final scores G are different, and the score of the infant formula powder added with the MFGM is higher than that of the infant formula powder added with the MFGM. This demonstrates that the evaluation at the ganglioside subclass level alone does not result in an effective evaluation, whereas the present invention provides for multidimensional scaling from the characteristic ganglioside subclass composition, characteristic ganglioside molecular composition, and overall distributionThe evaluation can obtain effective evaluation results in various cases.
It should be noted, however, that although the above two kinds of formulas are taken as examples for the relevant evaluation in this example, the method of the present invention is applicable not only to the evaluation of formulas but also to the evaluation of lipid raw materials as raw materials and auxiliary materials, and various dairy products and products.
Example 3
This example scored the actual samples using the method described in example 2 and objectively verified the results of the assessment method from a nutritional perspective.
In this example, the effects of breast milk (BF), normal infant formula (FF 1) and MFGM-added infant formula (FF 2) on infant development, and the effects of three feeding regimes on immune metabolism and hormone levels of 31-day-old piglets, were explored.
(1) Animal experiments were used for nutritional verification.
5 pregnant English-line large-white binary hybrid sows with similar gestation ages are selected for carrying out. The parturient SPF piglets (n=20) are naturally born for the SPF sows. Healthy piglets with similar weight and length are selected, and breast feeding is unified within 3 days after birth, so that the piglets can obtain enough maternal immunity and nutrition. The breast feeding was performed 3 days before the start of the experiment, and then the feeding experiment was formally performed until 31 days. After the initiation of the formal experiment, the random was divided into 3 groups, which were respectively breast milk group (BF group), ordinary infant formula (FF 1 group) and infant formula (FF 2 group) to which MFGM was added. The piglets and sows of the breast milk group are managed in the same pigsty, the breast feeding of the pigs is continued until weaning is carried out for 21 days, and then the piglets are fed with piglet feed; the formula powder group is fed with the reconstituted milk prepared from milk powder and water, and is fed by a milk bottle (a food trough for later period) for up to 31 days, and piglets are isolated from sows during the period, and are independently fed and managed by a cage. The sample collection time was 31 days post partum.
Collection of serum samples: feeding on days 7, 14, 21 and 31, and taking blood in an empty stomach. 3mL of anterior vena cava blood is collected by a non-anticoagulation blood collection tube, the blood collection tube is stood for 1h, and supernatant is obtained after centrifugation at 3000rpm and 4 ℃ for 20min, and the serum is obtained after centrifugation at-20 ℃ and is preserved for the determination of immune and hormone indexes and NMR metabolites.
Determination of immune and hormonal indicators: the serum immune factors and hormone level changes of piglets were measured using ELISA kit, mainly comprising 5-hydroxytryptamine (5-HT), dopamine (DA), epinephrine (EPI), norepinephrine (NE), parathyroid hormone-related protein (PTHrP), 2, 5-hydroxyvitamin D3 (HVD 3), igA, igG, igG1, early growth response factor 1 (EGR 1) and the like.
Determination of serum NMR metabolites: the sample was centrifuged at 13,000rpm for 2min and 540. Mu.L of the aqueous layer was transferred to a 2ml centrifuge tube. 60 μl DSS standard solution (Anachro, canada) was added. Then premix and transfer to a 5mm NMR tube (Norwell, usa). Spectra were collected using NMR spectroscopy (Bruker AV III 600 mhz). Subsequent signal processing was performed using Chenomx NMR Suite 8.1.1 (Chenomx inc., edmonton, canada). Then segment integration and baseline correction are performed. All spectra were referenced to the internal standard DSS and to the Chenomx compound library. A total of 67 metabolites were obtained from the NMR spectroscopy platform. These metabolites include various types of metabolites including vitamins/cofactors, organic acids, amino acid derivatives, ketones, nucleic acid components, amino compounds, amines, sugars, alcohols, and amides.
Serum immune factors and hormone levels of 31 day old piglets from different feeding regimes were tested and plotted (FIG. 4) for Kruskall-Wallis differences. It can be seen that the infant formula milk powder can normally promote the growth and development of piglets, and the infant formula milk powder added with the MFGM obviously increases the IgA level of the piglets and the content of serum betaine. IgA, igG, early growth response factor 1 (EGR 1) were significantly higher in both milk powder groups than in the breast milk group, and IgG1 were both higher in the FF2 group than in the FF1 group.
As can be seen from fig. 5 and 6, 67 metabolites were measured in total for all piglets, and the serum metabolite profiles of the breast milk group and the two infant formula groups were substantially identical before the first week of dietary treatment had yet to begin, and the 21-day-old piglet serum samples were significantly separated from the other time-spotted samples. Infant formulas supplemented with MFGM were found to significantly increase serum IgA levels in piglets by Kruskal-Wallis differential assay. The infant formula fed piglets had significantly increased levels of ascorbic acid (vitamin C), trimethylamine-N-oxide (TMAO) and galactose compared to breast feeding. Wherein the content of ascorbic acid and galactose in the FF2 group is higher than that in the FF1 group.
In conclusion, the experiment proves that the high-score FF2 group obviously increases the IgA level of piglets and the content of serum betaine when being fed, and the content of IgG, igG1, ascorbic acid and galactose is higher than that of the low-score FF1 group, so that the piglets fed by the FF2 group are better than the FF1 group in the aspects of immunity, oxidation resistance and nutrition metabolism.
(2) Clinical trials were used for nutritional verification.
Depending on the hospital, the right-born healthy term neonates are recruited, and their legal guardians need to carefully read the informed consent and inform the researchers of the decision to participate in the study. Volunteers were randomly and evenly divided into 3 groups, breast milk group (BF group), normal formula (FF 1 group) and experimental formula (FF 2 group), respectively. During the study period, the breast milk group is adhered to pure breast feeding, the milk powder group is adhered to feeding of the formula powder fed by eating, the common formula powder or the experimental formula powder is randomly distributed according to the random grouping, and other brands of milk powder cannot be eaten during the study period. Sample collection times were 1 month of age, 4 months of age, and 6 months of age, respectively.
As can be seen from fig. 7, at each month of age, the infants in FF2 group had a higher length than those in FF1 group, the gap from the breast milk group was gradually decreased, and at 6 months of age, the gap was substantially disappeared, and the infants in FF2 group and breast milk group had a significantly higher length than those in FF1 group. The weight of the infants in FF2 group had exceeded the weight of the FF1 group at 6 months of age and was similar to the weight of the breast milk group. From this, the feeding effect of FF2 group is better than that of FF1 group, and is more similar to that of breast milk.
The neuropsychological development of the above three groups of infants at 4 and 6 months of age was evaluated using an ASQ-3 childhood neurodevelopment evaluation system, and the results are shown in fig. 8. It can be seen that the FF2 group of infants had higher scores for communication, coarse movements, person-social, and problem solving than the FF1 group, and were closer to and not significantly different from the breast milk group, whereas the FF1 group of infants had significantly lower scores for coarse movements at 4 months of age than the breast milk group, and had significantly lower person-social movements at 6 months of age than the breast milk group. The above results demonstrate that higher scoring FF2 is significantly better than lower scoring FF1 in promoting neurological development in infants and is more similar to breast feeding.
In conclusion, through various experiments, the FF2 group with higher score is superior to the FF1 group with lower score in promoting the development of infants, enhancing the resistance of infants and promoting the neural development of infants, and is more similar to the breast milk group. Therefore, the method for evaluating the similarity between the milk product and the breast milk according to the ganglioside provided by the invention can objectively evaluate the nutrition level of the milk product or raw and auxiliary materials at the molecular level, and has a certain guiding significance for the subsequent simulation of the breast milk at the molecular level.
The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited thereto. Within the scope of the technical idea of the invention, a number of simple variants of the technical solution of the invention are possible, including combinations of the individual technical features in any other suitable way, which simple variants and combinations should likewise be regarded as being disclosed by the invention, all falling within the scope of protection of the invention.
Claims (10)
1. A method for multi-dimensional evaluation of similarity of a sample to breast milk, the method comprising the steps of:
(1) Establishing a local breast milk ganglioside database;
(2) Collecting a sample to be evaluated, and preprocessing;
(3) Measuring the content of the characteristic ganglioside subclasses and the content of the characteristic ganglioside molecules in the pretreated sample to be evaluated, and extracting peak areas, basal peak heights and peak numbers from a total ion flow diagram of the sample to be evaluated;
(4) The score G of the sample to be evaluated is calculated according to the following formula:
G = G s + G m + G t ；
wherein ,；
in the formula ,；H i is the content of characteristic ganglioside subclass i in breast milk, mg/L; h t Is the total ganglioside content in breast milk, mg/L; a is that i mg/L for the content of the characteristic ganglioside subclass i in the sample;
；
in the formula ,；R i is the correction coefficient of ganglioside, namely the sum of the proportion of the total ganglioside content in the sample of the characteristic ganglioside molecule content of all species which are common with breast milk in the sample; h n Is the content of characteristic ganglioside molecules n in breast milk, mg/L; h t Is the total ganglioside content in breast milk, mg/L; a is that n mg/L for the content of characteristic ganglioside molecule n in the sample;
G t = (S p + S h + S n )/3；
wherein ,；
in the formula ,P n peak area of peak n of ganglioside total ion flow graph in sample;P i is thatPeak area of peak i of ganglioside total ion flow graph in breast milk;
；
in the formula ,I n the height of basal peak of ganglioside total ion flow diagram in the sample;I i the height of basal peak of the ganglioside total ion flow diagram in breast milk;
；
wherein ,N n peak number of ganglioside total ion flow diagram in sample;N i peak number of ganglioside total ion flow diagram in breast milk.
2. The method of claim 1, wherein the preprocessing in step (2) comprises the steps of:
diluting the internal standard with a compound solution until the concentration of the internal standard is consistent with that of breast milk, adding the diluted compound solution into a sample to be evaluated, adding ultrapure water, methanol and dichloromethane to mix, adding the ultrapure water and the dichloromethane to mix, performing first centrifugation, and separating to obtain a first organic phase and a first aqueous phase; adding ultrapure water, methanol and dichloromethane into the first organic phase, mixing, performing secondary centrifugation, and separating to obtain a second organic phase and a second aqueous phase; mixing the first aqueous phase with dichloromethane, performing third centrifugation, and separating to obtain a third organic phase and a third aqueous phase; and combining the second water phase and the third water phase, drying with nitrogen, and re-dissolving with a complex solution to obtain a pretreated sample to be evaluated.
3. The method of claim 2, wherein the multiple solution is a solution having a volume ratio of methanol to water of (1-9): 1 or 4:1.
4. The method of claim 2, wherein the internal standard is a standard comprising c18:0gm3-d 5 and C18 Ganglioside GD3-d 3.
5. The method of claim 2, wherein the conditions of the first centrifugation comprise: centrifuging for 10-20min, or 15min; the centrifugal speed is 3000-8000r/min, or 6000r/min;
and/or, the conditions of the second centrifugation include: centrifuging for 5-15min, or 10min; the centrifugation rate is 2000-4000 Xg, or 3000 Xg;
and/or, the conditions of the third centrifugation include: centrifuging for 10-20min, or 15min; the centrifugation speed is 3000-8000r/min, or 6000r/min.
6. The method of claim 1, wherein in step (3), the characteristic ganglioside subclasses and the characteristic ganglioside molecular content are determined using ultra-high performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry.
7. The method of claim 6, wherein the method of ultra-high performance liquid chromatography is: a Waters BEH C18 chromatographic column with the specification of 100mm multiplied by 2.1mm and 1.7 mu m, the sample injection amount of 10 mu L, the flow rate of 0.3mL/min and the column temperature of 50 ℃; mobile phase A is 1mM ammonium acetate in methanol/water solution with the volume ratio of 1:9, and mobile phase B is 1mM ammonium acetate in methanol; the elution gradient is:
。
8. the method of claim 6, wherein the conditions of the quadrupole time-of-flight mass spectrum are: detecting in negative ion mode, mass spectrum detection parameters: the accumulation time is 0.25secs, the scanning range is 100-2000Da, the elution time is 14min, the heating air pressure is 40psi, the auxiliary heating air pressure is 35psi, the air curtain air pressure is 15psi, the ion source temperature is 400 ℃, the ion source spraying voltage is-4500V, the cluster removing voltage is-40V, and the collision energy is-40V.
9. The method of claim 1, wherein the characteristic ganglioside subclass in step (3) comprises GM3 and GD3.
10. The method of claim 1, wherein, the characteristic ganglioside molecules in step (3) include GM3 40:1;2, GM3 42:1;3, GD 3:1; 2, GM3 34:1;2, GD 3:40:2; 3, GM3 36:1;2, GM3 38:1;2, GM3 34:2;3, GD 3:1; 2, GM 3:2; 2, GD 3:0; 2, GD 3:42:1; 3, GM 3:0; 3, GD 3:2; 2, GM 3:1; 2, GD 3:42:0; 3, GM 3:0; 2, GD 3:40:2, GD 3:40:1; 2, GM 3:42; 3, GD3 36:1; 2) GM3 34:1, GM3 36:0, 2, GM3 34:0, 2, GM3 38:0, 2, GM3 40:2, 2, GD3 34:1, 2, GD3 40:0, 2, GM3 32:1, 2, GM3 38:2, 2, GM3 34:0, 3, GM3 40:0, 3, GM3 42:0, 2, GM3 36:2, 2, GM3 38:0, 3, GM3 36:1, 3, GM3 36:0, 3, GM 40:1, 3, GM 44:1, 2, GM3 32:0, 2, GM3 34:2, GM3 38:1, 3, GM3 32:1, 3, and GM3 36:2, 3.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2020232813A1 (en) * | 2019-05-20 | 2020-11-26 | 北京三元食品股份有限公司 | Method for extracting lipids in food and method for detecting lipids in food |
| CN113575696A (en) * | 2021-07-30 | 2021-11-02 | 江南大学 | Preparation method of structural lipid based on breast milk triglyceride composition |
| CN113640449A (en) * | 2021-10-13 | 2021-11-12 | 山东省产品质量检验研究院 | Rapid detection method for polylactic acid of bio-based degradable material |
| CN115267098A (en) * | 2022-07-29 | 2022-11-01 | 北京三元食品股份有限公司 | A method for evaluating milk product based on lipid component |
| CN116106231A (en) * | 2023-04-13 | 2023-05-12 | 北京三元食品股份有限公司 | Method for evaluating similarity between sample and breast milk in multiple dimensions |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2020232813A1 (en) * | 2019-05-20 | 2020-11-26 | 北京三元食品股份有限公司 | Method for extracting lipids in food and method for detecting lipids in food |
| CN113575696A (en) * | 2021-07-30 | 2021-11-02 | 江南大学 | Preparation method of structural lipid based on breast milk triglyceride composition |
| CN113640449A (en) * | 2021-10-13 | 2021-11-12 | 山东省产品质量检验研究院 | Rapid detection method for polylactic acid of bio-based degradable material |
| CN115267098A (en) * | 2022-07-29 | 2022-11-01 | 北京三元食品股份有限公司 | A method for evaluating milk product based on lipid component |
| CN116106231A (en) * | 2023-04-13 | 2023-05-12 | 北京三元食品股份有限公司 | Method for evaluating similarity between sample and breast milk in multiple dimensions |
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