CN105067604A - Method for rapidly and semi-quantitatively determining glucose content in fruits by using paper-based chip - Google Patents
Method for rapidly and semi-quantitatively determining glucose content in fruits by using paper-based chip Download PDFInfo
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Abstract
The present invention discloses a method for rapidly and semi-quantitatively determining the glucose content in fruits by using a paper-based microfluidic chip. The method comprises: preparing glucose standard solutions with different concentrations, adopting the paper-based microfluidic chip as a test platform, using ImageJ software to calculate the average gray values of each paper-based microfluidic chip reaction zone before and after the reaction so as to obtain the standard curve of the average gray value changes corresponding to different glucose concentrations, designing colourimetric cards of different grey gradients, obtaining the average gray value change of the glucose concentration in the fruit, and comparing with the colourimetric cards and the standard curve so as to semi-quantitatively determine the glucose content in the fruit. According to the present invention, the microfluidic paper-based chip sensor has advantages of short reaction time, good selectivity, good repeatability and the like; and the test platform can further be used for detecting the foodborne bacteria content in the food, and provides positive significance for the on-site rapid detection of the food safety.
Description
Technical field
The invention belongs to fruit quality express-analysis technical field, be specifically related to a kind of method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit.
Background technology
Along with growth in the living standard, consumer proposes higher requirement to fruit quality.Mostly adopting both at home and abroad fruit internal quality has abrasive laboratory chemical method to measure and the undamaged detection method based on spectrum, and these methods complete detection all in the quiescent state, cannot realize the requirement that real-time online detects.Therefore fruit internal quality on-line real-time measuremen technology be realize fruit commercial treatment and quality rapid classification be badly in need of solve a gordian technique difficult problem.
The content of fruit sugar is that its nutritive value one of characterizes, there is destruction sample, complicated operation, length consuming time in traditional sugar degree detection method, cost is high and cannot realize the shortcomings such as on-line checkingi, and the experimental apparatus that need be equipped with costly, and send professional to have operated chemical analysis, but the demand of the field quick detection to the field such as environment, food service industry can not be satisfied with.Therefore, a kind ofly to use simply, easily analytical equipment will become the direction of Future Development.
The features such as paper substrate microfluidic chip technology has easy processing, cost is low, analysis speed is fast, microminiaturized and portability.Paper substrate chip, as an important branch of micro-fluidic chip, obtains more concern and develops faster in research in the last few years.Compared with the micro-fluidic chip made with traditional materials such as glass, silicon, quartz, paper substrate micro-fluidic chip has that cost is low, technique is simple, portability is strong, be easy to the advantages such as process.Its as substrate with paper, forms plane or three-dimensional parent/hydrophobic microfine channel network and correlation analysis device by various process technology, builds " on paper microscale experiment room " (Lab-on-paper).The microminiaturization in Realization analysis laboratory, integrated and portability.
Paper substrate micro-fluidic chip equipment is many capillarities of water wettability based on filter paper itself and inside, and when without the need to outer power, the feature that testing sample solution flows at its surface by means of the capillary action of filter paper is made.Paper substrate micro-fluidic chip also have can single use, sample consumption is few, analysis speed is very fast, the simple to operate and advantage such as analysis while simultaneously can realizing many kinds of substance.At present, the application that the device of on-the-spot on-line checkingi is ideal just can obtain a direct analysis result only by a primary sample sample introduction exactly, thus has applicability and application prospect widely in the field such as chemistry, biology, medical science, food inspection.
Paper substrate micro-fluidic chip job operation is varied, reach its maturity.Main research and design goes out form parent/hydrophobic channel network on paper based substrate, is just achieved fluid and controls and analyze to detect.Conventional micro-fluidic paper substrate job operation has physical method and chemical method.
Summary of the invention
The object of the present invention is to provide a kind of method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit, and then using glucose as sugar metrics evaluation fruit internal quality; Also there is provided a kind of guide for method diabetic be easily understood simultaneously and select fruit.The present invention utilizes paper substrate micro-fluidic chip to invent and utilizes functionalization paper substrate microfluidic chip technology and colorimetric analysis technology to organically combine, set up the new method detecting sugar degree fast.Further, paper substrate micro-fluidic chip being applied to innovatively the detection of sugar in fruit has important practical significance, by detecting the quality of the measurable fruit of content of sugar in fruit.
Laser-engraving technique is applied in the present invention; In AutoCad software, design the shape of paper substrate chip, in advance paper substrate is cut into required figure with laser engraving machine, smooth surface, mellow and full paper chip can be obtained.Based on the photo of colour developing degree different on paper substrate after the dropping fruit syrup taken by digital camera, ImageJ software measurement is utilized to go out the gray-scale value of conversion zone, and then calculate the gray scale changes values of differential responses time paper substrate, the grey scale change value curve drawn by contrast standard concentration of glucose, infers the amount contained glucose in fruit syrup.
Based on above-mentioned thought, the present invention utilizes the fusion of design of material thought and paper substrate microfluidic chip technology, builds the Fast Detection Technique being applied to glucose in fruit.Its selective binding power is improved by the method for chemical modification, thus improve its concentration effect and sensitivity further, high flux can be carried out to glucose in fruit, the quick detection of high resolution, paper substrate microfluidic chip technology is applied to innovatively the detection of glucose in fruit and has important practical significance, by detecting the quality of the measurable fruit of content of glucose in fruit and then reaching Fast nondestructive evaluation fruit internal nutritional labeling, diabetes special population can be instructed the selection of fruit, to promoting China fruit brand position and strengthening the competitive power in its at home and abroad market, the aspect such as to increase farmers' income is of great immediate significance.
The object of the invention is to be achieved through the following technical solutions:
The present invention relates to a kind of method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit, described method comprises the steps:
A, the Glucose standards solution of preparation variable concentrations, respectively with paper substrate micro-fluidic chip for test platform (response matrix), the average gray value of the chromogenic reaction of iodine on each paper substrate micro-fluidic chip conversion zone in application ImageJ software measurement reaction front and back, make the typical curve of average gray value knots modification corresponding to different concentration of glucose, and the colorimetric card of different shade of gray is (by described normal concentration glucose R average after the reaction of paper substrate micro-fluidic chip, G average and B average mapping software draw out the glucose colorimetric card of variable concentrations, after statistical analysis, print over colour atla, ),
B, squeezed the juice by fruit to be measured, with paper substrate micro-fluidic chip for detection platform, before and after the reaction of application ImageJ software measurement, the average gray value of the chromogenic reaction of iodine on paper substrate chip conversion zone, calculates the average gray value knots modification that fruit to be measured is corresponding; Contrast described colorimetric card and typical curve, get final product the glucose content in sxemiquantitative fruit to be measured.
Preferably, described paper substrate micro-fluidic chip will have the polyglycol grafting of high-hydrophilic energy to paper substrate micro-fluidic chip surface, the functionalization paper substrate micro-fluidic chip of acquisition by the method for atom transition free radical polymerization reaction.
Preferably, the preparation of described paper substrate micro-fluidic chip specifically comprises the steps:
S1, on microslide, modify allyltrimethoxysilanis;
S2, coat PDMS by the substrate being carved with colored shape;
S3, on microslide, accurately carve colored shape structure with laser cutting method according to substrate shape;
S4, by atom transition free radical polymerization reaction by have high-hydrophilic can polyglycol grafting surperficial to cellulose paper, obtain hydrophilic cellulose paper;
S5, be made colored shape put into slide configurations by made of paper for hydrophilic cellulose, obtain functionalization paper substrate micro-fluidic chip, be applied to fluid detection.
Preferably, described colored shape refers to regular snowflake sheet-like morphology.
Preferably, in step S5, adopt laser engraving method that cellulose paper is made into colored shape.
Preferably, in steps A, described variable concentrations is followed successively by: 100mM, 50mM, 25mM, 12.50mM, 6.25mM, 3.12mM.
Preferably, in steps A, before and after the reaction of described application ImageJ software measurement, on each paper substrate micro-fluidic chip conversion zone, the average gray value of the chromogenic reaction of iodine is specially: glucose solution is added drop-wise to test zone, take pictures at distance paper substrate micro-fluidic chip 25cm place with camera at the beginning of reaction, the liquor kalii iodide of the 0.6mM of 10uL is dripped in paper substrate micro-fluidic chip central area, liquor kalii iodide can be flowed into test zone with hydrophobic channel by the capillary action of paper substrate, in same distance, paper substrate micro-fluidic chip is taken pictures again after question response 10min, gained photo of taking pictures twice imports in ImageJ software, calculate the average gray value of conversion zone.
The present invention has following beneficial effect:
(1) micro-fluidic chip has developed into one of state-of-the-art science and technology in the world gradually as core technology wherein.Paper substrate facture of microchip comparatively traditional microfluidic chip manufacturing is simple, easy to use, to the less pollution of environment.The basic operation such as selection, preparation, reaction, detection of the sample involved by the fields such as chemistry and biology is integrated in the paper chip of several square centimeters by paper substrate chip, the capillary action formed by microchannel and entering test zone to control fluid without the need to additionaling power, in order to complete different chemistry or bioprocesses, and its product is analyzed, what it was the biochemical analysis new situation start provides a new research platform.The present invention, by the detection adopting the paper substrate chip of microslide structure hydrophobic channel acquisition to be applied to glucose in fruit innovatively, has certain practical significance.
(2) glucose content in the disposable fruit detecting five kinds of different fruit or Different sources of the paper substrate chip prepared by the present invention, simple to operate, the reaction time is short, is applicable to being applied in the field quick detection of fruit.
Accompanying drawing explanation
By reading the detailed description done non-limiting example with reference to the following drawings, other features, objects and advantages of the present invention will become more obvious:
Fig. 1 is polar plot and the paper substrate chip sample figure of paper substrate chip used.
Fig. 2 is the glucose solution that concentration is respectively 100mM, 50mM, 25mM, 12.5mM, 6.25mM, 3.12mM, when time difference 2min, 4min, 6min, 8min, 10min, and the knots modification of photo gray-scale value before and after the micro-fluidic system reaction of paper substrate.
Fig. 3 is glucose solution paper substrate micro-fluidic chip system gray scale changes values curve and the colorimetric card produced thus when 10min that concentration is respectively 100mM, 50mM, 25mM, 12.5mM, 6.25mM, 3.124mM.
Fig. 4 is the testing result of glucose content in the fruit syrup of five kinds of Common Fruits;
Fig. 5 is five kinds of testing results from glucose content in the cider of the apple of Different sources.
Embodiment
Below in conjunction with embodiment, the present invention is described in detail.Following examples will contribute to those skilled in the art and understand the present invention further, but not limit the present invention in any form.It should be pointed out that to those skilled in the art, without departing from the inventive concept of the premise, can also make certain adjustments and improvements.These all belong to protection scope of the present invention.
The present invention is a kind of surperficial to paper substrate micro-fluidic chip based on polyglycol grafting, obtain functionalization paper substrate micro-fluidic chip, and with paper substrate chip for response matrix, the average gray value of the chromogenic reaction of iodine on the paper substrate of application ImageJ software measurement reaction front and back, make the typical curve of different glucose solution and paper substrate gray scale changes values, and then the glucose in sxemiquantitative fruit, there is following technological process and step:
The preparation of paper substrate micro-fluidic chip substrate: traditional paper substrate micro-fluidic chip produces paper substrate hydrophobic channel by the mode such as photoetching, batik, but these class methods may destroy the hydrophilic nmature of hard copy body, therefore we have developed a kind of novel paper substrate hydrophobic channel, prepare paper substrate micro-fluidic chip as hydrophobic material in conjunction with ordinary filter paper with PDMS (dimethyl silicone polymer), and optimize its process conditions.Its manufacturing process is mainly as follows: (1) modifies allyltrimethoxysilanis on microslide, (2) PDMS is coated by the substrate being carved with colored shape, (3) on microslide, colored shape structure is accurately carved with laser cutting method according to substrate shape, (4) by atom transition free radical polymerization reaction by have high-hydrophilic can polyglycol grafting surperficial to paper substrate micro-fluidic chip, obtain hydrophilic paper substrate; (5) with laser engraving, cellulose paper is made into decorative pattern shape and puts into slide configurations, namely can be applicable to fluid detection.With in the past on cellulose paper direct construction parent/hydrophobic channel method compared with, build hydrophobic channel with microslide and have following advantage: the parent/hydrophobic channel segmentation of paper substrate micro fluidic device obviously, the wall place of the groove of microslide can barrier liquid completely, forms a hydrophobic barrier; After microslide flower shape structure fabrication completes, reusable, only need with changing flower shape paper substrate chip after alcohol washes when reusing; When the method can ensure to repeat to test, the homogeneity of condition, all tests at every turn in same parent/hydrophobic channel.In addition, the paper substrate micro-fluidic chip that traditional parent of direct construction on cellulose paper/hydrophobic channel is formed is applied in the present invention, its detection time and accuracy of detection are much smaller than paper substrate chip of the present invention, its possible cause is that the impurity component producing interference in fruit syrup is more, the parent/hydrophobic channel of direct construction easily blocked or modification and then affect Detection results on cellulose paper.
The preparation of paper substrate micro-fluidic chip paper chip: according to the size of the hydrophobic channel designed and then the size of design paper substrate.By common filter paper, the method for laser engraving obtains the functionalization paper substrate micro-fluidic chip that simultaneously can detect six each and every one samples, and one of them is as blank region, and once experiment can detect the glucose content of five kinds of fruit.
The drafting of standard grayscale value curve of different glucose solution and the preparation of colorimetric card: the paper substrate chip after cutting through laser engraving machine does chemical modification to detect the glucose in fruit, brown can be presented based on the elemental iodine generating different quality on paper substrate to change to auburn color, the glucose content in measurable fruit.The glucose solution of preparation in early stage variable concentrations, comprise the glucose solution of 100mM, 50mM, 25mM, 12.50mM, 6.25mM, 3.12mM, produce the typical curve of average gray value knots modification calculated through ImageJ corresponding to different concentration of glucose and the colorimetric card by different shade of gray.Can find out that from data glucose solution is added drop-wise to paper substrate chip reaction upper 10 minute Absorbable organic halogens, according to the colour developing degree of paper substrate, contrast colorimetric card and gray-scale value knots modification get final product the amount of contained glucose in this fruit of sxemiquantitative.
The embody rule of the glucose content that method of the present invention detects in fruit is shown in following embodiment:
embodiment 1
Application paper substrate micro-fluidic chip detects the content of glucose in five kinds of Common Fruits, its process and step as follows:
By five kinds of peelings and corning of fruits not of the same race, retain edible portion, pulp is squeezed into juice by juice extractor, the fruit juice that liquid-transfering gun measures 0.6 μ L variety classes fruit is respectively added drop-wise to test zone, take pictures at distance paper substrate chip 25cm place with camera at the beginning of reaction, the liquor kalii iodide of 10uL (0.6mM) is dripped in paper substrate chip center region, liquor kalii iodide can be flowed into test zone with hydrophobic channel by the capillary action of paper substrate, in same distance, paper substrate chip is taken pictures again after question response 10min, gained photo of taking pictures twice imports in ImageJ software, calculate the average gray value of conversion zone, calculate the knots modification of the average gray value of conversion zone before and after reaction, different content of planting glucose in fruit can be measured by semidefinite by contrast standard curve and colorimetric card.
embodiment 2
Application paper substrate micro-fluidic chip detects the content of glucose in Different sources apple, its process and step as follows:
By the apple peel of five kinds of Different sources, retain edible portion, pulp is squeezed into juice by juice extractor, liquid-transfering gun measures 0.6 μ L, five kinds of ciders from Different sources apple and is added drop-wise to test zone, react first camera to take pictures at distance paper substrate 25cm place, the liquor kalii iodide of 10uL (0.6mM) is dripped in paper substrate chip center region, liquor kalii iodide can be flowed into test zone with hydrophobic channel by the capillary action of paper substrate, can reach stable after question response 10min, again take pictures at same distance paper substrate 25cm place, in the photo input ImageJ software that twice is taken, measure the average gray value of two photo test zones, calculate the knots modification of the average gray value of two same test zones of photo, the curve of the gray scale changes values of corresponding colorimetric card and Standard glucose solution, get final product the amount of glucose in the apple of sxemiquantitative Different sources source.
To the detection of gained sample in above-described embodiment and the result of experiment see Fig. 1 ~ 3 in accompanying drawing.
The flakes paper substrate chip of Fig. 1 (a) designed by the present invention is at the polar plot of AutoCad software, and the test zone diameter of design is 5.09mm, the long 8.09mm of hydrophobic channel, wide 2mm; Fig. 1 (b) is the paper substrate sample drawing of cellulosic filter paper after laser engraving machine cutting.
1. see accompanying drawing 2, as can be seen from the figure solubility is respectively the glucose solution of 100mM, 50mM, 25mM, 12.5mM, 6.25mM, 3.12mM when the reaction time is 2min, 4min, 6min, 8min, 10min, take pictures at distance paper substrate 25cm place and get figure, by ImageJ software measuring and calculating gray-scale value, and calculate average gray changes values, go out curve with Origin software development, experiment in triplicate.
See accompanying drawing 3, the glucose solution that Fig. 3 (a) is respectively 100mM, 50mM, 25mM, 12.5mM, 6.25mM, 3.124mM for concentration is reacting 10min, take pictures before and after the reaction of distance paper substrate chip 25cm place and get figure, ImageJ software calculates the average gray value of conversion zone, calculate its gray scale changes values, be X-axis with Origin software development with reaction density, the knots modification of the average gray value of test zone is the curve of Y-axis, and experiment in triplicate; Fig. 3 (b) is the colorimetric card of the large little makings according to the average gray changes values on Fig. 3 (a) typical curve, it is bronzing in colorimetric card, by comparing the depth of color (in figure from left to right, color shoals successively), thus draw the difference of glucose content in various fruit; This colorimetric card and typical curve can be used for Site Detection and the amount of glucose in sxemiquantitative fruit.
Fig. 4 is the testing result of glucose content in five kinds of Common Fruits (mangosteen, apple, muskmelon, white grape, lichee).As shown in Figure 4, the glucose content testing result of common commercially available fruit is known, and lichee is higher containing glucose amount.
Fig. 5 is the testing result of glucose content in the apple in five kinds of Different sources (Hebei, Gansu, Shanxi, Shandong, Shaanxi).As shown in Figure 5, the glucose content of Different sources apple there are differences, and wherein the apple in Luochuan, Shaanxi is relatively high containing grape.
Above specific embodiments of the invention are described.It is to be appreciated that the present invention is not limited to above-mentioned particular implementation, those skilled in the art can make various distortion or amendment within the scope of the claims, and this does not affect flesh and blood of the present invention.
Claims (7)
1. utilize a method for glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit, it is characterized in that, described method comprises the steps:
The Glucose standards solution of A, preparation variable concentrations, respectively with paper substrate micro-fluidic chip for detection platform, the average gray value of the chromogenic reaction of iodine on each paper substrate micro-fluidic chip conversion zone in application ImageJ software measurement reaction front and back, make the typical curve of average gray value knots modification corresponding to different concentration of glucose, and the colorimetric card of different shade of gray;
B, squeezed the juice by fruit to be measured, with paper substrate micro-fluidic chip for detection platform, before and after the reaction of application ImageJ software measurement, the average gray value of the chromogenic reaction of iodine on paper substrate micro-fluidic chip conversion zone, calculates the average gray value knots modification that fruit to be measured is corresponding; Contrast described colorimetric card and typical curve, get final product the glucose content in sxemiquantitative fruit to be measured.
2. the method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit according to claim 1, it is characterized in that, described paper substrate micro-fluidic chip will have the polyglycol grafting of high-hydrophilic energy to paper substrate micro-fluidic chip surface, the functionalization paper substrate micro-fluidic chip of acquisition by the method for atom transition free radical polymerization reaction.
3. the method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit according to claim 2, it is characterized in that, the preparation of described paper substrate micro-fluidic chip specifically comprises the steps:
S1, on microslide, modify allyltrimethoxysilanis;
S2, coat PDMS by the substrate being carved with colored shape;
S3, on microslide, accurately carve colored shape structure with laser cutting method according to substrate shape;
S4, by atom transition free radical polymerization reaction by have high-hydrophilic can polyglycol grafting surperficial to cellulose paper, obtain hydrophilic cellulose paper;
S5, be made colored shape put into slide configurations by made of paper for hydrophilic cellulose, namely can be applicable to fluid detection.
4. the method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit according to claim 3, it is characterized in that, described colored shape refers to regular snowflake sheet-like morphology.
5. the method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit according to claim 3, is characterized in that, in step S5, adopts laser engraving method that cellulose paper is made into colored shape.
6. the method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit according to claim 1, it is characterized in that, in steps A, described variable concentrations is followed successively by: 100mM, 50mM, 25mM, 12.50mM, 6.25mM, 3.12mM.
7. the method utilizing glucose content in paper substrate micro-fluidic chip method rapid semi-quantitative fruit according to claim 1, it is characterized in that, in steps A, before and after the reaction of described application ImageJ software measurement, on each paper substrate micro-fluidic chip conversion zone, the average gray value of the chromogenic reaction of iodine is specially: glucose solution is added drop-wise to test zone, take pictures at distance paper substrate micro-fluidic chip 25cm place with camera at the beginning of reaction, the liquor kalii iodide of the 0.6mM of 10uL is dripped in paper substrate micro-fluidic chip central area, by the capillary action of paper substrate, liquor kalii iodide is flowed into test zone with hydrophobic channel, in same distance, paper substrate micro-fluidic chip is taken pictures again after question response 10min, gained photo of taking pictures twice imports in ImageJ software, calculate the average gray value of conversion zone.
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| CN106442491A (en) * | 2016-09-07 | 2017-02-22 | 华东理工大学 | Method for rapidly detecting urine sugar and uric acid in solution |
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| CN106525840A (en) * | 2016-12-18 | 2017-03-22 | 福建神采新材料科技有限公司 | Annular Zhukun sodium copper chlorophyllin detection contrast disk color chart |
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| CN107894416A (en) * | 2017-12-05 | 2018-04-10 | 上海海洋大学 | A kind of method using nutrient composition content in mobile phone and functionalization paper substrate micro-fluidic chip detection fruit |
| CN109856193A (en) * | 2019-01-30 | 2019-06-07 | 浙江工业大学 | A kind of paper substrate micro-fluidic intelligent chip and system for examination of glucose concentration |
| CN111208184A (en) * | 2020-02-03 | 2020-05-29 | 盐城瑞力达科技有限公司 | Non-enzymatic glucose sensing and detecting system |
| CN111595842A (en) * | 2020-05-20 | 2020-08-28 | 中国科学院新疆理化技术研究所 | Preparation method of array type chemical colorimetric sensing chip for detecting explosives and drugs |
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