KR100396093B1 - Method and apparatus for analysing oxidation number 3 and 6 of chromium simultaneously - Google Patents

Abstract

The present invention relates to a method for analyzing chromium trivalent and chromium hexavalent, and more particularly, to a method for precisely real-time analysis of chromium trivalent and chromium hexavalent simultaneously at low cost by using a flow injection method and a chemiluminescence detection method. will be.

Therefore, the present invention, the step of mixing the chromium trivalent and chromium hexavalent mixed sample and water, and mixing the chemiluminescent reagent and oxidizing agent (S1); Transferring the mixed liquid of the mixed sample and water and transferring the mixed liquid of the chemiluminescent reagent and oxidant (S2); While each of the transferred mixed solution is mixed, the chromium trivalent and the chemiluminescent reagent reacts to emit light (S3); Detecting the generated light to quantify chromium trivalent value (S4); Discharging the sample and the reagent after the reaction (S5); Mixing a reducing agent instead of water in the mixing step, and repeating the transfer step (S2), the light emitting step (S3), the detecting step (S4), and the discharging step (S5) (S6 to S8); It provides a method for the simultaneous analysis of chromium trivalent and chromium hexavalent comprising the step of quantifying chromium hexavalent by comparing the amount of chromium trivalent quantified in the detection step, the lab-on-a-chip technology It is manufactured based on the present invention to provide a micro analysis substrate that can perform the method.

Description

Method for simultaneous analysis of chromium trivalent and chromium hexavalent and analytical substrate {METHOD AND APPARATUS FOR ANALYSING OXIDATION NUMBER 3 AND 6 OF CHROMIUM SIMULTANEOUSLY}

The present invention relates to a method for analyzing chromium trivalent and chromium hexavalent, and more specifically, it is possible to precisely analyze chromium trivalent and chromium hexavalent at the same time at a low cost in a short time in real time using a flow injection method and chemiluminescence detection method. And chromium trivalent and chromium hexavalent methods.

In general, chromium is an element of soil, volcanic ash and volcanic gas, which is a heavy metal. Chromium exists in nature in three states, unionized with chromium atoms, in the form of chromium trivalent or chromium hexavalent. Chromium trivalent is a form present in nature and unionized chromium and chromium hexavalent are generally produced in industrial processes. Chromium is mainly used to make iron alloys and other alloys. Chromium compounds produced in chemical plants are used for chromium plating, pigmentation, leather processing and wood processing.

Chromium hexavalent is irritating and can cause ulceration on the skin, short time exposure to high concentrations, irritating the nasal mucosa, and irritating the stomach. It also affects the kidneys and liver, and the US Department of Health and Human Services defines chromium hexavalent as a carcinogen. Chromium trivalents, on the other hand, do not have the same effect as chromium hexavalent and, rather, act as essential nutrients when consumed in moderation. Chromium in food is usually in the form of chromium trivalent. Exposure to unionized chromium is not common and its health effects are not well known.

Therefore, whether chromium exists in an unionized state, trivalent ionized or hexavalent is of great interest.

In general, as a method for analyzing chromium trivalent and chromium hexavalent, methods using ultraviolet spectroscopy, redox titration potentiometric analysis, and ion-selective electrodes are largely used.

In the case of chromium hexavalent analysis using ultraviolet spectroscopy, the absorption peak at 350 nm is measured when the concentration of chromium hexavalent is 100 ppm or more, and then analyzed using Beer's law, and diphenyl is less than 100 ppm. Carbazide (Diphenylcarbazide) was used to form a red complex and analyzed using a peak of 540 nm. In addition, when analyzing chromium trivalent, there is no distinct peak of its own, so oxidizing chromium trivalent to chromium hexavalent using a strong oxidizing agent, and then analyzing the same method as that of chromium hexavalent is used. come.

However, when analyzing the chromium trivalent and chromium hexavalent by this method, it is difficult to analyze at the same time, there is a problem such that it takes a lot of time to analyze the chromium trivalent and chromium hexavalent.

Redox titration potentiometric analysis is large and reproducible and not affected by concentration. However, it is difficult to automate the oxidation-reduction titration process, and the method using ion-selective electrode can be easily applied to the automated process. The problem is that the service life is short and considerable care must be taken in the management itself.

In addition, there are absorbance spectroscopy, atomic absorption spectroscopy, etc., but these methods have a large analytical device, require a lot of time and labor for the analysis of chromium trivalent and chromium hexavalent, and difficult real-time analysis in the field.

For real-time chromium analysis in the field, absorbance photometry using flow injection method was developed, but chromium trivalent and chromium hexavalent cannot be analyzed at the same time, many samples are required in preparation for analysis, and detection limit is 1000ppm. It is a problem that the precision is low because it does not satisfy the water quality standard 50ppb and the process control standard 500ppb to about 1ppm.

Ion Chromatography and Atomic Absorption Spectroscopy have been introduced to analyze chromium in concentrations as low as a few ppb. However, due to the large size of the analyzer, it is not portable and cannot be analyzed in the field in real time. There is a problem such as a long time to analyze.

Korean Patent Publication No. 1992-12903 describes a method for simultaneously analyzing chromium trivalent and chromium hexavalent using fluorescent X-ray analysis, but it is difficult to carry because the equipment is expensive and the device is large and it is difficult to carry in the field in real time. There is a problem in analyzing chromium trivalent and chromium hexavalent simultaneously.

The present invention was devised to solve the above problems, and its purpose is to simultaneously analyze chromium trivalent and chromium hexavalent at the same time at a low cost in real time in the field using a flow injection method and chemiluminescence detection method together. It is to provide a method for simultaneous analysis of chromium trivalent and chromium hexavalent.

Another object of the present invention is to provide a substrate for simultaneous analysis of chromium trivalent and chromium hexavalent using the above method.

1 is a plan view showing a substrate for simultaneous analysis of chromium trivalent and chromium hexavalent according to the present invention.

2 is a view showing a process for preparing a substrate for simultaneous analysis of chromium trivalent and chromium hexavalent according to the present invention.

3 is a view showing a device for the simultaneous analysis of chromium trivalent and chromium hexavalent in accordance with the present invention.

Figure 4 is a graph showing the chemiluminescence intensity according to the concentration of chromium trivalent.

5 is a graph showing chemiluminescence intensity according to chromium hexavalent concentration.

Explanation of symbols on the main parts of the drawings

10: Analysis substrate 13: First channel

16: second channel 17: third channel

18: discharge pipe 26: PDMS plate

27: glass plate 28: Tygon tubing

31: pulsation pump 34: Teflon tubing

35: Photomultiplier tube (PMT)

In order to achieve the object as described above, the present invention is a method for analyzing chromium trivalent and chromium hexavalent,

(a) mixing chromium trivalent and chromium hexavalent mixed samples and water through a first channel, and mixing a chemiluminescent reagent and an oxidant through a second channel (S1); (b) transferring the mixed solution of the mixed sample and water along the first channel and transferring the mixed solution of the chemiluminescent reagent and oxidant along the second channel (S2); (c) mixing the respective mixed liquids transferred in the transfer step through the third channel, wherein the chemiluminescent reagent and the oxidant react with chromium trivalent as a catalyst to emit light (S3); (d) quantifying chromium trivalent by detecting light generated in the light emitting step (S4); (e) discharging the sample and the reagent after reacting in the light emitting step (S5); (f) incorporating a chromium trivalent and chromium hexavalent mixed sample and a reducing agent for reducing chromium hexavalent chromium through the first channel and incorporating a chemiluminescent reagent and an oxidant through the second channel (S6). ; (g) reacting the mixed solution of the mixed sample and the reducing agent along the first channel and reacting to reduce the chromium hexavalent to chromium trivalent, and transferring the mixed solution of the chemiluminescent reagent and the oxidant along the second channel (S7). ); (h) repeating steps (c), (d) and (e) (S8); (i) it provides a method for the simultaneous analysis of chromium trivalent and chromium hexavalent comprising the step (S9) of quantifying chromium hexavalent by comparing the amount of chromium trivalent quantified in (d) and (h), respectively.

In addition, the present invention provides a substrate in which a channel is formed to analyze chromium trivalent and chromium hexavalent,

A first branch into which a sample containing a mixture of chromium trivalent and chromium hexavalent is injected, a second branch into which water or a reducing agent is selectively injected, the first branch and a second branch are formed into one, and the mixed sample and the water or reducing agent are transferred. Sample inlet including a first channel to react while being; Reagent inflow comprising a third branch into which a chemiluminescent reagent is injected, a fourth branch into which an oxidant is injected, a third branch where the third branch and the fourth branch meet to form one, and reacting while the chemiluminescent reagent and the oxidant are transported. part; A light emission reaction unit including a third channel where the first channel and the second channel meet and are formed as one, and each of the mixed liquids transferred through the first channel and the second channel reacts to emit light; It is connected to the third channel, and provides a substrate for simultaneous analysis of chromium trivalent and chromium hexavalent comprising an outlet including a discharge tube for discharging the mixed solution of the sample and the reagent after the reaction.

In particular, the substrate used to implement the analysis method according to the present invention is a lab-on-a-chip technology for forming microchannels in micro units using MEMS (Micro Electro Mechanical Systems) technology. It is possible to realize an ultra-compact analysis device by manufacturing based on.

Hereinafter, the present invention will be described in more detail with reference to the accompanying drawings.

1 is a plan view showing a substrate for simultaneous analysis of chromium trivalent and chromium hexavalent according to the present invention.

First, a sample mixed with chromium trivalent and chromium hexavalent is mixed through the first channel 13, and a chemiluminescent reagent and an oxidant are mixed through the second channel 16 (S1).

As shown in FIG. 1, a first branch 11 and a second branch 12 are connected to the first channel 13 so that the sample and water mixed with chromium trivalent and chromium hexavalent are different from each other. It is injected through a branch and mixed in the first channel 13.

Meanwhile, the third branch 14 and the fourth branch 15 are connected to the second channel 16 so that the chemiluminescent reagent and the oxidant are injected through different branches, respectively, and the second channel ( 16) are mixed.

As the chemiluminescent reagent injected in the step, luminol (luminol: 5-amino-2,3-dihydrophthalazine-1,4-dione), lucigenin (lucigenin: bis-N-methylacridinium), lophine (2, 4,5-triphenylimidazole), pyrogallol: 1,2,3-benzenetriol, gallic acid (3,4,5-trihydroxybenzoic acid) and the like can be preferably used.

In addition, as the oxidant injected in the above step, hydrogen peroxide, hypochlorite ions, ferricyanide, and the like may be preferably used.

Next, the mixed solution of the mixed sample and water is transferred along the first channel 13, and the mixed solution of the chemiluminescent reagent and the oxidant is reacted while being transferred along the second channel 16 (S2).

In this step, a pulsation pump, a plunger pump, a double-plunger pump, and a syringe pump are used to transfer a mixed solution of chromium trivalent and chromium hexavalent samples and water along the channel. (syringe pump). The pump is also used for the transfer of the chemiluminescent reagent and oxidant mixture.

Next, the mixed liquids respectively transferred in the transfer step S2 are mixed through the third channel, and the chemiluminescent reagent and the oxidant react with the chromium trivalent catalyst to emit light (S3).

Here, in the case where luminol is used as the chemiluminescent reagent and hydrogen peroxide is used as the oxidizing agent, the mixing reaction in which chromium trivalent is a catalyst is as follows.

Scheme 1

Luminol ^{_{+ 2H 2 O 2 + 2OH -}} + Cr 3+ ( catalyst) → 3- amino phthalate _{+ N 2 + 4H 2 O +} hυ

Next, the chromium trivalent is quantified by detecting the light generated in the light emitting step S3 (S4).

Light generated in Reaction Scheme 1 is detected using any one of a detection device such as a photomultiplier tube (PMT), a photodiode, a CD camera, and the like. That is, the detection device is attached to correspond to the third channel 17 of the substrate 10 to detect light.

Finally, the sample and reagent after the reaction in the light emitting step (S3) is discharged through the discharge pipe (S5).

Here, only chromium trivalent is quantified in the mixed solution of chromium trivalent and chromium hexavalent, and chromium hexavalent is not quantified.

To quantify chromium hexavalent, the above steps are repeated. However, unlike the above step, a reducing agent is injected instead of water.

That is, a sample in which chromium trivalent and chromium hexavalent are mixed and a reducing agent for reducing chromium hexavalent to chromium trivalent are mixed through the first channel 13, and a chemiluminescent reagent and an oxidant are mixed through the second channel (S6). ).

As in the mixing step (S1), the first channel 13 is connected to meet the first branch 11 and the second branch 12, chromium trivalent and chromium hexavalent mixed sample and chromium 6 Reducing agents for reducing valent trivalent to chromium trivalent are injected through different branches, respectively, and are mixed in the first channel 13.

The reducing agent for reducing chromium hexavalent to chromium trivalent includes potassium sulfite, sodium metabisulfite, glycolic acid, α-hydroxycarboxylate and α-carbonyl Carboxylate (alpha-carbonylcarboxylate) etc. can be used preferably.

Next, while the mixed solution of the mixed sample and the reducing agent is transported along the first channel 13, the chromium hexavalent is reduced to chromium trivalent, and the mixed solution of the chemiluminescent reagent and the oxidant reacts with the second channel 16. Reacts while being transported along (S7).

When potassium sulfite (K ₂ SO ₃ ) is used as the reducing agent, chromium hexavalent is reduced to chromium trivalent by the following reaction scheme.

Scheme 2

Cr ₂ O ₇ ^2- + 3K ₂ SO ₃ + 8H ⁺ → 2Cr ³⁺ + 6K ⁺ + 3SO ₄ ^2- + 4H ₂ O

In Scheme 2, the oxidation number of chromium of Cr ₂ O ₇ ^2- is 6. Reaction with potassium sulfite as the reducing agent reduces the number of chromium oxides to 3 and reduces the chromium to trivalent.

Next, while repeating the light emitting step (S3), the detection step (S4) and the discharge step (S5), it is possible to quantify the chromium trivalent from chemiluminescence (S8).

Finally, chromium hexavalent is quantified by comparing the amount of chromium trivalent quantified in the detection step (S4) with the amount of chromium trivalent quantified in the repetition step (S8) (S9).

Thus, through the steps (S1 ~ S9) can be analyzed at the same time chromium trivalent and chromium hexavalent, and also by continuous control the timing of injection of the reducing agent can also be a continuous analysis.

In particular, the present invention can miniaturize the analysis apparatus provided for the analysis according to the above method.

In other words, lab-on-a-chip, which applies MEMS (Micro Electro Mechanical Systems) technology to the bio field, which can process ultra-high density integrated circuits and micro gears by processing silicon, quartz, and silica. Based on on-a-chip technology, a substrate with fine channels is fabricated. The lab-on-a-chip, also called a microfluidics chip, allows the analysis of reactions with various biomolecules or sensors integrated in the chapter while flowing a small amount of analyte. Say Chip.

The configuration of an analytical substrate on which channels are formed to analyze chromium trivalent and chromium hexavalent will be described in detail with reference to the drawings.

The substrate for simultaneous analysis of chromium trivalent and chromium hexavalent according to the present invention, the sample inlet is formed to enter the sample to be largely analyzed, the reagent inlet is formed to introduce the reagent for analyzing the sample, And a light emitting reaction unit for generating light by reacting the sample and the reagent and a discharge unit for discharging the sample and the reagent after the reaction.

The sample inlet may include a first branch 11 into which a sample containing chromium trivalent and chromium hexavalent is injected, a second branch 12 into which water or a reducing agent is selectively injected, the first branch 11 and a second branch ( 12) is formed to meet and includes a first channel 13 to react while the mixed sample and water or reducing agent is transported.

In addition, a sample container 1 and a water or reducing agent container 2 are connected to ends of the first branch 11 and the second branch 12, respectively.

The reagent inlet is formed by forming a third branch 14 into which a chemiluminescent reagent is injected, a fourth branch 15 into which an oxidant is injected, and a third branch 14 and a fourth branch 15 into one. A second channel 16 through which reagents and oxidants are conveyed.

In addition, the chemiluminescent reagent vessel 4 and the oxidant vessel 5 are connected to the end portions of the third branch 14 and the fourth branch 15, respectively.

On the other hand, the light emitting reaction unit is formed by the first channel 13 and the second channel 16 is formed as one, and the mixed liquid transferred through the first channel 13 and the second channel 16 The mixed solution includes a third channel 17 that reacts and emits light.

In addition, the third channel 17 is installed so that all of the light emission range is connected to a detection device such as a photomultiplier tube (PMT).

The length of the third channel 17 preferably has a range of 1 μm to 100 cm. If the length is less than 1 μm, there is a problem that the production of the analysis substrate 10 itself is difficult, which is not preferable. If the length exceeds 100 cm, the reagent consumption is large, the transport of the sample is slow, and the analysis speed is slow. It is not desirable.

The discharge portion is connected to the third channel 17, and includes a discharge tube for discharging the mixed solution of the sample and the reagent after the reaction, and the discharge vessel 8 is connected to the end of the discharge tube 18.

Here, the widths and depths of the branches 11, 12, 14, and 15 and the channels 13, 16, and 17 are preferably 100 nm to 10 mm. If the width and depth is less than 100 nm, it is not preferable that the production of the analytical substrate itself is difficult. If the width and depth exceed 10 mm, the consumption of reagents is large, the transport of samples is slow, and the analysis speed is also slow. It is not desirable.

In addition, the analysis substrate 10 in which the branches and channels are formed may be manufactured by selecting one or more materials such as rubber, silicone rubber, plastic, glass, silica, and the like. ), Any one of methods such as etching, molding, pressing, machining, and laser processing can be used.

Hereinafter, a process in which the analytical substrate 10 is manufactured by a molding method using polydimethylsiloxane (PDMS) among the above-described methods will be described.

2 is a view showing a process for preparing a substrate for simultaneous analysis of chromium trivalent and chromium hexavalent according to the present invention.

First, the negative photoresist 21 is coated on the silicon wafer 22. Rubber, silicon-based rubber, plastic, glass, silica and the like may be used as the material of the wafer. There are various methods such as spin coating and dip coating, but the spin coating method is more preferable. In addition, the number of coating is not limited, but is preferably 1 to 2 times. When not coated, there is a problem that the height of the channel is lowered, and when the number of coatings exceeds two times, there is a problem that it is difficult to make the height of the channel constant.

After the negative photoresist 21 is coated on the silicon wafer 22, the photomask 23 is covered and exposed to ultraviolet rays. When the exposed silicon wafer 22 is developed, an embossed mold 24 having a desired pattern is obtained.

PDM polydimethylsiloxane (PDMS) 25 is poured into the finished mold, crosslinked, and then removed to obtain a PDMS plate 26 having a desired intaglio pattern.

The hole for inserting the container is drilled, and the glass plate 27 having a clean surface is attached to the PDMS plate 26 after surface treatment with an arc discharge formed using a Tesla coil. At this time, the bonding through the surface treatment of the PDMS plate using Tesla coil can be made in a glass plate, a silicon wafer and the like.

Finally, Tygon tubing 28 fits snugly into the hole and secured with an adhesive to complete the substrate for simultaneous analysis of chromium trivalent and chromium hexavalent.

3 is a view showing a device for the simultaneous analysis of chromium trivalent and chromium hexavalent in accordance with the present invention.

As shown in FIG. 3, the pulsation pump 31 is equipped with a 9 V battery 33 as a pump driving energy source, and the pulsation pump 31 and the chromium trivalent and chromium hexavalent substrates for simultaneous analysis 10 according to the present invention. ) Is inserted into the Tefron tubing (34) by inserting a capillary tube.

One end of the Teflon tubing 34 is manufactured by fitting a good Tygon tubing with good elasticity in order to fit well with the reservoir (reservoir) of the analysis substrate 10 according to the present invention.

Meanwhile, in order to observe the mixing of the sample solution, the chromium hexavalent reducing agent and the luminescent reagent, a color CCD camera may be installed on the analysis substrate 10 to check the flow of the fluid. Pulsating pumps create pulsations in the sample inlet, which interferes with the stability of the detected signal. Although pulsation cannot be eliminated, it is possible to design the device so that pulsation occurs less. When the formation of bubbles is observed by checking the flow of the fluid with a color CD camera, bubbles may be removed by increasing the flow rate using the pulsation pump 31.

Light generated from the analysis substrate 10 of the present invention may be detected by a photomultiplier tube (PMT) 35.

EXAMPLE

In order to carry out the analytical method according to the present invention, an analytical substrate 10 was prepared and connected with other analytical devices, from which chromium trivalent and chromium hexavalent were quantified.

First, in order to manufacture the analytical substrate 10, the negative photoresist 21 SU-8 is coated on the silicon wafer 22 twice with a height of 100 to 150 µm, and the photomask 23 is covered with ultraviolet rays. After exposure, the silicon wafer 22 was developed to form a pattern.

The polydimethylsiloxane (25) was poured into the patterned embossed frame (24), cross-linked, and then removed, thereby obtaining a PDMS plate (26) having a desired engraved pattern. After the surface treatment with the formed arc discharge, the glass plate 27 whose surface was cleaned was attached to the PDMS plate 26.

The Tygon tubing 28 was cut to a length of 5 mm, fitted with holes, and fixed with an adhesive to complete a 2.5 cm × 5.0 cm plastic analysis substrate 10.

Next, a teflon having a diameter of 0.3 mm, an outer diameter of 1.58 mm, and a length of 20 cm was inserted into a pulsation pump 31 driven by a 9V dry battery 33 to insert a capillary tube having an outer diameter of 0.3 mm and an inner diameter of 95 µm. The tubing 34 was connected to the analytical substrate 10 of the present invention, and one side of the Teflon tubing 34 was fitted with a good elasticity Tygon tubing having an internal diameter of 1.58 mm to fit the reservoir of the chip well.

Light generated by the chemical reaction in the analysis substrate 10 of the present invention was detected by the photomultiplier tube 35. Instrument control and data processing were controlled with a computer 36 using a Lap view 5.0 program and a DAQ Card-1200 multifunction I / O card.

Chromium chloride hexahydrate and potassium chromate were dissolved in distilled water to prepare a sample in which chromium trivalent and chromium hexavalent were mixed. As a chemiluminescent reagent, a boric acid buffer solution of pH 11.5 was used. 0.2 M luminol and 6.8 × 10 ⁻³ M hydrogen peroxide were used. As a chromium hexavalent reducing agent, 0.015 M gallium sulfate having a pH of 3.0 was used. Luminescence was measured by flowing at a rate of / min.

The quantitative determination of chemiluminescence intensity was measured while changing the concentration of chromium trivalent to 10, 50, 100, 250, 500, and 1000 ppb, which is shown in FIG. 4.

Although the chemiluminescence intensity did not increase linearly as the concentration increased, it was confirmed that linearity was obtained by taking the logarithm of the detected value. At this time, the linear concentration range was 50 to 500 ppb, and the detection limit was about 10 ppb.

In addition, the quantitative determination of chemiluminescence intensity was measured while changing the concentration of chromium hexavalent to 10, 50, 100, 250, 500, and 1000 ppb, which is shown in FIG. 5.

At this time, the linear concentration range was 10 to 1000 ppb, and the detection limit was about 1.3 ppb.

The graphs shown in FIGS. 4 and 5 are calibration curves showing quantitative quantities, and the concentrations can be predicted by comparing the intensity of light generated in an actual sample with the graphs.

Although the preferred embodiments of the present invention have been described above, the present invention is not limited thereto, and various modifications and changes can be made within the scope of the claims and the detailed description of the invention and the accompanying drawings. Naturally, it is within the scope of the present invention.

According to the simultaneous analysis method of chromium trivalent and chromium hexavalent according to the present invention, by producing a micro-analytical substrate, by using a flow injection method and a chemiluminescence detection method, in real time in the field at a low cost in a short time with a chromium trivalent It is effective to analyze chromium hexavalent simultaneously at the same time.

In addition, by applying such an analytical substrate to the inflow and discharge of steel plants, chromium plating plants, etc., automatic water quality measurement network, portable measurement equipment, etc., it contributes to the small size and light weight of the equipment, and enables the real-time analysis. There is.

Claims (11)

In the method of analyzing chromium trivalent and chromium hexavalent, (a) incorporating a chromium trivalent and chromium hexavalent mixed sample and water through a first channel and incorporating a chemiluminescent reagent and an oxidant through a second channel; (b) transferring the mixed liquid of the mixed sample and water along the first channel and transferring the mixed liquid of the chemiluminescent reagent and oxidant along the second channel; (c) mixing the respective liquids transferred in the transfer step through the third channel, and causing the chemiluminescent reagent and the oxidant to react with the chromium trivalent catalyst to emit light; (d) quantifying chromium trivalent by detecting light generated in the light emitting step; (e) discharging the sample and reagent after reacting in the light emitting step; (f) incorporating a sample mixed with chromium trivalent and chromium hexavalent through the first channel and a reducing agent for reducing chromium hexavalent to chromium trivalent and incorporating a chemiluminescent reagent and an oxidant through the second channel; (g) reacting the mixed solution of the mixed sample and the reducing agent along the first channel to react to reduce the chromium hexavalent to chromium trivalent and to transfer the mixed solution of the chemiluminescent reagent and the oxidant along the second channel; (h) repeating steps (c), (d) and (e); (i) quantifying chromium hexavalent by comparing the amounts of chromium trivalent respectively quantified in (d) and (h) Simultaneous analysis method of chromium trivalent and chromium hexavalent. The method of claim 1, Step (b) and (g) is a chromium trivalent and chromium hexavalent simultaneous analysis method characterized in that the mixed liquid is transferred using a pulsation pump, a plunger pump, a double-plunger pump or a syringe pump. The method of claim 1, The chemiluminescent reagent is a method for simultaneous analysis of chromium trivalent and chromium hexavalent, characterized in that selected from the group consisting of luminol, lucigenin, ropin, pyrogallol and gallic acid. The method of claim 1, Said oxidant is selected from the group consisting of hydrogen peroxide, hypochlorite ions and ferricyanide. The method of claim 1, The reducing agent is a method of simultaneous analysis of trivalent and chromium hexavalent, characterized in that selected from the group consisting of potassium sulfite, sodium metabisulfite, glycolic acid, α-hydroxycarboxylate and α-carbonylcarboxylate. . In a substrate where a channel is formed to analyze chromium trivalent and chromium hexavalent, A first branch into which a sample containing chromium trivalent and chromium hexavalent is injected, a second branch into which water or a reducing agent is selectively injected, the first branch and a second branch meet and are formed as one, and the mixed sample and water or reducing agent are transported. Sample inlet including a first channel to react while being; Reagent inflow comprising a third branch into which a chemiluminescent reagent is injected, a fourth branch into which an oxidant is injected, a third branch where the third branch and the fourth branch meet to form one, and reacting while the chemiluminescent reagent and the oxidant are transported. part; A light emission reaction unit including a third channel where the first channel and the second channel meet and are formed as one, and each of the mixed liquids transferred through the first channel and the second channel reacts to emit light; A discharge part connected to the third channel and including a discharge pipe through which the mixed solution of the sample and the reagent after the reaction is discharged; A substrate for the simultaneous analysis of chromium trivalent and chromium hexavalent. The method of claim 6, The analysis substrate is a chromium trivalent and chromium hexavalent simultaneous analysis substrate, characterized in that made of at least one selected from the group consisting of rubber, silicone rubber, plastic, glass and silica. The method of claim 6, The branch and the channel is a substrate for simultaneous analysis of chromium trivalent and chromium hexavalent, characterized in that the width ranges from 100nm to 10mm. The method of claim 6, The branch and the channel is a substrate for simultaneous analysis of chromium trivalent and chromium hexavalent, characterized in that the depth ranges from 100nm to 10mm. The method of claim 6, The chromium trivalent and chromium hexavalent chromium substrate for simultaneous analysis, characterized in that the third channel of the emission reaction portion has a length in the range of 1㎛ to 100cm. The method of claim 6, The analysis substrate is a chromium trivalent and chromium hexavalent simultaneous analysis substrate, characterized in that produced by any one method of etching, molding, pressing, machining and laser processing.