A dual-range responsive pH optical probe

Abstract

A dual-range response pH optical probe, the absorption spectrum of hematoporphyrin monomethyl ether shows a dual-response characteristic to the medium pH, that is, hematoporphyrin monomethyl ether can respond to the pH in two different ranges, so that the determination of pH in different ranges can be realized by using a single probe, that is, 'one needle two uses'. Hematoporphyrin monomethyl ether also has a ratio response characteristic to pH. The ratio type pH optical probe can overcome the influence of many non-pH factors. The present application utilizes the photometric dual-range response characteristic of hematoporphyrin monomethyl ether to pH to establish a new method for determining pH by using hematoporphyrin monomethyl ether as an optical probe. The present application is used for the determination of pH of practical samples such as small-volume samples, viscous samples, food samples with large acidity span, biological samples and environmental water samples, and satisfactory results are obtained, which shows the practical application advantages of the pH optical probe and opens up the application of light absorption type porphyrin compounds in analytical science.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of spectrophotometry, in particular to a dual-interval response pH optical probe. BACKGROUND

[0002] Acidity determination is an important daily work [Liu Q., et al., Food Analytical Methods, 2017, 10(1): 111-117]. In the field of environmental protection, pH is an important indicator of water quality, which can characterize specific pollution [El-Gendy N.S., et al., Desalination and water treatment, 2016, 57(12): 5514-5528]. In medicine, the pH of human blood and urine is usually maintained within a certain range, and if it fluctuates, it is a pathological phenomenon [Dunn W.J., et al., Australian Critical Care, 2016, 29(1): 41-45.]. Therefore, pH monitoring is an important practical work in the fields of food, environmental health, medical clinic, etc. [Safavi A., et al., Analytica Chimica Acta, 1998, 367(1-3): 167-173].

[0003] Visual colorimetry and glass electrode method are the conventional methods for pH determination. The visual observation method is insufficient in accuracy and precision, and the glass electrode method has many difficulties in the determination of small volume samples and high viscosity samples. The optical probe pH determination method based on the principle of molecular spectroscopy has been widely used due to its high sensitivity, good selectivity, wide application range, and less reagent consumption. In recent years, the development and research of new probes and new methods have been highly valued [Si Y., et al., Biosens & Bioelectronics, 2016, 75(1): 320-327]. The pH optical probe refers to a substance whose absorption or fluorescence characteristics have a significant correlation with the pH of the system, and thus can be used to indicate the acidity and alkalinity of the medium. The determination of pH is realized by the change of fluorescence or absorption intensity or spectrum of the optical probe within a certain pH range. The optical probe is usually stable in nature, easy to apply, and has high sensitivity, and can observe the pH change in time and space. However, the conventional pH optical probe can only respond to the pH in one interval. SUMMARY

[0004] The present application aims to solve the above-mentioned problems in the prior art, and provides a pH determination method based on an optical probe that can respond to the acidity in two different intervals. This method can realize the determination of pH in different intervals with a single probe, and has the effect of "one needle for two uses".

[0005] To achieve the above object, the present application adopts the following technical solutions:

[0006] 1) adding blood porphyrin monomethyl ether aqueous solution with fixed concentration and volume into a series of containers;

[0007] 2) adding buffer solution with different pH values in sequence into the above series of containers, and setting the volume;

[0008] 3) scanning the absorption spectrum of the pH solution system, and measuring the absorbance at the absorption peak or a pair of wavelengths.

[0009] The response range of the blood porphyrin monomethyl ether to pH includes two ranges of pH 1.89-4.05 and pH 4.99-7.09.

[0010] Blood porphyrin monomethyl ether aqueous solution with fixed concentration and volume is added into a series of containers in sequence, and the number of the reaction containers is not less than 3; the pH value of the buffer solution falls within the corresponding linear range of the working curve, and the corresponding linear range of the working curve refers to the linear working curve range of the measured pH corresponding to the concentration of the blood porphyrin monomethyl ether aqueous solution used.

[0011] The added blood porphyrin monomethyl ether aqueous solution has a final concentration of 1.0*10 -6 -1.2*10 -5 mol / L.

[0012] After setting the volume of the reaction system, the absorbance is measured in the wavelength range of 300-450 nm.

[0013] When the absorbance ratio is used for measurement, the wavelength pairs of 401 nm / 412 nm (pH 1.89-4.05) and 393 nm / 414 nm (pH 4.99-7.09) are selected respectively.

[0014] The molecular formula of the blood porphyrin monomethyl ether is: 35 40 N4O6

[0015] The structural formula of the blood porphyrin monomethyl ether is:

[0016]

[0017] ​The reaction relates to the reaction in the water phase with different acidity. The absorption spectrum of hematoporphyrin monomethyl ether (HMME) in the aqueous solution has significant correlation with the pH of the medium, and presents the characteristics of "double response", that is, the HMME can respond to the pH in two different intervals, and the response intervals are not overlapped and the response behaviors are different, which is significantly different from the conventional pH optical probe, because the conventional pH probe can only respond to the pH in one interval. The response of the hematoporphyrin monomethyl ether to the pH has the characteristics of dual-function, and the "one needle two uses" can be realized.

[0018] Further, the response of the HMME to the pH has the characteristics of ratio response, and compared with the conventional probe, the ratio type optical pH probe can overcome the influence of many non-pH factors such as the change of the local concentration of the chromophore to cause the error of the measurement, and the more accurate determination of the pH is realized.

[0019] Accordingly, the hematoporphyrin monomethyl ether is used as a new type of pH optical probe, the double-interval response characteristics of the hematoporphyrin monomethyl ether to the pH are utilized, a new method for determining the pH by the optical probe is established, and the method is applied to the determination of the pH of the small volume sample, the viscous sample and the food, the biological sample and the environmental water sample with large pH span, and satisfactory results are obtained, which shows that the method has good practicability.

[0020] Compared with the prior art, the beneficial effects of the technical scheme of the present application are:

[0021] 1) The single porphyrin optical probe is used to realize the determination of the pH in two different intervals, which is significantly different from the traditional optical pH probe, has not been reported before, and is first proposed in the present application.

[0022] 2) The sensitivity is high. The probe has obvious response even if the final concentration is as low as 1.0×10 -6 mol / L.

[0023] 3) The feasibility of the HMME probe in the determination of the pH of the special sample such as the small volume sample and the high viscosity sample is proved by the actual application, which shows that the present application can be applied not only to the determination of the pH of the conventional large volume and low viscosity sample, but also to the determination of the sample which is difficult to be determined by the traditional pH determination method, and has outstanding advantages.

[0024] 4) The operation is simple. BRIEF DESCRIPTION OF DRAWINGS

[0025] Figure 1 The figure is the UV-Vis absorption spectrum of hematoporphyrin monomethyl ether in the pH 1.84-4.15 interval.

[0026] Figure 2 The figure is the UV-Vis absorption spectrum of hematoporphyrin monomethyl ether in the pH 4.99-7.4 interval.

[0027] Figure 3 Logarithmic linear response relationship of hematinic methyl ether absorbance and absorbance ratio to pH in acidic region.

[0028] Figure 4 Logarithmic linear response relationship of hematinic methyl ether absorbance and absorbance ratio to pH in weak acidic-weak alkaline region. DETAILED DESCRIPTION

[0029] In order to make the technical problems, technical solutions and beneficial effects of the present application more clear, explicit and understandable, the present application is further described in detail below in combination with the drawings and examples.

[0030] Determination of constant liquid cell (4.0 mL liquid cell): 90 μL of 1.0 x 10 -4 mol / L hematinic methyl ether aqueous solution was added into a 5.0 mL centrifuge tube, and then buffer solution with different pH was added to make the final volume of the solution 3.0 mL and the final concentration of the probe 3.0 x 10 -6 mol / L. 3.0 mL buffer solution with the same pH was added into another 5.0 mL centrifuge tube as a reference solution for determination of constant liquid cell. The UV-Vis absorption spectrum (300-450 nm) of hematinic methyl ether in different pH medium was scanned.

[0031] Determination of micro liquid cell (400 μL liquid cell): 30 μL of 1.0 x 10 -4 mol / L hematinic methyl ether aqueous solution was added into a 1.0 mL centrifuge tube, and then buffer solution with different pH was added to make the final volume of the solution 1.0 mL and the final concentration of the probe 3.0 x 10 -6 mol / L. 1.0 mL buffer solution with the same pH was added into another 1.0 mL centrifuge tube as a reference solution for determination of micro liquid cell. The UV-Vis absorption spectrum (300-450 nm) of hematinic methyl ether in different pH medium was scanned.

[0032] The present application is described in detail below in combination with the drawings and tables.

[0033] 1) UV-Vis absorption spectrum of hematinic methyl ether in the pH 1.84-4.15 region

[0034] Figure 1 UV-Vis absorption spectrum of hematinic methyl ether in the pH 1.84-4.15 region.

[0035] The maximum absorption wavelength of HMME is at 401 nm in the pH 1.84-4.15 interval, and the peak height gradually decreases with the increase of pH, and two isosbestic points appear at 370 nm and 412 nm. The data on the peak of each curve is the pH value. The concentration of the used hematoporphyrin monomethyl ether is 3.0 x 10 -6 mol / L.

[0036] 2) UV-Vis absorption spectrum of hematoporphyrin monomethyl ether in the pH 4.99-7.4 interval

[0037] Figure 2 The UV-Vis absorption spectrum of hematoporphyrin monomethyl ether in the pH 4.99-7.4 interval.

[0038] In the pH 4.99-7.4 interval, the maximum absorption wavelength of HMME is blue-shifted to 393 nm, and the peak gradually increases with the increase of pH, and two isosbestic points appear at 358 nm and 414 nm. The data on the peak of each curve is the pH value. The concentration of the used hematoporphyrin monomethyl ether is 3.0 x 10 -6 mol / L.

[0039] 3) Logarithmic linear response relationship of absorbance and absorbance ratio of hematoporphyrin monomethyl ether in the acidic interval to pH

[0040] The absorbance of hematoporphyrin monomethyl ether at the absorption peak (401 nm) in the strong acidic interval (pH 1.89-4.05) decreases with the increase of pH, and the negative logarithm of absorbance (-lgA) is positively correlated with pH ( Figure 3 -A). Further, the ratio of absorbance at the absorption peak to that at the isosbestic point (412 nm) is calculated, and the logarithm of the ratio [i.e. lg(A 401 / A 412 )] is calculated, and it is found that the ratio is negatively correlated with pH ( Figure 3 -B), indicating that hematoporphyrin monomethyl ether can be used as both an absorption intensity probe and an absorption ratio probe. The above investigation is repeated in various acidic buffer systems (potassium chloride-hydrochloric acid, potassium hydrogen phthalate-hydrochloric acid, disodium hydrogen phosphate-citric acid, and phosphate universal buffer), and it is found that the linear response relationship of hematoporphyrin monomethyl ether to pH is consistent.

[0041] 4) Logarithmic linear response relationship of absorbance and absorbance ratio of hematoporphyrin monomethyl ether in the weak acidic-weak basic interval to pH

[0042] In the weak acidic-weak basic range (pH 4.99-7.09), the absorption peak of hematoporphyrin monomethyl ether is blue-shifted to 393 nm, and an isosbestic point appears at 414 nm. The absorbance at the absorption peak increases with the increase of pH. The logarithm of the absorbance at 393 nm and the absorbance ratio of 393 nm to 414 nm are processed, and the similar phenomenon is found, i.e. -lgA 393 and lg(A 393 / A 414 ) are linearly related to pH (A Figure 4 -A, B). The linear relationship of the response behavior is just the opposite to that in the acidic range. The above investigation is repeated with various acidic buffer systems (sodium bicarbonate-citric acid, Britton-Robinson and phosphate universal buffer), and the linear relationship is consistent.

[0043] 5) Related parameters of the spectrophotometric response of hematoporphyrin monomethyl ether to pH in a microcell

[0044] The absorption spectrum of hematoporphyrin monomethyl ether in phosphate universal buffer solution is scanned with a 400 μL microcell, and the spectral behavior is not different from that obtained with a constant quartz cell. In the acidic range (pH 1.99-4.04), the negative logarithm of the absorbance at the absorption peak (-lgA 401 ) is positively linearly related to pH, and the logarithm of the absorbance ratio of the absorption peak to the isosbestic point [lg(A 401 / A 412 )] is negatively linearly related to pH. In the weak acidic-weak basic range (pH 5.02-7.00), the negative logarithm of the absorbance at the absorption peak (-lgA 393 ) is negatively linearly related to pH, and the logarithm of the absorbance ratio of the absorption peak to the isosbestic point [lg(A 393 / A 414 )] is positively linearly related to pH, which is not different from the result obtained with a constant cell, indicating that hematoporphyrin monomethyl ether can be used for the determination of a micro-volume sample. The specific results are shown in Table 1.

[0045] Table 1

[0046]

[0047] 6) Logarithmic linear characteristic parameters of the absorbance and absorbance ratio of hematoporphyrin monomethyl ether to pH in a high-viscosity medium

[0048] The pH of the system was determined by adding 50% glycerol to the buffer solution at different pH to form a high viscosity buffer system, using HMME as a probe. The absorption spectrum of HMME in the microcuvette was scanned, and the obtained spectrum was compared with that without glycerol. It was found that the positions of the absorption peak and the isosbestic point only slightly shifted. In the acidic range of pH (2.33-3.77), the negative logarithm of absorbance (-lgA 403 ) at the absorption peak (403 nm) was positively linearly related to pH, and the logarithmic value of the ratio of absorbance at the absorption peak to that at the isosbestic point [lg(A 403 / A 414 )] was negatively linearly related to pH. In the weakly acidic to weakly alkaline range of pH (4.32-6.14), the maximum absorption wavelength of the spectrum was 396 nm, and the isosbestic point was 420 nm. The negative logarithm of absorbance (-lgA 396 ) at the absorption peak (396 nm) was negatively linearly related to pH, and the logarithmic value of the ratio of absorbance at the absorption peak to that at the isosbestic point [lg(A 396 / A 420 )] was positively linearly related to pH. The results were consistent with those in the buffer system without glycerol, indicating that HMME can be used for the determination of the pH of a high viscosity sample. The specific results are shown in Table 2.

[0049] Table 2

[0050]

[0051] 7) Determination results of actual samples

[0052] Using hematoporphytin monomethyl ether as a probe, it was applied to the determination of the pH of food (white vinegar), environment (rainwater) and medical (urine) samples.

[0053] I. Determination of the pH of white vinegar

[0054] The pH of the purchased white vinegar sample (without any treatment) was determined by a pH meter as a reference value. Hematoporphytin monomethyl ether (3.0×10 -6 mol / L) was added to the sample to determine the absorbance, and then the pH value of the white vinegar was calculated according to the positive linear relationship between the negative logarithm of absorbance (-lgA) at the absorption peak (401 nm) of hematoporphytin monomethyl ether in the acidic range (pH 1.89-4.05) and pH.

[0055] II. Determination of the pH of rainwater

[0056] The pH value of the environmental rainwater sample collected on a rainy day was determined by a pH meter as a reference value. Hematoporphytin monomethyl ether (3.0×10 -6The pH of the rainwater is calculated according to the pH response relationship of the hemin monomethyl ether to pH in the weak acid-weak base interval (pH 4.99-7.09) based on the absorbance measured at 405 nm.

[0057] III. Determination of pH of urine of normal person

[0058] The morning urine of a normal person is taken, and the pH value is directly measured by a pH meter as a reference value without any treatment. The sample is added with hemin monomethyl ether (3.0×10 -6 The pH of the morning urine is calculated according to the pH response relationship of the hemin monomethyl ether to pH in the weak acid-weak base interval (pH 4.99-7.09) based on the absorbance measured at 405 nm.

[0059] As shown in Table 3, the results of determination of pH of three actual samples by the method of the present application are consistent with the results of determination by a pH meter, which indicates the practicability of the method.

[0060] Table 3

[0061]

[0062] In the present application, the absorption spectrum of the hemin monomethyl ether shows a "double response" characteristic to the pH of a medium, i.e., the hemin monomethyl ether can respond to the pH in two different intervals, so that a single probe can be used to determine the pH in different intervals, i.e., "one needle for two uses". The hemin monomethyl ether also has a ratio response characteristic to pH. The ratio-type pH optical probe can overcome the influence of many non-pH factors. The present application utilizes the double-interval response characteristic of the hemin monomethyl ether to pH in luminosity, and establishes a new method for determining pH by using the hemin monomethyl ether as an optical probe. The present application is used for determination of pH of actual samples such as small-volume samples, viscous samples, food samples, biological samples and environmental water samples with a large span of acidity, and satisfactory results are obtained, which shows the practical application advantages of the pH optical probe and opens up the application of the light-absorption type porphyrin compound in analytical science.

Claims

1. A dual-zone responsive pH optical probe, characterized in that: The dual-range response pH optical probe adopts hematoporphyrin monomethyl ether, which has two response ranges of pH 1.89-4.05 and pH 4.99-7.09 to pH; The application of the dual-range response pH optical probe comprises the following steps: 1) adding hematoporphyrin monomethyl ether aqueous solution with a fixed concentration and volume into a series of containers; 2) adding buffer solutions with different pH values in sequence into the above series of containers, and adding water to constant volume; 3) scanning the absorption spectrum of the pH solution system, and measuring the absorbance at the absorption peak or a pair of wavelengths; When a working curve is made or a sample is determined, the absorbance or absorbance ratio is logarithmized, and then the pH is plotted.

2. A dual-zone responsive pH optical probe as claimed in claim 1, wherein: The containers are at least 3.

3. A dual-zone responsive pH optical probe as claimed in claim 1, wherein: The pH values of the buffer solutions fall within the corresponding working curve linear range, which is determined by the concentration of the hematoporphyrin monomethyl ether aqueous solution used and the linear working curve range for determining the pH.

4. A dual-zone responsive pH optical probe as defined in claim 1, wherein: The absorbance is determined in the wavelength range of 300-450 nm.

5. A dual-zone responsive pH optical probe as defined in claim 3, wherein: The added hematoxylin monomethyl ether aqueous solution is the corresponding amount of porphyrin solution with a final concentration of 1.0 x 10 -6 ~1.2 x 10 -5 mol / L.

6. A dual-zone responsive pH optical probe as defined in claim 1, wherein: When the absorbance ratio is used for determination, the wavelength pairs 401 nm / 412 nm and 393 nm / 414 nm are selected, respectively.

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