| 其他摘要 | Biogenic organic acids such as formic and acetic acids are universal occurrences in atmospheric environment, and each of them in the troposphere is an important trace volatile organic carbon component which has great impacts on climate, environment, ecosystems and human health. In order to study the biogeochemical cycles of the organic acids, a more direct method is needed for discerning the specific sources; carbon isotopic analyses of the organic acids was investigated as such a method. Therefore, to explore stable carbon isotope of organic acids (δ13C value) analytical method can move forward the studies of organic acids on biogeochemical cycles. This preliminary study presents the stable carbon isotope compounds of formic and acetic acids analysis method use solid phase needle trap micro extraction combined with gas chromatography/combustor/isotope ratio mass chromatography (GC/C/IRMS). The thesis has laid a foundation on the methodology for an in-depth research to explore "the carbon isotopic composition of different organic acids’ sources", "the existence of unknown sources of organic acids "and" the contribution of different sources in atmospheric environment".
Research focusing on the following five aspects, first of all, the thesis has reviewed and summed up the study evolution of low molecular weight organic acids on the world, in this part, organic acid concentration levels in various sample forms and distribution in different sites were abstracted in to tables. Secondly, in order to obtain an appropriate enrichment and separation method which adapt to GC/C/ IRMS interface, the thesis has reviewed the low molecular weight organic acids concentration analysis method. At the third, research has demonstrated that low molecular weight organic acids in aqueous solution can be extracted by NeedlEx, many conditions such as pH, purge gas volume were optimized to improve the extraction efficiency. Fourthly, trace formic and acetic acids in atmospheric samples such as in rainwater or gas aerosol were extracted by NeedlEx and δ13C values of them were analyzed by GC/C/ IRMS. Fifthly, δ13C values of formic and acetic acids emission from natural products or automobile exhaust were analyzed as potential sources for isotopic analysis. The most important content is: to application the most appropriate, current method (In terms of needle trap solid phase micro extraction, NeedlEx) for extraction trace organic acids from aqueous solution, and then analysis their δ13C values by GC/C/ IRMS. During the course of the method established, main results were obtained as follows:
1. To establish a stable carbon isotopic composition analysis method for volatile organic compound standard samples such as formic and acetic acids
Quartz tube connect with static sealing flange technology can obtain a high vacuum effectively, and it avoid possiblely CO2 pollution and sample volatile leakage apart from atmosphere when inject samples. After load acid in the tube individually, then closed it with gas torch that burns ethyne and combust it into CO2 at 850℃, after refined again, the CO2 can be analysis by gas isotope mass spectrometry (Finnigan MAT-252). These prepared courses can be effectively avoided isotope fractionation cause by evaporation and environmental CO2 impact on the determination values. Take the market available high purity formic and acetic acids as the two standard samples, preparation of their isotopic analysis for 5 repeats, then use the Finnigan MAT-252 to analysis the value of their δ13C value. The results showed that it is highly reproducible for using this method, the measured value of relative standard deviation of δ13C for each acid is 0.07% (formic acid, n = 5) and 0.04% (acetic acid, n = 5) respectly. On the other hand, other data sets of the same standards were measured by GC/C/IRMS directly, but there were no significant difference between the results of the two methods statistically, except for the accuracy are obviously less than the former. High precision and low cost shoud makes the method can be applied to other volatile organic compounds’ δ13C value calibration. Therefore, it is significative for accurate determination of the corresponding δ13C values of the volatile organic compound in actual sample, and provides a basis conditions for isotopic analysis.
2. Conditions were optimized for gas chmatography and isotope ratio mass spectrometry for online separation and isotopeic measurement for formic and acetic acids.
In order to separation formic and acetic acids form NeedlEx without any derivative step, Stabilwax-DA capillary column (30m×0.25mm I.D., 0.25m D.F.; Restek, Bellefonte, PA, USA) which was specially developed for analysising the volatile free acids was selected. Helium and nitrogen were supplied by passing through the molecular sieve and used as carrier and makeup gas, respectively. GC spareation was programmed as follows: initial injection port temperature set at 200℃, splitless injection; up a two-stage procedure, the main gradient conditions were as follows: initial oven temperature at 65℃, and hold it for 1min; temperature gradient 1: ramp to 80℃ at a rate of 2℃/min; temperature gradient 2: ramp to 170℃ at a rate of 15℃/min and hold for 2mins. Under these conditions, there is a gap of 0.79 minutes between acetic and formic acids’ retention time, and each standard deviation of retention time for formic and acetic are less than 0.05min, it could be considered that two of the acids was separated completely, and the conditions is suitable for determination of carbon isotopes in the next step. Formic and acetic acids separated from Gas chromatography were send into an interface maintained at 350℃, and then followed by a combuster at 850℃, all of the acids were converted into CO2 , after eliminate H2O by Nafion membrane, the isotopic signatures of the CO2 were obtained by calibrating against a referenced CO2 standard which was relatived to Vienna Pee Dee Belemnite.
3. Conditions for solid phase micro extraction of formic and acetic acids form aqueous phase were optimized, and an extraction device was designed to be used for the experiment.
A needle extraction device (NeedlEx) coupled with a gas aspirating pump was intended to perform sampling and preconcentration of target compounds from aqueous sample before gas chromatographic analysis, therefore, the NeedlEx for atty acids can be used to extract organic acids in aqueous solution. In order to improve the efficiency and reduce environmental pollution, the conditions for the solid phase micro extraction technology were optimized, and we have designed a device of dynamics headspace -needle trap for sample preparation, and couple with GC/C/IRMS to determination the formic and acetic acids’ δ13C value in aqueous solution.
The devices’ design was based on the principle of purge and trap, and it’s well known that increase the volume of purge and trap gas would increase the amount of analyte. Our results show that the total amount of organic acid in NeedlEx has been rapid increase with the extraction gas volume reached as 400mL, and then the increase velocity will slow down.In order to extract the most quality of organic acids in aqueous sample and reduce the labor intensity for extraction. 1000 and 2000mL extraction gas has been attempted; statistics ofδ13C value for each acid show that there was no significant difference between the two volumes, so we adopt 1000mL extraction gas for our experiment for the workload balance.
It’s also well known that organic acid anions are hydrophilic, and these forms for acids are too difficult to volatile into the headspace, only the states of free organic acids volatile into the headspace could be extracted by hydrophobic sorbent of NeedlEx partly. In order to forward direction of the dissociation reaction to the favorable undissociated both compounds, we can see almost all of acids that occur in the solution are in the neutral form when the pH is less than 2. many of the different conditions of acidification effects on the extraction were assessed, but we generally used 500μL 4mol / L of phosphoric acid as acidification condition in experiment, results show that acidification could improve the the amount of analyte, take 10μg/mL formic and acetic acid standard solution for example, increase rate reach up to 30.12% and 14.46% for formic and acetic acid respectively. However, there was not any significant different on δ13C value between acidification and non acidification in generally, but because of the precision of δ13C test value for each acids has a positive correlation with the quality of analyte. Therefore, precision of δ13C value after acidification is superior than without acidification. For example, standard deviation of δ13C value for 10μg/mL standard solution after acidification was equal to 1.22‰, but the non acidification one was up to2.93‰.
It is report that extraction temperature is one of the important parameters for the evaluation of extraction efficiency in SPME and other extraction techniques.But Increase of temperature accelerates the entrance of liquid into the pores of sorbent, and The liquid–liquid intermolecular attraction, which is responsible for surface tension, leads to capillary condensation in pores.If the capillary forces are stronger than adsorption, as is in the case of higher temperature, the needle performance decreases. Based on the evaluation, ambient temperature (25℃) was chosen as the optimization extraction temperature in the following experiments. Additionally, samples were stirred on a magnetic mixer at 2000 round/min m to establish equilibrium between the headspace and the aqueous.
Using the optimized conditions described above, the validation procedures of the NeedlEx method were performed for the quantitative and isotope analysis of the target compounds in standard sample. The samples for the calibration study were prepared by spiking pure water with analytes over concentration ranges of 10~300μg/mL. The results showed that concentration of not less than 10μg/mL can be determined the δ13C value otherwise the peak area will be too small to calculate the δ13C value, but Linearity which was evaluated in terms of the correlation coefficients of the regression equations of the calibration curves was very well. The last concentration for which the δ13C value was within this iterative interval and for which the standard deviation was less than 1‰ (triplicate measurements) should be 50μg/mL for acetic acid and less than 100μg/mL for formic acid 4. Formic and acetic acids in the rainwater samples must be preconcentration by solid phase extraction (SPE) or else they couldn’t be analysed by the above method.
It is well known that concentration of formic and acetic acids in rainwater are againsted a few to several dozens of μmolL-1, it is means that NeedLEx can not be directly applied in extraction of the acids in rainwater for isotopic analysis. So that an anion-exchange cartridge of SPE (LC-SAX; specifications: 500mg/3mL; exchange capacity: 0.2meq/g) was used to preconcentration the aicds in 1L aqueous standard sanples. Calibration study were prepared by spiking pure water with analytes over concentration ranges of 50~2000μg/L too. Compare the average of δ13C values for each acid of all most concentration with theoretic calues, we found that there is no significance difference between two of them for fomic and acetic acid, probabilities of most δ13C values for each concentration were greater than the significant level of 0.05(except 200μg /L for acetic acid), it prove that there was no significant difference between the determination and the theoretical value.
However, In addition to organic acids, there were other anionics in precipitation; the components complex system will seriously reduce the anion-exchange capacity of organic acids in cartridge. Accordingly, seven low molecular weight carboxylic anions and inorganic anions were determined by ion chromatography(IC), and corresponding exchange capacity of several cartridges were selected to implement the extraction action, and the trapping with NeedlEx and isotopic analysis.
5. A preliminary analysis of the δ13C values for formic and acetic acids in the rainwater or gas phase was studied.
Guiyang, a representational city of southwest China as there’s high concentration of organic acid in the atmosphere, and it’s favorable for using the method to study the δ13C values of organic acids in this region. In late August 2008 to mid-October 2008, six precipitations were collected in the city, concentration of formic and acetic acids were range form 5.75 to 22.43 μmol/L and 5.43 ~ 13.09μmol/L determined by IC. And Correspondingδ13C values were range form -29.08 ‰ to -25.72 ‰ for formic acid and from -30.40 ‰ to-26.23 ‰ for the acetic acid. The measuredδ13C values points that both of the acids towards terrestrial sources for the six precipitations.
Combine with the concentration analysis, the determination of formic/acetic ratio-Henry's law coefficient tell us that most of fomic and acetic acids were source from photochemical oxidation of unsaturated organic compounds in atmosphere. Futher more, δ13C values of the acids tell us that the unsaturated organic compounds are major source from vegetations, human activities and biomass burning.
Previous studies pointed out that the high degree of correlation between formic and acetic acids was result as the same source of them. Similarly, there is a significant linear correlation (R2=0.87) of the two targets in the six precipitations of Guiyang. Assuredly, δ13C value with similar changes in scope -25.72 ‰ to -30.40 ‰ can be sure that there is a homology sources model of the formic and acetic acids in the rainwater samples.
The devices of NeedlExes are also suitable for extracting the free organic acids of gas phase in atmosphere. So thatδ13C values of fomic and acetic acids in atmosphere for three sites were been analyzed. The results showed that δ13C values of acetic acid in atmosphere were in the scope of -31.03‰ to -26.37‰, and mean equal to -28.74‰ from urban to suburbs area. On the other handδ13C values of formic acid in atmosphere are form -29.42‰ to -22.97‰, mean equal to -27.12‰ at the same sites. All theδ13C values of the acids in atmospheric gas phase pointed towards that both of the two acids had the same terrestrial sources.
We also benefit as well as the method of δ13C value measurement from formic and acetic acids in automobile exhaust and formic acid emission from formicine ants. It is show that δ13C values of -23.25 ± 1.25 ‰ for formic acid and -22.43 ± 0.43 ‰ for acetic acids as the results of automobile exhaust. Formicine ants, on the other hand, ranged of -22.43±0.43‰. As shown earlier, measured atmospheric formic and acetic acid in Guiyang had a composition less depleted in 13C, therefore, formic and acetic acids emission from automobile exhaust or formicine ants would not be the major source of atmospheric organic acids.
6. Expansion of applying research of the isotopic analysis method on fraudulent food labeled with 13C in acetic acid.
Different photosynthetic routes such as the Calvin cycle (C3), Hatch-Slack cycle (C4) will lead to the products of them have different δ13C value, so that the stable carbon isotope ratio analysis play a special role in determining the authenticity of foodstuffs. In this study, δ13C values of acetic acid in vinegar were found to be closely related with the raw materials, take wheat bran, rice vinegar for the raw materials of vinegar for example, the δ13C values of them are around -27 ‰. But 13C of the acetic acids fermented with sorghum, barley and soybean are more abundant, which δ13C values are around -19 ‰. Other production of the rice vinegar has a composition less depleted in 13C, and δ13C value of acetic acid is around -29 ‰. Significant different δ13C values of acetic acid in various vinegars made that the method of isotopic analysis provides a good solution for quality inspection. |
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