Study On Molecular Composition And Structure Of Fulvic Acid And Humic Acid By High Resolution Mass Spectrometry

Humic substances is a complex macromolecule organic matter and an important component in natural organic matter.It plays a significant role in the environmental behavior,bioavailability and toxicity of various pollutants in environmental media,and has great application prospects in agriculture,medicine and medicine field.However,the molecular composition and chemical structure of humic substances is not very clear,and its composition and structure are easily affected by the origin,age,climate and biological conditions.In this paper,the composition and structure of fulvic acid and humic acid from different sources were characterized in detail on molecular level by the electrospray ionization combined with Fourier cyclotron resonance mass spectrometer?ESI FT-ICR MS?.In addition,forest soil humic acids were fractionated to six sub-fractions by using different concentrations of sodium pyrophosphate solution and sodium hydroxide solution according to the solubility of humic acids at different pH values,and their similarities and differences in composition and structure were compared by ESI FT-ICR MS.The main research conclusions are as follows:1.The molecular compositions of four representative samples of fulvic acid?river water,lake water,prairie soil and forest soil?were comprehensively analyzed,and the molecular characteristics and their differences of different fulvic acid were revealed.As a result,2708-4438 molecular formulas were assigned in the four fulvic acid samples,and CHO,CHOS₁,CHON₁,CHON₁S₁,CHON₂ and CHON₃compounds were identified in fulvic acid samples.CHO was the most abundant compound,and CHON₁S₁ and CHON₃ compounds were existed as unique to soil FA.The ratio of lignin components in soil fulvic acid was lower than that in aquatic fulvic acid,while the ratio of condensed aromatic structure components was greater than that in aquatic fulvic acid.The molecular aromaticity of the samples was as follows:river water<lake water<prairie soil<forest soil;and the aromaticity of various compounds was as follows:CHOS₁<CHO<CHON₁<CHON₁S₁<CHON₂<CHON₃,which shows that the aromatic character increases with increasing N atom number.The distribution of average DBE and average carbon number of CHO compounds decrease first and then increase as the oxygen atoms increased.Some of the molecules of O₄,O₆ and O₁₀ species compounds are probably cyclic compounds containing more carboxyl groups and hydroxyl groups.In addition,we also analyzed the mechanism of ammonia as an ionizing agent in high resolution mass spectrometry.Under the condition of ammonia,the proportion of CHO compound of four fulvic acid increased,CHOS₁,CHON₂ and CHON₃ compounds decreased,and the proportion of condensed aromatic structure components decreased,but the proportion of lipids,proteins/peptides and lignin components increased.2.Systematic comparison on molecular composition of four humic acid samples?river water,forest soil,peatland and leonardite?with different humification degree were performed.In total,767-2775 molecular formulas were matched in four humic acid samples,CHO,CHOS₁,CHON₁ and CHON₂ compound were identified in humic acid samples.CHO compound was the most abundant compound in each sample,and compared to the humic acid with low humification degree,humic acid with high humification degree contains more CHON and less CHO compound,and more condensed aromatic structure components.The aromaticity of various compounds is CHO<CHOS₁<CHON₁<CHON₂,and the aromaticity of CHON compounds is the strongest.The distribution of the average DBE and the average carbon number of CHO compound have a strong linear relationship with the number of oxygen atoms in the range of O₇-O₁₆.Some compounds of O₇ are probably aromatic compounds containing carboxyl and hydroxyl groups.3.The molecular composition characteristics and differences of soil humic acid sub-fractions extracted by Na₄P₂O₇ solution and NaOH solution were analyzed.The number of matched molecular formulas of six humic acid sub-fractions was from1293-2103.With the increase of elution sequence,the proportion of CHO and CHON₁compounds are increased and decreased respectively.The ratio of lipids and condensed aromatic structure components are ascent and descent respectively,and lignin compounds increased with the elution sequence under the same extraction agent,indicating the different concentrations of Na₄P₂O₇ solution and NaOH solution could successfully separate the sub-fractions with different composition and structure from the soil humic acid.Except for the last sub-fraction,the aromaticity of all compounds obeys the rule of CHO<CHOS₁<CHON₁<CHON₂<CHON₃,and the aromaticity of CHON compounds is the highest.The proportion of CHO compounds in Na₄P₂O₇ solution extraction is less than that in NaOH solution extraction and the content of CHON compounds in Na₄P₂O₇ solution extraction is higher than that in NaOH solution extraction.The average DBE of CHO compounds in Na₄P₂O₇ solution extraction is proportional to the number of oxygen atoms.4.The differences of molecular composition between fulvic acid and humic acid from the same source were compared.For the same origin of fulvic acid and humic acid,fulvic acid contains more species composition than humic acid.The proportion of CHO compounds in fulvic acid is lower than that of humic acid,while the proportion of CHOS₁ and CHON compounds is significantly higher than that of humic acid.In addition,the ratio od lignin component in fulvic acid is higher than that of humic acid,while the ratio of lipids component is lower than that of humic acid.