Chemical Degradation of Humic Substances for Structural

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Downloaded by FORDHAM UNIV on January 2, 2013 | http://pubs.acs.org Publication Date: December 15, 1988 | doi: 10.1021/ba-1988-0219.ch001

Chemical Degradation of Humic Substances for Structural Characterization L. B. Sonnenberg, J. D. Johnson, and R. F. Christman Department of Environmental Sciences and Engineering, School of Public Health, University of North Carolina-Chapel Hill, Chapel Hill, NC 27514

Chemical degradations of humic substances are reviewed, with emphasis on the identity of the products and how these compounds relate to the overall structure of the humic macromolecule. The methods described include (1) reduction using metals, hydrogen, and metal hydrides; (2) oxidation with copper oxide, permanganate, and chlorine; and (3) alkaline hydrolysis. Experimental results are reported for the reductive and solvolytic degradation of isolated aquatic humic acid using metallic sodium dissolved in liquid ammonia. A number of highly oxygenated compounds, many that commonly result from oxidative procedures, are identified in the reduction mixture. The presence of these structures in the original humic macromolecule is suggested.

DEGRADATIVE TECHNIQUES CAN BE EFFECTIVE TOOLS for structural elu-

cidation of complex natural polymers (I). Prediction of the types of bonds that are unstable to a particular method of degradation, coupled with identification and quantification of products, can indicate what substructures are included in the macromolecule and how they are linked together. Ideally, these techniques will produce untransformed subunits in a mechanistically predictable manner. Degradative methods have long been used to examine the subunits that exist in humic substances and their mode of attachment to the parent molecule. Aquatic humic material has been degraded hydrolytically and oxidatively by a variety of methods. However, because of the uncertainty about

0065-2393/89/0219-0003$06.25/0 © 1989 American Chemical Society

In Aquatic Humic Substances; Suffet, I., et al.; Advances in Chemistry; American Chemical Society: Washington, DC, 1988.


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AQUATIC HUMIC SUBSTANCES

the reactions that are taking place in these complex reaction mixtures, com pounds identified may not represent structures in the parent molecule. The low weight yields generally obtained in oxidative and hydrolytic work (1-25%) also make extrapolation to the maeromolecular structure difficult. As yet no single analytical method, degradative or instrumental, can provide data for unequivocal characterization of humic structure. Therefore, the use of several techniques, chemical and instrumental, and the comparison and corroboration of data obtained from each of these methods is a sound ap proach to the complex issue of humic composition. Although seldom applied to aquatic humic material, reductive degra dation is a potentially important element in a multifaceted approach to its characterization. Reduction of humic substances can be viewed as a com plement to oxidation. The two processes are similar, in that structural fea tures affecting their outcome are likely to be similar. Because the fundamental mechanisms of the initial step of cleavage are different, how ever, the sites of fission and secondary reactions are not likely to be the same. In this chapter, some important results of oxidative and hydrolytic deg radation of aquatic humic material are reviewed. Because reductive degra dation has not been commonly applied to aquatic humic substances, a review of methods and results for the reduction of nonaquatic humic materials and humiclike compounds is given. The application of a new reductive degra dative method to aquatic humic acid is reported.

Oxidation and Hydrolysis of Aquatic Humic Materials Some degradative procedures that have produced measurable quantities of subunits from aquatic humic and fulvic acid include alkaline hydrolysis (2) and oxidation using copper oxide (3-6), potassium permanganate (2, 7), and chlorine (8-10). Classes of compounds that represent the majority of the compounds produced by each method are given in Table I. Liao (2, J I) exposed isolated humic and fulvic acid from two aquatic sources (Black Lake, North Carolina, and Lake Drummond, Virginia) to 0.5 Ν N a O H for 1.5 h to get under 2% by weight of acidic ether-extractable products. Although fission of certain activated ethers (as in lignin) and of esters might be expected, ester cleavage was postulated (2,12). The alcohols expected from the cleavage of both of these bond types were not identified, a result suggesting that the alcoholic fragments may be on a very large molecule. Distributions of hydrolysis and permanganate oxidation products were similar, but permanganate gave 20-25% weight yields of acidic ether-ex tractable products. A n exception to the similarity in product identities was the formation of phenylglyoxylic acids in the permanganate procedure (2). Analysis of permanganate degradation products indicated the presence of


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SONNENBERG ET AL.

Degradation for Structural Characterization

Table I. Major Classes of Common Degradation Products from Aquatic Humic Material KMn0 Product Classes NaOH CuO Aliphatic monoacids + + diacids ++ + ++ triacids + + Carboxylic benzenes ++ + ++ phenols-anisoles ++ ++ ++ furans + + Phenolic-anisolic + Phenylglyoxylic acids + Aromatic aldehydes ++ ketones ++


4

Cl

2

++ ++ + +

NOTE: References from which this table is compiled are given in the text. "Single pluses indicate only that the class of products was found in the reaction products. 'Two pluses indicate that the class of compounds was present as major reaction products.

aromatic compounds with three to six substituents (alkyl, ketone, carboxylic acid, or hydroxyl), short-chain aliphatics, polycyclic ring structures, and carbohydrates. These substructures were thought to be joined to the humic macromolecule through carbon-carbon bonds (2), because permanganate cleaves double bonds and alkyl side chains of aromatic compounds. Reuter et al. (7) obtained about 12% yields of similar products for the permanganate treatment of nonmethylated humic material. Premethylated humic substances gave a higher yield with a different product distribution. Reuter et al. concluded that the Satilla River humus they degraded was more aliphatic in nature than previously believed. They based their conclu sion on the prevalence of oxalic acid when premethylated humus was de graded in methylene chloride containing crown ether (for permanganate dissolution). The oxalic acid identified in the reaction mixture was also judged to indicate a high degree of unsaturation and oxygenation of the aliphatic portion of the macromolecule. Phenolic acids were determined to be the dominant form of aromatic compounds on the basis of the same experiments. The production of most compounds (including furanmonocarboxylic acid) was explained in terms ofhumic structure. However, furandicarboxylic acid, which was also found by Liao et al. (II), was considered an artifact of the oxidative treatment of premethylated humic material because its distribution varied with treatment conditions. Although treatment of humic substances with copper oxide is a milder degradative procedure than permanganate oxidation, it has produced a num ber of identifiable compounds. Christman and Ghassemi (3) provided con clusive evidence for the presence of aromatic structures in aquatic humic material by using copper oxide degradation. Phenolic products, both ligninand non-lignin-derived, were considered structural nuclei. In addition, alkyl


5

6


and ether linkages were postulated on the basis of these oxidative data. Ertel and Hedges (4) obtained 0.8-5.2% of the original starting carbon in aquatic humic material in the form of lignin-derived phenols. The suite of phenols that were released from humic matter, including those with carboxylic, alkyl, and aldehydic functional groups, were related to the biogeochemical nature of the humic material investigated (4, 13). Norwood (5, 6), who degraded aquatic humic material with copper oxide to carry out structural studies, found both aliphatic (5) and aromatic (5, 6) material. Although the presence of phenolic structures is confirmed through copper oxide degradation results and solid-state C N M R spectra, their relative contribution to the overall structure of fulvic acid is small for the aquatic humic material examined, as shown by solid-state C N M R spectra (5, 6). In fact, C N M R data reported by Norwood et al. (6) suggested that aliphatic carbon is 51% of the total humic carbon; only 21% of the total carbon is aromatic. When alkaline copper oxide was applied to soil humic acid and the results were compared to the products of alkaline hydrolysis, it was found that the weight yields were similar, but the oxidation products were smaller and more easily identified (14). Degradation by copper oxide appears to be the only oxidation method reviewed to give noncarboxylic phenols. Although it is likely that these compounds exist as such in the parent molecule, decarboxylation of o-phenolic acids can occur with copper oxide treatment (3, 15). Chlorination of aquatic material has been conducted to release humic subunits from the parent molecule, as well as to investigate its role in by product formation during disinfection of drinking water and wastewater. Chlorination studies at high p H (8, 16) (using aquatic humic acid) and at neutral p H (10) (using aquatic fulvic acid) under high chlorine-to-carbon molar ratios gave an array of chlorinated and nonchlorinated products dis tributed as indicated in Table I. The weight yield of products at high chlorineto-carbon ratios is intermediate to those of base hydrolysis and permanganate oxidation (22). Structural inferences made from chlorination experiments include the following: (1) a highly cross-linked structure is consistent with the formation of aromatic compounds that are highly substituted with carboxyl groups from oxidative carbon-carbon cleavage (16); (2) unsaturated short-chain acids are derived from ring cleavage of anisolic and phenolic compounds (16); (3) saturated acids arise from saturated side chains (12); and (4) fused ring systems in the humic macromolecule may produce phenylglyoxylic acids (10). As shown in Table I, a high degree of oxygenation is found in the major products of oxidative and hydrolytic degradation of aquatic humic material. The presence of polycarboxyl functional groups is a striking feature of the aliphatic, benzoic, phenolic, and furan products. It is not surprising that oxidation and hydrolysis primarily produce compounds with high oxidation states. Whether these functional groups originate from the degradative processes or are present in the original molecule is unclear. However, data


1 3

1 3

1 3


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SONNENBERG ET AL.

Degradation for Structural Characterization7

from permanganate and hydrolysis studies have been used to demonstrate method-specific structural relationships of humic material (12). Models of structures that could produce the amounts and types of compounds found for each degradation method are shown in Charts I and II.


M

j

1

R-OX-(CHJ-R'

M=macromolecule

if R'=COOH, X=2 (II)

( ) = relative weight

if R'= C H , X= 14 3

(I)

R* Alk., or Ar.

Chart I. Hypothetical structural relationships in aquatic humic material based on degradation products of alkaline hydrolysis. (Reproduced with permission from ref 12. Copyright 1989 John Wiley à- Sons.)

OH

N=3 Chart II. Hypothetical structural relationships in aquatic humic material based on degradation products of permanganate. (Reproduced with permission from ref. 12. Copyright 1989 John Wiley & Sons.) In Aquatic Humic Substances; Suffet, I., et al.; Advances in Chemistry; American Chemical Society: Washington, DC, 1988.

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Reductive Degradation A class of reactions used in lignin and soil humic studies but seldom applied to aquatic humic work is reductive degradation. Reduction can be achieved by a number of methods, including catalytic hydrogénation and the use of metals, metallic hydrides, and nonmetallic reducing compounds such as those containing phosphorus and sulfur. Several of these procedures have been employed to evaluate the properties ofhumic and humiclike material. Zinc dust distillation of peat humic acid (17, 18) and soil humic acid (19) and zinc dust fusion of the latter humic material (19) yielded polyaromatic hydrocarbons. Interestingly, Chesire and various co-workers (17, 18) saw evidence of a polyaromatic hydrocarbon core in peat humic acid with per manganate oxidation followed by decarboxylation. These same investigators found complex aromatic compounds when peat humic acid was treated with hydrogen iodide and red phosphorus (J 7, 18). Hydrogénation of lignin with copper chromite and Raney nickel catalysts has given evidence of guaiacyl and syringyl nuclei linked by β- and ^-ethers, as well as by propyl groups (20-23). Condensed aromatic ring systems with side chains linked through oxygen bridges were postulated by Kukharenko and Savelev (24, 25) to be constituents of the neutral fraction ofhumic acids hydrogenated with a nickel catalyst. Volatile and nonvolatile carboxylic acids and phenols were other products of the hydrogénation. Felbeck (26) de graded organic matter from a muck soil by using high-pressure hydrogenolysis followed by hydrogénation with Raney nickel. Results suggested that at least one carbon-carbon double bond exists for every four carbon atoms in the nonhydrolyzable fraction. A n n - C ^ or n - C ^ hydrocarbon, detected in one vacuum distillate, may have arisen from chains of unsaturated het erocyclic compounds connected by carbon-carbon bonds (26).

Alkali Metal Reduction of Humic Materials The reductive procedure most used for the degradation of humic material entails the use of sodium amalgam, usually in alkaline solution. Mostly soil humic acid has been degraded, although one application to aquatic organic matter is discussed. The different procedures followed and results obtained are summarized in Table II and Chart III. Many of the sodium amalgam investigations were undertaken to differ entiate sources ofhumic material, both on a geographic basis and with respect to their biogenesis. To these ends, a great deal of emphasis has been placed on the array of phenols produced, as indicated in Chart III. One of the first groups of investigators of sodium amalgam degradation, Burges et al. (27, 28), assigned most of the phenolic products to two groups: those derived from lignin and those derived from either flavonoids or soil microbial proc esses. The differences in thin-layer chromatograms of reaction products were deemed to be of value as a "fingerprint technique" for humic origin



25-50 mg 250 mg 2g 25-40 mg 0.5 g ig

30 20 50 25 50 100 5 5 3 5 3 5 0.15 4.0 0.8 30.0-50.0 3.0 5.0

3 3 4 3-4 1-2 3

4

Time (b) 3

Yields" (% wt) 1 x 10-14 >1 x 30-35 1 x 2-5 4 x 10-15 1 x 10-12 2 x 5-8 3 x 32 1 x 8-20 2 x 8-20 1 x 12-16 (methylated extracts)

fe

"Yields are based on weights of ether-extractable products. The number of sequential degradations required to produce the yields is given. AH, acid hydrolyzed; UH, unhydrolyzed.

UH soil'' AH soil AH soil soil, peat AH soil AH soil

2.1-0.8

39 45 46 33 42 43

3

29

50

0.7-2.0 g

Reference 28

AH soil

Table II. Sodium Amalgam Reaction Conditions and Yields Na--Hg ReductantHumic Acid Amount Strength Suhstrate Amount Source (wt ratio) ω (%) 0.5 g AH soil 50 3 3.0 6



(29)

(39)

(46)

(33,39)

(28,39,42,45,46)

(46)

(28,33,39)

(28,33)

(28,46)

(28,33,45,46)

(33,39,46)

(28,33,39,45,46)

(39)


(28)

(29)

(28,39,46)

1. SONNENBERG ET AL.


so

S

ε "S CO

a. eo


CO

IS s Ο.

•S Si

"S 80% match of low-resolution El spectra with library spectra. ^Identified by low-resolution EI, CI,fragmentationanalysis, and manual match of literature spectra. •"Identified by both methods in footnotes a and h.


Cl

2

1. SONNENBERG ET AL.


21


Conclusions Oxidative, hydrolytic, and reductive degradations of humic substances re lease a number of identifiable compounds. Because the products from these disparate processes are so similar, these compounds probably represent subunits of the humic macromolecule. The presence of highly oxidized com pounds, such as polycarboxylic phenols, in a reduction mixture corroborates information obtained from oxidative and hydrolytic studies. The supposition that these types of compounds exist as such in the original macromolecule is supported. The modes of attachment of these subunits to the parent molecule are not yet completely understood. However, a mechanistic inter pretation of the products formed from various degradation methods suggests that the major covalent linkages involved are ether and alkyl bonds activated by unsaturation, and esters. Much of the information gleaned from individual chemical degradation studies is qualitative, because of the low and variable product yields often obtained. More comprehensive product isolation and the performance of repetitive degradation experiments may be useful. These techniques could maximize yields and thereby minimize constraints on structural interpre tations. Comparison of the results obtained from a variety of methods, both instrumental and chemical, can provide new and corroborative structural information. Reductive degradation should be an important element i n a comprehensive effort to characterize humic structure.

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;


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44. 45. 46. 47. 48. 49. 50.


51. 52. 53. 54. 55. 56. 57. 58. 59. 60. 61.


RECEIVED for review August 19, 1987. ACCEPTED for publication December 4, 1987.


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Chemical Degradation of Humic Substances for Structural

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