Organic Marine Geochemistry - American Chemical Society

7 The Biogeochemistry of Chlorophyll J. William Louda and Earl W. Baker

Downloaded by UNIV OF MISSOURI COLUMBIA on November 27, 2013 | http://pubs.acs.org Publication Date: April 21, 1986 | doi: 10.1021/bk-1986-0305.ch007

Organic Geochemistry Group, Florida Atlantic University, Boca Raton,FL33431

In this paper we attempt a first approximation of chlorophyll diagenesis, stressing the generation of DPEP-series geoporphyrins. Samples analyzed include viable and dead unialgal cultures, sediment traps, surface to near-surface (0-2m) sediments and long cores (5-1,000m) obtained from DSDP/IPOD and industry. Results reveal that chlorophyll(-a) loses Mg and phytol through the actions of cellular senescence and autotrophic recycling in the water column and surface sediments. Following deposition these 'pheo-pigments' undergo the competing reactions of allomerization, yielding purpurin and chlorin acids in oxic conditions, or loss of the 10-carbomethoxy moiety, forming pyro-phorbides in anoxic settings. These key reactions appear to fate subsequent diagenesis to either pigment destruction or fossilization, respectively. DPEP-series geoporphyrins are thought to be the result of the stepwise defunctionalization and aromatization of the pyro-phorbide type precursors. The phenomenology of chlorophyll geochemistry and tentative identification of several intermediates are described. The present study is based on the assumption that geoporphyrins are the diagenetic products of biotic tetrapyrrole pigments. This we take as a reasonable premise, given the structural complexity of this class of biomarkers (see Figure 1), and was concluded years ago by the late Professor Alfred Treibs (1-5). However, the development of strong precursor-product relationships in organic geochemistry requires not only statement of plausible end-members (e.g. chlorophyl1-a and DPEP-series geoporphyrins) but must include description of the intervening reactions and intermediate structures. These investigations, albeit preliminary, are designed to ultimately f i l l the existing gaps in knowledge between biotic 0097-6156/ 86/0305-0107506.00/ 0 © 1986 American Chemical Society

In Organic Marine Geochemistry; Sohn, M.; ACS Symposium Series; American Chemical Society: Washington, DC, 1986.

108

O R G A N I C M A R I N E GEOCHEMISTRY

structures (6-8) and the growing data base of product ( v i z . geoporphyrins) structures (9-17).


Experimental Samples. Unialgal cultures of the diatom Synedra sp. were purchased from Carolina B i o l o g i c a l Supply. Sediment trap samples and near surface sediments (0-2m) from the Peruvian upwelling region, c o l l e c t e d during Cruise 73 Leg 2 of the R/V Knorr, were provided by Woods Hole Océanographie I n s t i t u t i o n (18-19). Nearsurface sediments from the Guaymas Basin i n the Gulf of C a l i f o r n i a , collected by the R/V Washington Leg 3-1978, were provided by Scripps I n s t i t u t e of Oceanography (20). Sediments and sapropel from Big Soda Lake (Nevada, U.S.A.. 21_) and Mangrove Lake (Bermuda. 24) were obtained from the United State Geological Survey. Deeply buried (e.g. 15-1000 m., sub-bottom) marine sediments were obtained from the DSDP/IP0D program, Leg 64 (20-23). An immature marine shale of C a l i f o r n i a (Pliocene/ Miocene. 24·) was provided by Mobil Research and Development. Except for viable diatoms, a l l samples were frozen upon c o l l e c t i o n and maintained so u n t i l e x t r a c t i o n . Solvents, Extraction and Separation of Pigments. A l l solvents were freshly glass d i s t i l l e d and ethers were freed of peroxides over highly activated (0% H2O) basic alumina. A l l procedures were i n dim yellow l i g h t and extracts/isolates maintained frozen under N2, whenever p o s s i b l e . Chromatographic separation u t i l i z e d m i c r o c r y s t a l l i n e c e l l u l o s e , Sephadex LH-20, and s i l i c a gel in normal and reverse phase modes. Extraction and chromatography i s detailed elsewhere (20). P u r i f i c a t i o n and I d e n t i f i c a t i o n of Isolates. In the case of extremely immature samples containing bacteriophytin-a (e.g. Big Soda Lake, Mangrove Lake), i n t e r f e r i n g carotenols were removed via phase separation into 90% aqueous methanol (25-26). Low-pressure high-performance liquid-chromatography (LPHLC, Ace-Glass) using 13-24 s i l i c a i n normal (Whatman #LPS-1) and C-18 reverse phase (Whatman #LRP-1) modes was employed for f i n a l pigment p u r i f i c a t i o n and co-chromatographic t e s t s . Isoc r a c t i c elution employed methanol/acetone/water (90:5:5, v/v/v) f o r pheophorbide free acids and methanol/acetone (95:5 v/v) f o r pheophytins during RP-LPHPLC. The presence of more than trace (ca 0 . 5 - 1.0%) water in the reverse phase mode lead to exceedingly long elution times (> 1 nr.) with phytylated pigments. This fact served as a t e s t f o r the presence of the phytyl ester. Non-polar pigments, pheophytins and decarboxylated species, were p u r i f i e d over normal phase s i l i c a with increasing percentages of acetone in petroleum ether. Mass spectrometry was performed on a DuPont #21-491B i n s t r u ment operated at the lowest i o n i z i n g voltage (e.g. 4.5-12.0 eV, 40-60 A) possible per sample. E l e c t r o n i c absorption spectra were recorded on a PerkinElmer 575 instrument calibrated with hoi mi urn oxide. The absorpt i o n spectra of native pigments, sodium borohydride reduction


7.

LOUDA AND BAKER

Biogeochemistry of Chlorophyll

109

products (27-28), and the copper chelates of each (20) were compared to numerous authentic chlorophyll derivatives"Tstandards) i n order to c l a s s i f y the chromophore and i t s auxochromes.


Results and Discussion Previously we have divided tetrapyrrole diagenesis into e a r l y - , middle-and late-stages (4-5). These d i v i s i o n s encompass the defunctionalization of dihydroporphyrins (e.g. phorbides), aromatization and c h e l a t i o n , in that order. The geoporphyrins formed during mid-and late-diagenesis are therefore free-base and metal 1o(e.g. N i ; , V = 0) porphyrins, respectively. Since t h i s study deals primarily with early-diagenesis, the following section is presented only to reveal the nature of these i n i t i a l l y formed geoporphyrins. Products of Diagenesis, Immature Geoporphyrins. The progress of the tetrapyrrole diagenetic continuum i s such that the a r b i t r a r i l y defined stages (4-5) can and do overlap. Thus, i t i s often p o s s i ble to i s o l a t e more than one pigment type (e.g. free-base and metallo-porphyrins) from the same stratum. Shown in Figure 2 are the mass spectral histograms, or carbon-number d i s t r i b u t i o n , of (a) the defunctionalized phorbides ( i . e . 7,8-dihyroDPEP-series), (b) the free-base DPEP series and (c) the nickel DPEP-series isolated from a Miocene/Pliocene marine shale of C a l i f o r n i a . While not t o t a l l y i d e n t i c a l , maxima at C31 and the ranges of pseudohomologs (C27 to C34) reveal t h e i r s i m i l a r o r i g i n . Past studies (4,5,29) have shown that the DPEP- series can be geochemically generated with C30, C31, or C32 maxima. These pigments, DPEP-series porphyrins with a limited carbon number range, represent early diagenetic end products. The metalloporphyrins ( v i z . n i c k e l , vanadyl) c h a r a c t e r i s t i c of catagenesis appear to form via p a r a l l e l diagenesis with the exceptions of the degree of a l k y l a t i o n and the immediate organic environment. That i s , carbon-numbers up to C40, C50 and beyond e x i s t in the vanadyl porphyrins (5,30-33) and these pigments appear to a r i s e from an inextractable organically bound state (4,29,34,35). An example of the series and carbon-number d i s t r i bution of the vanadyl porphyrins from a 'moderately mature' petroleum i s given as Figure 3. The complexity of geoporphyrin arrays becomes evident upon examination of Figure 3. That i s , since t h i s spectrum was averaged from low voltage (4.5eV) scans y i e l d i n g only parent ions, at least twenty-nine compounds (14 DPEP and 15 ETIO), not counting isomers which are known to exist (13-15,9-12), must be present. Re-examination of Figure 2 reveals that t h i s spreading of carbon number d i s t r i b u t i o n s amongst the tetrapyrroles probably begins early in the diagenesis of these pigments and occurs within the free or solvent-extractable species (e.g.Ni porphyrins), as well as the presumably bound forms thought to y i e l d the vanadyl pigments. The maturational aspects of metalloporphyrins are covered elsewhere (6-7). Phytoplankton cultures (Synedra, Bacillariophyceae) and water-column d e t r i t u s (sediment trap samples) were analyzed in order to describe the pre-depositional a l t e r a t i o n of chlorophyll and therein type the immediate precursor complement to early diagenesis.



110

ORGANIC MARINE GEOCHEMISTRY

a)PHORBIDES.DPEPsehes: (RING-Ε PRESENT)/C# Pheophytin-a PPa Ρ Pa-allomer 7E7DP-PYRO-PPa 9-OD-7E7DP-PYRO-PPa MESO-PYRO-PPa DOMPPa DPE(78dideH) DPEP(7,8dkJeH) BacteriopheoptTytin-a (3/Wihydro) flCHLORI NS.ETIO-series: (RING-Ε ABSENT)/C# PUR PURIN-18 PURPURIN- 7 CHLORIN-e6 CHLORIN-p6 ETKDPORPHYRIN-III (78dideH)

R1 2

R2 7_

V V V

R4 9

R3 10

Pr-phy KCOOCH3 Pr H.COOCH3 Pr HQCOOCH3

V V Ε Ε Ε Ε

Ε Ε Pr Pr Pr Ε

COCH3 Pr-phy

R1 2 V V V V E E

=0 =0 =0

Η, Η KH Η, Η Η,Η Η,Η Η,Η

=0 Η,ΟΗ =0 Η,Η Η,Η Η,Η

HCOOCH3

=0

R3 R4 R2 7 6 Pr 0=C—Ο—OO Pr COCOOH COOH Pr CH^COOH COOH Pr COOH COOH H E

Figure 1. Structures of tetrapyrrole pigments mentioned in text. Code: V=vinyl; E=ethyl; Pr=propionic a c i d ; Phy=phytol (as phytyl e s t e r ) ; DP=despropio-; PD=oxydeoxo; pp-a=pheophorbide-a ; D0MPP-a= deoxomesopyropheophorbi de-a; DPE=deoxophyl1oerythri η ; DPEP=deoxophylloerythroetioporphyrin ( c f . ^ ]_). 9

26 28 3 0 32 34 CARBON NUMBER

Figure 2. Mass spectral histograms of tetrapyrrole pigments c h a r a c t e r i s t i c of mid-/late-diagenesis. (a) free-base 7,8-dihydro-DPEP-series; (b) free-base DPEP-series; and (c) nickel DPEP (ETIO- omitted)-series. Sample; Pliocene/Miocene shale of marine o r i g i n ( c f . 24).


7.

Biogeochemistry of Chlorophyll

LOU DA AND BAKER

Phytoplankton Cultures. A viable unialgal culture of the diatom Synedra sp. was s p l i t i n t o 2 aliquotes. The f i r s t portion (VIABLE") was extracted immediately and analyzed. The second part ("DEAD") was purged with nitrogen, sealed and stored in the dark at room temperature (20-22°C) for 2 months before analysis. The e l e c t r o n i c spectra of the crude extracts of "VIABLE" and "DEAD" diatoms, given as Figure 4, reveals the t o t a l conversion of chlorophyll-a to pheopigments through the loss of Mg. That i s , Soret absorption has s h i f t e d hypsochromically (429 to 411 nm) i n concert with a bathchromic s h i f t (663 to 667 nm) i n the position of band I ('red') absorption. Chromatographic analyses revealed that 94+% of the a-series pigments i n the "VIABLE" diatoms was c h l o r o p h y l l - a . This pigment was below detectable l i m i t s (

Organic Marine Geochemistry - American Chemical Society

Recommend Documents