Paleoecological considerations of the sediment-water interface of Florida Bay as derived from chlorophylls, chlorophyll derivatives, and carotenoids.

Paleoecological Considerations of the Sediment-Water Interface of Florida Bay as Derived from Chlorophylls, Chlorophyll Derivatives, and Carotenoids.

Topical Area: Paleoecology

J. William Louda, Joseph W. Loitz, Earl W. Baker, Organic Geochemistry Group, Department of Chemistry and Biochemistry, Florida Atlantic University, Boca Raton, FL; David T. Rudnick, South Florida Water Management District, West Palm Beach, FL

There exists much popular and scientific literature which details changes in the condition of Florida Bay. Natural fluctuations in the myriad of biomes which comprise the overall ecosystem of the Bay are likely overlaid, and may be overwhelmed, by perturbations due to the activities of man. A few of the better known more widely publicized anthropogenic influences include: the construction of the Flagler railway system, now the "Overseas Highway", which included much inter-key pass filling; increasing development of the Keys with concomitant increases in sewerage seepage from antiquated septic systems emplaced in highly porous limestones; and drastic alteration of the timing, volume and locations of the fresh water discharge from the Kissimmee /Lake Okeechobee/ Everglades watershed into the Bay. Ostensibly, as results of these changes, potentially coupled with natural cycles, certain problems in the ecology of the Bay have occurred. Specifically, salinity increased in certain regions, sea grasses became infested with a slime mold (Labyrinthula sp) and/or died, and algal blooms, notably of high cyanobacterial biomass, have occurred (e.g. 2^nd Florida Bay Science Conference, 1996).

The lipophilic pigments, the chlorophylls and carotenoids are intimately linked to the process of photosynthesis. These pigments either directly participate in the energy absorption process (e.g. chlorophyll-a, bacteriochlorophyll-a; fucoxanthin, et cetera), scavenge activated oxygen species (anti-photodynamic activity: e.g. violaxanthin/antheraxanthin, diatoxanthin/ diadinoxanthin), or serve as direct sunscreens (e.g. zeaxanthin).

Within the last decade or so, a strong resurgence in the potential of the photosynthetic pigments as chemotaxonomic markers has occurred. The ability to separate and identify these pigments, ranging from the highly polar carboxylic acids (e.g. chlorophyllides, chlorophylls-c) to the very non-polar terpenoid esters (e.g. pheophytins), in a single analysis has indeed opened the door to pigment based chemotaxonomy. The present studies are ultimately aimed at using the chlorophylls, chlorophyll derivatives, carotenoids and their relationships to authochthonous organic matter to assess present and past ecosystems in qualitative and quantitative manners. Therefore, we address the present photoautotrophic biota using pigment-based chemotaxonomy and, using this and other data, apply pigment analysis to sedimentary organic matter in attempts to reveal past ecologies, both qualitatively ('taxa') and quantitatively ('paleoproductivity').

Samples investigated during the present studies included: cultured phytoplankton from Florida Bay (Synechococcus elongatus, Cyclotella choctawathceeana, the "2-micron picosphere": ex. C. Tomas); water samples forming a transect across Florida Bay (ex. Florida Bay Interlaboratory Chlorophyll Calibration Study, W. Kruczynski); macroalgal and higher plant samples (e.g.Thalassia testudinum); and 4-inch sediment cores ( 0.6-1.2 meters) collected in liaison with the USGS (W.Orem, E.Shinn, C.Holmes, R.Halley, S.Ishman). Sample handling, preparation and analyses were made in dim light to darkness with oxygen and heat excluded as possible. Seston (> 0.45 mm: GF/F), phytoplankton and macroalgae/higher plant material was extracted with 90% acetone(aq.). Sediments were extracted with 100% tetrahydrofuran, shown in our laboratory to be 6-12x as effective in the extraction of pigments from Florida Bay marls relative to the more common solvents (acetone, methanol).

In order to estimate the (Divisional) taxonomic makeup of a population of oxygenic photoautotrophs, one employs the in vivo molar ratios of chlorophyll-a (CHL-a) to a selected taxon specific pigment biomarker. For example, on a world-wide basis, the CHL-a/fucoxanthin (FUCO) value of diatoms (DIATS) averages about 1.2:1. Thus, if one finds 1 mole of FUCO, 1.2 moles of chlorophyll-a can reasonably be expected. Extension of this principle with markers for chlorophytes ( CHL-a / CHL-b~ 3.6), dinoflagellates ( CHL-a / peridinin [PERI] ~ 2.3), and cyanobacteria (CHL-a/ zeaxanthin [ZEA] ~ 5.0, CHL-a/ myxoxanthophyll [MYXO] ~ 5-12) can then lead to regression formula with which to estimate the divisional makeup of a community. In the present study, we investigated 3 major phytoplankton species, given above, from Florida Bay waters and adjusted our existing regression formulae with site specific data. First, the "2 micron picosphere" [2m-PICO"] was shown to be a cyanobacterium by the presence of MYXO, ZEA, CHL-a, echinenone and b-carotene. S. elongatus was found to contain only ZEA as a possible chemotaxonomic marker. The CHL-a to ZEA and MYXO values for "2m-PICO" were 13.88 and 8.87, respectively (ZEA/MYXO=1.56). S. elongatus had CHL-a/ZEA = 4.16. Here, we choose to estimate the 2 populations separately. That is, the "2m-PICO" would be estimated from MYXO directly. However, S. elongatus and similar non-MYXO containing cyanobacteria, would be estimated from ZEA after correcting for a contribution by MYXO-containing species. Thus, a 'corrected ZEA [ZEA'] was defined here as [ZEA'] = ([ZEA]- 1.56x[MYXO]). Diatoms would be estimated from C. choctawatcheeana CHL-a/FUCO values ( 0.91) which were corrected slightly upwards (1.1) to allow for a wider distribution of diatom species. In the end, we arrived at the following regression formula for use in Florida Bay waters.

TOTAL = CYANOBACTERIA + CHLOROPHYTES + DIATOMS + DINOFLAGELLATES

S = (7.5x[MYXO]+5.0[ZEA']) + (3.6x[CHL-b]) + (1.1x[FUCO]) + ( 2.3x[PERI])

Analyses of 3 water samples forming a SE-NW transect across Florida Bay formed the first test of this formula. The samples analyzed were from Twin Key Basin (TKB), Rabbit Key Basin (RKB), and Sandy Key Basin (SKB). Following HPLC/PDA analysis of the pigments in each sample, conversion to molar values and application of the formula, we found that these 3 samples contained cyanobacteria / diatoms / dinoflagellates in the following ratios: TKB= 58/41/01, RKB= 76/24/00, and SKB= 36/48/16, for samples collected on 11/20/96. This distribution matches fairly closely the reported cyanobacterial spread across the Bay and even reflects the more or less open exchange with oceanic flora at the peripheral sites, SKB and TWB, respectively. That is, the most diatoms and dinoflagellates, summing to more than the cyanobacterial population, were found at the SKB site which is in contact with the open Gulf of Mexico.

Analysis of fresh Thalassia testudinum revealed the expected 'higher plant' pigments, namely CHL-a (53%), CHL-b (13%), lutein (LUT: 21%), b-carotene (3%), and neo-/viola-/anthera-xanthins (9%). Analysis of brown turtle grass collected on the sediment surface revealed a pattern of pigments unmistakably belonging to diatoms (CHLs-c₁/-c₂, FUCO, diadinoxanthin) and cyanobacteria (lg.amts. ZEA), in that order, with only a trace of lutein remaining. CHL-b was totally absent. It appears that diatoms and cyanobacteria form a periphytonous growth on/in the dead Thalassia leaves. Whether the species are different than the truly pelagic and benthic forms in the Bay would require examination by those trained in phytoplankton systematics.

Analyses of sediments revealed that the surficial sediments (e.g.0-2 cm) contained a diatomaceous cyanobacterial biofilm / mat underlain by purple-S bacteria. Large amounts of ZEA indicate a predominance of cyanobacteria in the surface sediments. Chlorophylls-c₁/-c₂,, FUCO, diadinoxanthin (DDX), and fucoxanthinol (F-ol), the most common degradation product of FUCO are all prevalent in surficial sediments yet are entirely absent after only about 5-10 cm additional burial. Purple-S bacteria were evidenced by the presence of bacteriochlorophyll-a (BCHL-a), bacteriopheophytin-a (BPPT-a), and 2 isomers of spirilloxanthin.

Certain diagenetic alterations can be followed downhole in the lime-marls of Florida Bay. All BCHL-a converts to BPPT-a and pyroPPT-a. All polar pigments required for the chemotaxonomic assessment of diatoms (CHLs-c₁/-c₂; FUCO, DDX, F-ol) are destroyed or complexed into intractable geopolymers. ZEA and, to a certain extent, LUT are preserved downhole. A gradual downhole generation of the cis-isomeric forms of ZEA and b-carotene was observed. The most dramatic alteration occurred within the chlorophyll-a population. That is, some CHL-a was found to be converted to pheophytin-a(PPT-a), its epimer (PPT-a') and eventually to pyroPPT-a. However, the majority of CHL-a was found to funnel through pyropheophorbide-a (pPPB-a) and into, via a (Dieckman-like) dehydration reaction, cyclopheo- phorbide-a-enol. This non-fluorescent compound, in which the 17-propionic acid moiety forms a C-C bound, via water loss, with the 13²-C on the carbocyclic ring, is the overwhelmingly most abundant CHL-a derivative in the lime-marls of Florida Bay.

In general, the downhole distribution of pigments and their relationships to a widely fluctuating

organic carbon content (2-6%,dry wt.) indicates a qualitatively time-stable ecosystem which reacts quantitatively to water column productivity and nutrient flux.

This study covers a bit under 2 funded years. Analyses of remaining samples and data reduction continues. Future directions include identification of certain unknown minor components, mainly carotene-diols and hydroxy-carotenones, and estimation of oxygenic versus anoxygenic inputs to the sedimentary organic matter.