Marine detection

ImmunO lSS iy. Chemiluminescence compounds (eg, acridinium esters and sulfonamides, isoluminol), luciferases (eg, firefly, marine bacterial, Benilla and Varela luciferase), photoproteins (eg, aequorin, Benilld), and components of bioluminescence reactions have been tested as replacements for radioactive labels in both competitive and sandwich-type immunoassays. Acridinium ester labels are used extensively in routine clinical immunoassay analysis designed to detect a wide range of hormones, cancer markers, specific antibodies, specific proteins, and therapeutic dmgs. An acridinium ester label produces a flash of light when it reacts with an alkaline solution of hydrogen peroxide. The detection limit for the label is 0.5 amol. 

Other appHcations of firefly hioluminescence include measurement of the activity of bacteria in secondary sewage treatment activated sludge (296,297), detection of bacteria in clean rooms and operating rooms, measurement of bacteria in bottled foods, beverages (298), and pharmaceuticals (299), determination of the antimicrobial activity of potential dmgs (300), determination of the viabiHty of seeds (301), and measuring marine biomass concentrations as a function of ocean depth or geographical location (302). 

Numerous high pressure Hquid chromatographic techniques have been reported for specific sample forms vegetable oHs (55,56), animal feeds (57,58), seta (59,60), plasma (61,62), foods (63,64), and tissues (63). Some of the methods requite a saponification step to remove fats, to release tocopherols from ceHs, and/or to free tocopherols from their esters. AH requite an extraction step to remove the tocopherols from the sample matrix. The methods include both normal and reverse-phase hplc with either uv absorbance or fluorescence detection. AppHcation of supercritical fluid (qv) chromatography has been reported for analysis of tocopherols in marine oHs (65). 

A number of other chemicals suspected of having endocrine disrupting potential also occur at high levels in the tissues of marine mammals. For example, tribiityltin compounds are present in the tissues of the Steller sea lion (Eumetopias juhatus) from Hokaido, Japan, and in stranded bottlenose dolphins (Tursiops truncatus) found along the US Atlantic and Gulf coasts.Additional chemicals detected include PAHs, toxaphene and chlordane. ... 

Fig.7-10. Separation of amino acids after derivatization with OPA and A -isobu-tyryl-L-cysteine. Column Superspher 100 RP-18 (4 pm) LiChroCART 125-4, mobile phase 50 mM sodium acetate buffer pH 7.0/sodium acetate buffer pH 5.3/methanol, flowrate 1.0 ml min temperature 25 °C detection fluorescence, excitation 340 nm/emission 445 nm. Sample amino acid standard mixture. (Merck KGaA Application note W219189 reproduced with permission from H. P. Fitznar, Alfred-Wegener-Institute for Polar and Marine Research.)...

Quality control tests are intended to detect produced materials which deviate from manufacturing specifications, and thus may result in questionable performance. The materials are usually subjected to spectrographic analysis which is the primary quality control check. The exposure tests are necessarily of short duration (hours or days), in which the test conditions attempt to reflect the environment of operation, for example using artificial seawater for a marine application. Since a property that is reproducible and indicative of a consistent quality anode is all that is required, there is no attempt to mirror, except in the crudest fashion, current density profiles. 

Widder, E. A., Latz, M. I., and Case, J. F. (1983). Marine bioluminescence spectra measured with an optical multichannel detection system. Biol. Bull. 165 791-810. 

A second source of plutonium, dispersed more locally, is liquid effluent from fuel reprocessing facilities. One such is the fuel reprocessing plant at Windscale, Cumbria in the United Kingdom where liquid waste is released to the Irish Sea(6). Chemical analysis of this effluent shows that about one percent or less of the plutonium is in an oxidized form before it contacts the marine water(7). Approximately 95 percent of the plutonium rapidly adsorbs to particulate matter after discharge and deposits on the seabed while 5 percent is removed from the area as a soluble component ). Because this source provided concentrations that were readily detected, pioneering field research into plutonium oxidation states in the marine environment was conducted at this location. 

The atmosphere may be an important transport medium for many other trace elements. Lead and other metals associated with industrial activity are found in remote ice caps and sediments. The transport of iron in wind-blown soil may provide this nutrient to remote marine areas. There may be phosphorus in the form of phosphine, PH3, although the detection of volatile phosphorus has not been convincingly or extensively reported to date. 

The use of high performance liquid chromatography (HPLC) for the study of paralytic shellfish poisoning (PSP) has facilitated a greater understanding of the biochemistry and chemistry of the toxins involved. HPLC enables the determination of the type and quantity of the PSP toxins present in biological samples. An overview of the HPLC method is presented that outlines the conditions for both separation and detection of the PSP toxins. Examples of the use of the HPLC method in toxin research are reviewed, including its use in the determination of the enzymatic conversion of the toxins and studies on the movement of the toxins up the marine food chain. 

Experimental evidence indicates that many marine bacteria produce TTXs. However, TTX production by some bacteria has not been validated since TTX and anhydro-like TTX are described as "difficult to detect" by using HPLC and GC-MS methods, and show no activity in the mouse bioassay. 

Bioluminescence can be used for spedfic detection of separated bioactive compounds on layers (BioTLC) [46]. After development and drying the mobile phase by evaporation, the layer is coated with microorganisms by immersion of the plate. Single bioactive substances in multicomponent samples are located as zones of differing luminescence. The choice of the luminescent cells determines the specificity of detection. A specific example is the use of the marine bacterium Vibrio fischeri with the BioTLC format. The bioluminescence of the bacteria cells on the layer is reduced by toxic substances, which are detected as dark zones on a fluorescent background. BioTLC kits are available from ChromaDex, Inc. (Santa Ana, CA). 

Other kinds of bloassays have been used to detect the presence of specific allelochemical effects (8), effects on N2 fIxatlon (9), the presence of volatile compounds (10) and of Inhibitory substances produced by marine microalgae (11). Putnam and Duke (12) have summarized the extraction techniques and bioassay methods used In allelopathy research. Recent developments In high performance liquid chromatography (HPLC) separation of allelochemlcals from plant extracts dictates the need for bloassays with sensitivity to low concentrations of compounds contained In small volumes of eluent. Einhellig at al. (13) described a bloassay using Lemna minor L. growing In tissue culture cluster dish wells that maximizes sensitivity and minimizes sample requirements. 

Some typical applications in SFE of polymer/additive analysis are illustrated below. Hunt et al. [333] found that supercritical extraction of DIOP and Topanol CA from ground PVC increased with temperature up to 90 °C at 45 MPa, then levelled off, presumably as solubility became the limiting factor. The extraction of DOP and DBP plasticisers from PVC by scC02 at 52 MPa increased from 50 to 80 °C, when extraction was almost complete in 25 min [336]. At 70 °C the amount extracted increased from 79 to 95 % for pressures from 22 to 60 MPa. SFE has the potential to shorten extraction times for traces (<20ppm) of additives (DBP and DOP) in flexible PVC formulations with similar or even better extraction efficiencies compared with traditional LSE techniques [384]. Marin et al. [336] have used off-line SFE-GC to determine the detection limits for DBP and DOP in flexible PVC. The method developed was compared with Soxhlet liquid extraction. At such low additive concentrations a maximum efficiency in the extractive process and an adequate separative system are needed to avoid interferences with other components that are present at high concentrations in the PVC formulations, such as DINP. Results obtained... 

Applications Off-line SFE-GC-MS has been used for the analysis of additives in polyurethanes [52], Marin et al. [53] have used off-line SFE-GC to determine the detection limits for DOP and DBP in flexible PVC. The method was compared with Soxhlet liquid extraction. 

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