Fluorescence detection and surface plasmon resonance spectroscopy could help make water safer. “Water, water, every where, nor any drop to drink,” wrote Samuel Taylor Coleridge more than two centuries ago. And nowadays, there is far less water available for a burgeoning global population, and what remains is increasingly likely to be chock-full of chemicals and other toxic materials no one should have to imbibe. In fact, water pollution is one of the problems that most affect human life, said Ibrahim Abdulhalim, professor and chairman of the electro-optic engineering department at Ben Gurion University of the Negev in Beersheba, Israel. “As time passes, the problem becomes more severe.” Chemists and engineers worldwide are looking into more and better ways to find new water sources, to filter available water for healthful consumption and to recycle wastewater before it re-enters rivers, lakes and oceans. Before you can use (or reuse) water, though, you must first know what’s in it that has to come out. Researchers continually refine fluorescence detection and surface plasmon resonance (SPR) spectroscopy to pinpoint dangerous chemicals and elements, with the ultimate hope of identifying toxins in water as close to their source as possible so that the water can be cleaned or the toxins prevented from entering the water sources in the first place. In this surface plasmon resonance (SPR) imaging setup, a monochromatic laser beam generates an evanescent wave at a single incidence angle on a thin layer of metal. When the momentum of the photons matches the momentum of the metal electrons, a dark line related to the surface plasmon resonance appears on the bright area of the captured images. Courtesy of SPIE. Endocrine disruptors have a strong effect on wildlife and are thought to have a telling effect on people as well, especially in children and infants. These chemicals generally act as hormone mimics that can cause undesired biochemical reactions in people and animals. For example, bisphenol A (BPA), which is used in the manufacture of some plastics, has come under scrutiny as an endocrine disruptor that may have an adverse health effect on young children exposed to it from drink bottles and microwavable dishes. High-performance liquid chromatography and gas chromatography/mass spectroscopy typically are used to determine the presence of BPA and other endocrine disruptors in food, water and urine, but they require lengthy sample preparation steps and large samples for testing. To shorten test time inexpensively, Abdulhalim and his Ben Gurion colleagues Alina Karabchevsky, Lev Tsapovsky and Robert S. Marks developed a biosensor based on SPR imaging principles. Silver or gold To create an SPR-based detection system, they deposited a 70-nm-thick layer of silver onto a glass slide, then topped it with a 21-nm-thick protective layer of SiO2. Using gold to amplify SPR signals is more common, but Abdulhalim’s group chose silver because it adheres better to glass, without additional materials. The SiO2 prevents any weakening of the glass-silver bond via interactions with oxygen, H2S or other substances found in natural water sources. After placing a sample (they tested both BPA and the hormone estrone) on the coated slide, they used a 637-nm laser from Edmund Optics to excite the silver, which, in turn, energized the sample. The resulting SPR signals, which were detected with a CCD sensor also from Edmund Optics, indicated the presence of either BPA or estrone. To highlight the SPR signal, and therefore gain sensitivity, the researchers used an algorithm based on a principle called the Radon transform, which calculates signal density along straight lines of different slopes, according to Abdulhalim. Besides BPA and estrone, the SPR-based technique is useful for detecting testosterone, drugs such as carbamazepine, and various bacteria and metals, Abdulhalim said. His group is working on a multichannel version of the SPR sensor, which would detect two or more polluting substances in water samples. “The possibility of miniaturizing an SPR sensing device, [its] low detection limit and [its] fast detection speed are among the main advantages [of the technique],” Abdulhalim said, “although there is a place for improving all these parameters.” He added that SPR sensor improvements will come through improved optics and improved system parameters, although most researchers choose to work on only one of these areas. Metallic silver is not used solely for coatings in SPR research. Silver nanoparticles, renowned for their antimicrobial and electrical properties, are used in many common consumer goods as well, from deodorants to sneakers and vacuum cleaners. But even as the material was gaining ground in everyday products, questions about its possible effects on human and animal health cropped up. Silver nanoparticles Currently, the total amount of silver found in natural water sources is about 0.2 to 0.3 µg/l. Water specifically treated with silver, for the purposes of destroying bacteria or algae, can harbor as much as 50 µg/l or more. Ongoing concern exists over the artificial increase in the production and use of the particles in households and hospitals, especially. Mining operations are a large source of silver particles in waterways, of course, but increasingly, so, too, are consumer products. Specially made odor-fighting socks contain silver nanoparticles, for example, but these leach out bit by bit with every washing, eventually finding their way to wastewater treatment plants and beyond. The effects that colloidal silver particles might have on the ecosystem is largely guessed at, but some researchers and government regulators fear that a large influx of such nanoparticles into rivers and streams could have a catastrophic effect on microbial populations, which would lead to a chain reaction affecting the entire food chain. Researchers at Florida Institute of Technology (FIT) in Melbourne wondered, however, whether the silver particles found in rivers and lakes necessarily were all a result of manufacturing and mining. Could some of it arise naturally? Some of the reflected signals captured by the camera are shown, with extracted surface plasmon resonance lines along the images. DL = experimental detection limit. Courtesy of SPIE. Humic acid is a prominent component in soil, peat and similar material through which water runs – from farms and mines, for example, on the way to rivers and streams. Humic acid, of which there are many varieties, originates through the decomposition of lignin and other plant materials. The material acts as a chemical-reducing agent, oxidizing to balance free ions it encounters. Virender K. Sharma of FIT and his colleagues sought to find what effect, if any, humic acid had on the formation of colloidal silver nanoparticles from silver ions in the soil. The group used an Agilent Technologies UV-VIS spectrometer to study the interactions of humic acid and silver ions in water from several natural sources, as well as at room temperature and at 90 °C. Ultimately, they found a characteristic surface plasmon resonance at about 400 nm that indicated the formation of silver nanoparticles in colloidal form. They found that the peak wavelength at which surface plasmon resonance occurs varies with the diameter of the forming nanoparticles as well as with the amount of agglomeration that has occurred at the time of measurement. They also saw that the peak spread a bit when the concentration of humic acid was low, but that that of silver ions remained steady, indicating that the individual particle sizes were dependent on the quantity of humic acid present. Tests at the higher temperature increased the rate at which the nanoparticles formed. The investigators propose that silver nanoparticles formed via humic acid are very stable for long periods and conceivably could be found far from their points of origin. It remains to be seen whether such stability is a good thing or an ecodisaster in the making. Finding mercury Even more troublesome than silver in the environment is mercury. The metal’s usefulness in fluorescent lightbulbs is only a recent, and small, sample of its omnipresence. The metal is, however, one of the most toxic materials one might encounter on any given day. In its monomethyl form, it is a powerful neurotoxin that can cause severe brain damage, yet it often is found in predatory fish that gobble up tiny prey before themselves turning up on our dinner plates. The US Environmental Protection Agency has set 0.3 µg/g as the maximum level of mercury allowable in fish, including freshwater and shellfish. Until now, however, the only way to determine the amount of mercury in fish tissue was through time-consuming lab tests. Signal interference resulting from saltwater makes testing more difficult in common tests that seek to separate the mercury from the tissue first. Combining bovine serum albumin, 3-mercaptopropionic acid and rhodamine 6G with gold nanoparticles produces a fluorescence-based probe that can ferret out mercury contamination in water. Reprinted with permission of Environmental Science & Technology. To develop a faster, yet still reliable, method of mercury testing – critical for timely analysis in the field – Chih-Ching Huang’s group at the National Taiwan Ocean University in Keelung looked to gold nanoparticles. The team concocted a fluorescence assay composed of bovine serum albumin (BSA) and rhodamine-6G bonded to gold nanoparticles via 3-mercaptopropionic acid with which to test solutions that were similar to the water that would be extracted from sea life. The BSA prevented the overaggregation normally caused by the high levels of salt in the samples. More importantly, when the gold nanoparticles made contact with mercury ions, they released rhodamine-6G molecules into the solution. The resulting increase in fluorescence emission indicated the presence of mercury in the sample. Surface plasmon resonance imaging can be used to search for multiple pollutants (or other materials) in water sources. Here, triple-channel SPR imaging of three analytes was conducted with a single sensor, showing oil, glue and deionized water (DI). (a) shows the original image in gray scale, and (b) shows the processed image with extracted locations of SPR lines. Courtesy of SPIE. Masking the presence of other heavy metals in the samples, using reagents such as ethylenediamine or nanowires made of tellurium increased the sensitivity of the researchers’ gold nanoparticle-based system. The investigators found that the system is useful for quickly determining the presence of mercury in seawater, and river and tap water, as well as in actual fish tissue. They also showed that the assay is effective for several forms of mercury, including phenylmercury and monomethylmercury, and that cadmium, lead and silver also may be tested with the technique. With such improvements in the ability to single out the most worrisome pollutants – and filtering techniques to match – the available amount of drinkable water is sure to rise.