Support of the Geoluminescence Dating Research Laboratory has been provided by Baylor University and grants from the National Science Foundation (GSS-166023), National Geographic (#9990-1), and Atlas Sand. Discovery and learning in this lab were enhanced by our many collaborators, students, and visitors that have brought new perspectives, ideas, and approaches.

<ol>
	<li>Murray, A. S., Wintle, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+single+aliquot+regenerative+dose+protocol:+Potential+for+improvements+in+reliability.">The single aliquot regenerative dose protocol: Potential for improvements in reliability.</a> Radiation Measurements. 37 (4-5), 377-381 (2003).</li><li>Wintle, A. G., Murray, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+review+of+quartz+optically+stimulated+luminescence+characteristics+and+their+relevance+in+single-aliquot+regeneration+dating+protocols.">A review of quartz optically stimulated luminescence characteristics and their relevance in single-aliquot regeneration dating protocols.</a> Radiation Measurements. 41, 369-391 (2006).</li><li>. RISO Luminescence Analyst, Version 4.57 Available from: <a target="_blank" href="https://users.aber.ac.uk/ggd">https://users.aber.ac.uk/ggd</a> (2021)</li><li>Aitken, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> An introduction to optical dating: the dating of Quaternary sediments by the use of photon-stimulated luminescence. , (1998).</li><li>Duller, G. A. T., Wintle, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+review+of+the+thermally+transferred+optically+stimulated+luminescence+signal+from+quartz+for+dating+sediments.">A review of the thermally transferred optically stimulated luminescence signal from quartz for dating sediments.</a> Quaternary Geochronology. 7 (1), 6-20 (2012).</li><li>Brown, N. D., Forman, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluating+a+SAR+TT-OSL+protocol+for+dating+fine-grained+quartz+within+Late+Pleistocene+loess+deposits+in+the+Missouri+and+Mississippi+river+valleys,+United+States.">Evaluating a SAR TT-OSL protocol for dating fine-grained quartz within Late Pleistocene loess deposits in the Missouri and Mississippi river valleys, United States.</a> Quaternary Geochronology. 12, 87-97 (2012).</li><li>Forman, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Luminescence+Dating+in+Paleoseismology.">Luminescence Dating in Paleoseismology.</a> Encyclopedia of Earthquake Engineering. , 1371-1378 (2015).</li><li>Lepper, K., McKeever, S. W. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+objective+methodology+for+dose+distribution+analysis.">An objective methodology for dose distribution analysis.</a> Radiation Protection Dosimetry. 101 (1-4), 349-352 (2002).</li><li>Tsukamoto, S., Duller, G. A. T., Wintle, A. G., Muhs, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessing+the+potential+for+luminescence+dating+of+basalts.">Assessing the potential for luminescence dating of basalts.</a> Quaternary Geochronology. 6, 61-70 (2011).</li><li>Li, S. H., Chen, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Studies+of+thermal+stability+of+trapped+charges+associated+with+OSL+from+quartz.">Studies of thermal stability of trapped charges associated with OSL from quartz.</a> Journal of Physics D-Applied Physics. 34 (4), 493-498 (2001).</li><li>Fu, X., Li, S. H., Li, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optical+dating+of+aeolian+and+fluvial+sediments+in+north+Tian+Shan+range,+China:+Luminescence+characteristics+and+methodological+aspects.">Optical dating of aeolian and fluvial sediments in north Tian Shan range, China: Luminescence characteristics and methodological aspects.</a> Quaternary Geochronology. 30, 161-167 (2015).</li><li>Hu, G., Li, S. -. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Simplified+procedures+for+optical+dating+of+young+sediments+using+quartz.">Simplified procedures for optical dating of young sediments using quartz.</a> Quaternary Geochronology. 49, 31-38 (2019).</li><li>Porat, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+magnetic+separation+for+purifying+quartz+for+luminescence+dating.">Use of magnetic separation for purifying quartz for luminescence dating.</a> Ancient TL. 24 (2), 33-36 (2006).</li><li>Mejdahl, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thermoluminescence+dating+of+sediments.">Thermoluminescence dating of sediments.</a> Radiation Protection Dosimetry. 17, 219-227 (1986).</li><li>Fain, J., Soumana, S., Montret, M., Miallier, D., Pilleyre, T., Sanzelle, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Luminescence+and+ESR+dating-Beta-dose+attenuation+for+various+grain+shapes+calculated+by+a+Monte-Carlo+method.">Luminescence and ESR dating-Beta-dose attenuation for various grain shapes calculated by a Monte-Carlo method.</a> Quaternary Science Reviews. 18, 231-234 (1999).</li><li>Prescott, J. R., Hutton, J. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cosmic+ray+contributions+to+dose+rates+for+luminescence+and+ESR+dating:+large+depths+and+long-term+time+variations.">Cosmic ray contributions to dose rates for luminescence and ESR dating: large depths and long-term time variations.</a> Radiation Measurements. 23, 497-500 (1994).</li><li>Peng, L., Forman, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=LDAC:+An+excel-based+program+for+luminescence+equivalent+dose+and+burial+age+calculations.">LDAC: An excel-based program for luminescence equivalent dose and burial age calculations.</a> Ancient TL. 37, 2 (2019).</li><li>Lafuente, B., Downs, R. T., Yang, H., Stone, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+power+of+databases:+the+RRUFF+project.">The power of databases: the RRUFF project.</a> Highlights in Mineralogical Crystallography. , 1-30 (2015).</li><li>Wintle, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Luminescence+dating:+laboratory+procedures+and+protocols.">Luminescence dating: laboratory procedures and protocols.</a> Radiation Measurements. 27, 769-817 (1997).</li><li>Aitken, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Thermoluminescence Dating. , (1985).</li><li>Porat, N., Faerstein, G., Medialdea, A., Murray, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Re-examination+of+common+extraction+and+purification+methods+of+quartz+and+feldspar+for+luminescence+dating.">Re-examination of common extraction and purification methods of quartz and feldspar for luminescence dating.</a> Ancient TL. 33 (1), 22-30 (2015).</li><li>And&#242;, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gravimetric+Separation+of+Heavy+Minerals+in+Sediments+and+Rocks.">Gravimetric Separation of Heavy Minerals in Sediments and Rocks.</a> Minerals. , 15 (2020).</li><li>Nelson, M., Rittenour, T., Cornachione, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sampling+methods+for+luminescence+dating+of+subsurface+deposits+from+cores.">Sampling methods for luminescence dating of subsurface deposits from cores.</a> Methods and Protocols 2. 88, 1-15 (2019).</li><li>Munsell Color. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Munsell Soil Color Charts: with Genuine Munsell Color Chips. , (2010).</li><li>USDA Natural Resources Conservation Service Soils. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Keys to soil taxonomy. 20, (2014).</li><li>User Manual. Malvern Instruments Ltd Available from: <a target="_blank" href="https://www.malvernpanalytical.com/en/products/product-range/mastersizer-range/mastersizer-3000">https://www.malvernpanalytical.com/en/products/product-range/mastersizer-range/mastersizer-3000</a> (2013)</li><li>Wentworth, C. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+scale+of+grade+and+class+terms+for+clastic+sediments.">A scale of grade and class terms for clastic sediments.</a> Journal of Geology. 30 (5), 377-392 (1922).</li><li>Mejdahl, V., Christiansen, H. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Procedures+used+for+luminescence+dating+of+sediments.">Procedures used for luminescence dating of sediments.</a> Boreas. 13, 403-406 (1994).</li><li>Madsen, A. T., Duller, G. A. T., Donnelly, J. P., Roberts, H. M., Wintle, A. G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronology+of+hurricane+landfalls+at+Little+Sippewisset+Marsh,+Massachusetts,+USA,+using+optical+dating.">Chronology of hurricane landfalls at Little Sippewisset Marsh, Massachusetts, USA, using optical dating.</a> Geomorphology. 109, 36-45 (2009).</li><li>Durcan, J. A., Duller, G. A. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+fast+ratio:+A+rapid+measure+for+testing+the+dominance+of+the+fast+component+in+the+initial+OSL+signal+from+quartz.">The fast ratio: A rapid measure for testing the dominance of the fast component in the initial OSL signal from quartz.</a> Radiation Measurements. 46, 1065-1072 (2011).</li><li>Wang, Y., Chen, T., Chongyi, E., An, F., Lai, Z., Zhao, L., Liu, X. -. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quartz+OSL+and+K-feldspar+post-IR+IRSL+dating+of+loess+in+the+Huangshui+river+valley,+northeastern+Tibetan+plateau.">Quartz OSL and K-feldspar post-IR IRSL dating of loess in the Huangshui river valley, northeastern Tibetan plateau.</a> Aeolian Research. 33, 23-32 (2018).</li><li>Murray, A., Olley, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Precision+and+accuracy+in+the+optically+stimulated+luminescence+dating+of+sedimentary+quartz:+A+status+review.">Precision and accuracy in the optically stimulated luminescence dating of sedimentary quartz: A status review.</a> Geochronometria. 21, 1-16 (2002).</li><li>Murray, A. S., Wintle, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Luminescence+dating+of+quartz+using+an+improved+single-aliquot+regenerative-dose+protocol.">Luminescence dating of quartz using an improved single-aliquot regenerative-dose protocol.</a> Radiation Measurements. 32 (1), 57-73 (2000).</li><li>Timar-Gabor, A., Wintle, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=On+natural+and+laboratory+generated+dose+response+curves+for+quartz+of+different+grain+sizes+from+Romanian+loess.">On natural and laboratory generated dose response curves for quartz of different grain sizes from Romanian loess.</a> Quintenary Geochronology. 18, 34-40 (2013).</li></ol>

The Baylor University Geoluminescence Dating Research Laboratory, within the Dept. of Geosciences and the associated personal do not have any conflicts of interest or financial interest that can affect the design of experiments or analysis, protocols, outcomes of the research or educational activities conducted in the lab. This Lab, including all the technology within and software, is used solely to conduct research, discovery, education, and mentoring.

Quartz mineralogical purity is critical for OSL dating. However, quartz spectral purity is equally important and is usually enhanced with the careful concentration of quartz grains. Ideally, quartz grains under blue LED light (470 nm &#177; 20 nm) stimulation for 40 s should emit &#8805; 90% of the luminescence within the first ~0-2.5 s of stimulation, termed the fast component, with &#60; 10% of light emission between ~2.5 and ~15 s (medium component), and a final low emission post ~15 s, (slow component) (Figure 8). A luminescence emission dominated by a fast component is preferred because it is rapidly solar reset (in seconds) and shows high sensitivity to applied &#946; radiation in the laboratory, enhancing equivalent dose determinations. An important metric to assess the dominance of fast components for OSL dating of quartz is the calculation of a &#34;fast ratio&#34;29,30 with an example shown by Equation 2 and in Figure 8. A fast ratio of &#62;20 for quartz shine down curve is considered a robust luminescence emission suitable for OSL dating29 (see Figure 8A). Separates that have contamination with K-feldspars and plagioclase or feldspathic inclusions often yield fast ratios of &#60;10 (see Figure 8B,C) and are unsuitable for SAR quartz dating protocols.
Fast Ratio <img alt="Equation 2" src="/files/ftp_upload/62706/62706eq02.jpg" /> (Equation 2)
Where L1: Fast component emission for ~0-2.5 s 
L2: Medium component emission ~2.5-15 s L3: Slow component emission ~ 15-40 s
An important test on the spectral purity of isolated quartz grains is the response of aliquots to infrared excitation from LEDs (845 nm &#177; 4 nm). Most quartz grains yield a low or negligible luminescence emission with IR stimulation at or within a few hundred counts of background emissions. A metric has been developed to assess IR-based emissions, called the IR depletion ratio, which is calculated as a SAR ratio (Lx/Tx) for irradiated (5-10 Gy) quartz grains stimulated with IR LEDs and then blue LEDs. Specifically, the ratio of IR luminescence divided by blue emissions should be &#60;5%, which indicates a spectrally pure quartz fraction amenable for OSL dating (Figure 8A). However, there are instances that mineralogically pure quartz grains can yield errant luminescence emissions with IR stimulation. This IR signal may reflect adhering lithic fragments or feldspathic inclusions in quartz. In such instances, quartz grains should be dated by feldspar protocols31. These protocols with modifications can be used to separate and confirm the purity of other minerals for OSL dating, such as k-feldspar, plagioclase, and olivine and pyroxene for other planetary applications.
The ability to isolate a &#62;99% quartz separate and confirm the purity at the grain level is a prerequisite for accurate luminescence dating. Single-grain and ultra-small aliquot (10-50 grains) dating requires additional verification that the luminescence emissions of all grains were from quartz. In turn, the application of thermal transfer approaches that can yield credible OSL ages up to one million years is predicated on pure quartz signals from mineral grains6. A mono-mineralogic quartz separate is foundational for applying OSL-SAR protocols, which provides a sequence of ages for deciphering the depositional history of eolian and fluvial systems for the late Quaternary1,2,32,33 (Figure 1 and Figure 2). Contamination of quartz aliquots by the errant K-feldspar grains or feldspathic inclusions in quartz or adhering lithic fragment yields a mixed dosimetric signal and prone to anomalous fading often yields underestimates4. However, a pure quartz separate does not absolutely ensure spectral purity and appropriate emissions for quartz dating. Effective OSL dating requires careful and complete isolation of quartz grains and OSL associated metrics to verify a pure quartz separate mineralogically and spectrally2,33,34.

Optically stimulated luminescence (OSL) geochronology yields the time from the last light or heat exposure after sediment erosion, deposition and burial; and further exposure to light or heat. Thus, natural sedimentary processes or heating events (&#62;300 &#176;C) reduces the previously inherited luminescence signal to a consistently low level. In the past two decades, there have been substantial advances in luminescence dating, such as single aliquot and grain analysis of specific mineral grains, like quartz. These experiment-based dating protocols with blue or green diodes can compensate effectively for sensitivity changes induced in the laboratory, rendering OSL ages for the past ca. 500 ka1,2,3.
Silicate minerals such as quartz and potassium feldspar have varying crystal lattice-charge defects; some formed at the time of mineral crystallization and others due to subsequent exposure to ionizing radiation, resulting in geochronometric potential. These defects are probable locations of electron storage with trap-depth energies of ~1.3-3 eV. A subpopulation of contained electrons in lattice-charge defects of quartz grains is a source for time-diagnostic luminescence emissions with excitation by blue light. Thus, this luminescence emission increases with time, above the solar or heat reset level with exposure to ionizing radiation during the burial period. This signal is reduced to a low, definable level (&#34;zeroed&#34;) with subsequent sunlight exposure with sediment erosion, transport, and deposition. This luminescence &#34;cycle&#34; occurs in most depositional environments on Earth and other planets. Thus, OSL dating of sedimentary quartz grains provides a depositional age, reflecting the time elapsed since the last light exposure with deposition and burial (Figure 1).
Luminescence dating is a dosimetric-based technique that yields age estimates for selected mineral grains, like quartz, from eolian, fluvial, lacustrine, marine, and colluvial sediments associated with enumerable contexts for geomorphic, tectonic, paleontologic, paleoclimatic, and archaeologic research2,4,5,6,7. OSL dating is also being evaluated to constrain surface processes on other planets, particularly on Mars8,9. Often, the most used mineral in OSL dating on Earth is quartz, reflecting its natural abundance, an inherent sensitivity as a geochronometer, signal stability, and rapid resetting with sunlight exposure (seconds to minutes)4,10,11,12. However, the accuracy of OSL dating is compromised if the quartz extract is impure, particularly if contaminated by potassium and other feldspars, which can have luminescence emissions ten to hundred-fold brighter than quartz and can yield age underestimates13. Therefore, the absolute (&#62;99%) purity for extracts of quartz grains from sediment is pivotal for accurate OSL dating. Thus, the focus of this contribution is to provide detailed procedures for isolating highly purified quartz grain separates from a variety of polymineral sediments. This requires integration of knowledge of mineralogy, crystal chemistry; optical and Raman imaging, to effectively apply laboratory protocols, to render OSL ages on quartz grains from carefully sampled strata from retrieved sediment cores. The sediment cores were collected by a push and percussion coring method, which retrieved intact sediment down to a depth of 20-25 m.
The OSL time-sensitive signal is reset relatively rapidly with minutes to hours of sunlight exposure. The geological OSL signal accumulates from this solar reset level. Although, the OSL emissions of quartz are considerably variable, reflecting original crystalline structure, lattice impurities, sensitization with luminescence resetting cycles14 (Figure 1). Thus, there is inherent variability in the dose sensitivity of quartz, and dating protocols need to be devised for specific mineralogic and sedimentary provenance. Fortunately, the emergence of single aliquot regenerative (SAR) dose protocols for quartz1,2 yielded systematics to redress variability in the OSL emissions and metrics to evaluate laboratory changes in apparent OSL sensitivity. Sediment grains function as long-term radiation dosimeters when concealed from further light exposure, with the luminescence signal serving as a measure of radiation exposure during the burial period. The radiation dose that is equivalent to the natural luminescence emission of isolated quartz grains is referred to as the equivalent dose (De: in grays, Gy), which is the numerator of the OSL age equation (Equation 1). The denominator is the Dose rate (Dr: Grays/yr.), defined by contributing &#945;, &#946;, and &#947; radiation, originating from the radioactive decay of daughter isotopes in the 235U, 238U, 232Th decay series, 40K, and with lesser contributions from the decay of 85Rb and cosmic and galactic sources.
OSL age (yr) = <img alt="Equation 1" src="/files/ftp_upload/62706/62706eq01.jpg" /> (Equation 1)
Where, D&#945;&#160;= alpha dose D&#946; = beta dose D&#947; = gamma dose Dc = cosmic dose and w=water attenuation factor.
Another method for U and Th determinations in the laboratory or the field is gamma spectrometry, with the Germanium variant able to quantify U and Th isotopic disequilibrium with suitable adjustments to the dose rate. The beta and gamma components of the environmental dose rate need to be modified for mass attenuation15. However, there is an effectively insignificant alpha dose for grains &#62;50 &#181;m with the outer 10-20 &#181;m of grains removed by treatment with undiluted HF during preparation. A critical component in dose rate assessment is the quantification of the cosmic and galactic dose during the burial period, which is calculated for specific points on Earth with adjustments for longitude, latitude, elevation, burial depth, and density of overlying sediment16,17.
Sediments that contain &#62;15% quartz are usually relatively straightforward for separating out a high purity quartz fraction. However, sediments with &#60;15% quartz often require added time to ensure needed mineralogic purity for OSL dating. Approximately 500-1000 quartz grains are needed for this analysis, but often thousands of grains are separated for duplicate analyses, archiving to expand a calibration library, and future advancements. The mineralogic composition of sediment samples is initially assessed, grain by grain, by petrographic analysis through a binocular microscopic (10-20x) and associated imagine analysis. The mineralogy of individual grains is tested further by Raman spectroscopy to measure grain spectra using an excitation laser (455 nm, 532 nm, 633 nm, or 785 nm) and statistically compare grain emissions to known mineral spectra from the RRUFF System Database18.
Once the visual and spectral inspection is satisfactory, the purity of the OSL signal is further checked, utilizing an automated luminescence reader system. Three to five aliquots of the sample are exposed to infrared excitation (IR = 1.08 watts at 845 nm &#177; 4 nm), which preferentially stimulates feldspar minerals, and this emission is compared to emissions by blue light excitation (Bl = 470 nm &#177; 20 nm), which preferentially stimulates quartz. If the ratio IR/Bl &#8805; 5%, the test indicates feldspar contamination and acid digestions are repeated. If the ratio IR/Bl &#60;5%, then the samples are deemed quartz fraction satisfactorily for dating.
Single aliquot regeneration (SAR) protocols on quartz grains is an often-used approach in OSL dating sediments with procedures tailored for a specific sample, a study site, or an area. The reproducibility of these protocols is determined by giving quartz grains a known beta dose (e.g., 30 Gy) and evaluating what heat pretreatment recovers this known dose (Figure 2). In practice, determining a De with the SAR protocols involves the calculation of a ratio between the natural luminescence and the luminescence from a known test dose (Ln/Tn ratio), which is compared to the luminescence emissions for regenerative doses divided by the luminescence from the same test dose (Lx/Tx) (Figure 2). A correction, a consistently applied test-dose (e.g., 5 Gy), has been devised to compensate for quartz grain(s) sensitivity changes with measurement through SAR cycles. Often the OSL emissions increase by &#62;5% with each successive SAR cycle, though given the same dose (e.g., 5 Gy)7.
At least forty aliquots of quartz or 500 grains are analyzed with TL/OSL reader system, with blue light excitation. The luminescence data generated is analyzed by software associated with the Ris&#248; TL/OSL-DA-20 reader system. The De and Dr values and age estimates are calculated using the Luminescence Dose and Age Calculator (LDAC)17. This platform applies statistical models to determine equivalent dose (De) values and render corresponding OSL age with constrained errors.
The extracted light-shielded sample from a core is prepared for two reasons: 1) To obtain a mineralogic fraction of quartz grains with a purity of &#62;99%, and 2) To isolate grains of specific size fraction, e.g., 150-250 &#181;m, for assessment of the environmental Dr for OSL dating17. In many sedimentary settings, quartz grains are common; but mixed with other silicate and non-silicate minerals, rock fragments, and organic matter. Previously, procedures were briefly outlined, indicating some specific steps and reagents needed to isolate pure quartz grains in the context of OSL dating 13,19,20,21,22,23. This contribution has benefited greatly from these previous approaches. This paper outlines revised, and more detailed protocols using petrographic imaging and Raman technology to monitor grain mineralogy and render highly pure (&#62;99%) quartz extracts for luminescence dating. These quartz isolation protocols have been developed after preparing hundreds of samples from diverse geological environments in the Americas, Eurasia, China, and Africain, the Baylor Geoluminescence Dating Research Laboratory, reflecting analytical experience over thirty years, and are not definitive methods, with suitable variations used by other labs. These are not static protocols, and modifications and additions for improvement are welcomed.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>10 mL pipette</td>
			<td>VWR</td>
			<td>53044-139</td>
			<td></td>
		</tr>
		<tr>
			<td>100 mL graduate cylinder</td>
			<td>VWR</td>
			<td>24774-692</td>
			<td></td>
		</tr>
		<tr>
			<td>100% China bristles brush</td>
			<td>Subang</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>2&#39; Macro MC7 PVC Liner</td>
			<td>Macro-Core</td>
			<td>46125</td>
			<td></td>
		</tr>
		<tr>
			<td>Analytical balance</td>
			<td>Sartorius 1207 MP2</td>
			<td>2107</td>
			<td></td>
		</tr>
		<tr>
			<td>Bransonic Ultrasonic cleaner</td>
			<td>VWR</td>
			<td>97043-958</td>
			<td></td>
		</tr>
		<tr>
			<td>Calgonate Hydrofluoric Acid Burn Relief Gel, Calgonate</td>
			<td>VWR CALGEL25</td>
			<td>101320-858</td>
			<td></td>
		</tr>
		<tr>
			<td>Concentrated (48&#8211;51%) hydrofluoric acid (HF)</td>
			<td>VWR</td>
			<td>BDH3042</td>
			<td></td>
		</tr>
		<tr>
			<td>Core MC7 Soil Sampling System</td>
			<td>Macro-Core</td>
			<td>216883</td>
			<td></td>
		</tr>
		<tr>
			<td>Deionized water (DIW)</td>
			<td>Baylor University</td>
			<td>DIW Faucet</td>
		</tr>
		<tr>
			<td>Geoprobe</td>
			<td>Enviroprobe</td>
			<td>6620DT</td>
			<td></td>
		</tr>
		<tr>
			<td>Hydrochloric acid 36.5&#8211;38.0% ACS, VWR Chemicals BDH</td>
			<td>VWR</td>
			<td>BDH3032-3.8LP</td>
			<td></td>
		</tr>
		<tr>
			<td>Hydrogen peroxide&#160; (H2O2) 25%</td>
			<td>VWR Chemicals BDH</td>
			<td>BDH7814-3</td>
			<td></td>
		</tr>
		<tr>
			<td>Hydrogen peroxide 12%</td>
			<td>VWR Chemicals BDH</td>
			<td>BDH7814-3</td>
			<td></td>
		</tr>
		<tr>
			<td>Inductively coupled plasma mass spectrometry-ICP-MS</td>
			<td>ALS Laboratories, Reno, NV</td>
			<td>ME-MS81d</td>
			<td></td>
		</tr>
		<tr>
			<td>Laser diffraction particle size analyzer Malvern Mastersizer 3000</td>
			<td>Malvern Panalytical</td>
			<td>Mastersizer 3000</td>
			<td></td>
		</tr>
		<tr>
			<td>Lead hydrometer with range 2.00&#8211;3.00 g/cm3</td>
			<td>Thomas Scientific</td>
			<td>13K065</td>
			<td></td>
		</tr>
		<tr>
			<td>LOW PRESSURE SODIUM 35W CLEAR Sodium Vapor Lamp for Thomas Duplex Safelights</td>
			<td>Interlighht</td>
			<td>WW-5EGX-9</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic rods and wands</td>
			<td>Alnico V Magnet</td>
			<td>Magnetic wands #21R584. Magnetic Stir Bar #21R590</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic Stirrer Stainless Steel Magnetic Mixer with stir bar. Max Stirring Capacity 3000 ml</td>
			<td>INTLLAB</td>
			<td>MS-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic Stirrer Stainless Steel Magnetic Mixer with stir bar. Max Stirring Capacity 3000 mL</td>
			<td>INTLLAB</td>
			<td>MS-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic Stirrer Stainless Steel Magnetic Mixer with stir bar. Max Stirring Capacity 3000 mL</td>
			<td>INTLLAB</td>
			<td>MS-500</td>
			<td></td>
		</tr>
		<tr>
			<td>MC5 PVC Liner</td>
			<td>Macro-Core</td>
			<td>600993</td>
			<td></td>
		</tr>
		<tr>
			<td>MC5 Soil Sampling System (LWCR)</td>
			<td>Macro-Core</td>
			<td>204218</td>
			<td></td>
		</tr>
		<tr>
			<td>Neodymium magnets</td>
			<td>MIKEDE</td>
			<td>24100000</td>
			<td></td>
		</tr>
		<tr>
			<td>Nylon mesh</td>
			<td>Gilson Company, INC</td>
			<td>500 &#956;= NM-B #35&#160; 450 &#956;= NM-1 #40-10 350 &#956;= NM-B #45 250 &#956;= NM-B #60 150 &#956;= NM-2 #100-10 100 &#956;= NM-C #140&#160; 63 &#956;= NM-C #230 45 &#956;= NM-3 #325-10&#160; 38 &#956;= NM-D #400</td>
			<td></td>
		</tr>
		<tr>
			<td>Optifix Dispensers, MilliporeSigma HCl bottle dispenser</td>
			<td>VWR</td>
			<td>EM-10108048-1. Serial F93279E</td>
			<td></td>
		</tr>
		<tr>
			<td>Optifix Dispensers, MilliporeSigma HF bottle dispenser</td>
			<td>VWR</td>
			<td>EM-10108048-1. Serial 005499</td>
			<td></td>
		</tr>
		<tr>
			<td>Plastic beaker</td>
			<td>VWR</td>
			<td>89172</td>
			<td></td>
		</tr>
		<tr>
			<td>Powdered POLY-GEE Brand Sodium Polytungstate (SPT-Na6 (H2W12O40) _H2O)</td>
			<td>Geoliquids, INC.</td>
			<td>SPT001</td>
			<td></td>
		</tr>
		<tr>
			<td>Premier binocular microscope</td>
			<td>VWR</td>
			<td>SMZ-05/Stereo Zoom Microscope/EA</td>
			<td></td>
		</tr>
		<tr>
			<td>Quartz Griffin Beakers, Chemglass</td>
			<td>VWR</td>
			<td>89028</td>
			<td></td>
		</tr>
		<tr>
			<td>REDISHIP Protector Premier Hood</td>
			<td>VWR</td>
			<td>&#160; 89260-056</td>
			<td></td>
		</tr>
		<tr>
			<td>RIS&#248; TL/OSL DA-20</td>
			<td>Ris&#248; National Laboratory, Denmar</td>
			<td>TL/OS-DA-2</td>
			<td></td>
		</tr>
		<tr>
			<td>Rockwell F80 Sonicrafter electric saw</td>
			<td>Rockwell</td>
			<td>RK5121K</td>
			<td></td>
		</tr>
		<tr>
			<td>Spectroscopy analyzer: DXR Raman microscope</td>
			<td>Thermoscientific DXR Raman microscope</td>
			<td>IQLAADGABFFAHCMBDI</td>
			<td></td>
		</tr>
		<tr>
			<td>Squirt bottle</td>
			<td>VWR</td>
			<td>10111</td>
			<td></td>
		</tr>
		<tr>
			<td>Tetrasodium diphosphate decahydrate 99.0&#8211;103.0%, crystals, BAKER ANALYZED ACS, J.T. Baker (Na4P2O7 10H2O) &#62; 95%,</td>
			<td>VWR</td>
			<td>JT3850-1</td>
			<td></td>
		</tr>
		<tr>
			<td>Thomas Duplex Super SafeLight Sodium Photographic Darkroom Light USA</td>
			<td>Freestyle</td>
			<td>Model: 42122</td>
			<td></td>
		</tr>
	</tbody>
</table>

isolation of quartz grains for optically stimulated luminescence (osl) dating of quaternary sediments for paleoenvironmental research

Optically stimulated luminescence (OSL) dating quantifies the time since mineral grains were deposited and shielded from additional light or heat exposure, which effectively resets the luminescence clock. The systematics of OSL dating is based on the dosimetric properties of common minerals, like quartz and feldspar. The acquired luminescence with exposure to natural ionizing radiation after burial provides a depositional age for many Quaternary sedimentary systems, spanning the past 0.5 Ma. This contribution details the procedures for separating pure quartz grains of a known range of particle sizes to facilitate luminescence analysis with small or single grain aliquots. Specifically, protocols are given for the needed data and interpretations for effective OSL dating of terrestrial sediment cores or sample tubes from exposures. These cores, 5-20 m long in 1.2 m sections, are split lengthwise and crown-cut leaving 80% of core volume undisturbed, which facilitates sampling of light-protected sediment for OSL dating deep within the core. Sediment samples are then subjected to a series of physical separations to obtain a certain grain-size interval (e.g., 150-250 &#181;m). Magnetic minerals are removed in wet and dry states using magnets. A series of chemical digestions starts with soaking in H2O2 to remove organic matter, followed by HCl exposure to remove carbonate minerals, followed by density separation. Subsequently, grains are soaked in HF for 80 min and after in HCl to render solely quartz grains. The mineralogic purity (&#62;99%) of the quartz extract is quantified with grain petrographic assessment and Raman spectroscopy. Repeating this quartz isolation procedure may be necessary with sediment that contains &#60;15% quartz grains. Excitation of the purified quartz grains by LED-derived blue and IR light allows calculations of the fast and IR depletion ratios, which are metrics to assess the dominance of luminescence emissions from quartz.

The laboratory procedures outlined are focused on enhancing the separation of pure quartz grains (700 to 50 &#181;m size) needed for OSL dating without inadvertent light resetting in the laboratory (Figure 1). A pure quartz separate, mineralogically and optically, is a prerequisite for applying SAR and TT-OSL dating procedures (Figure 2). These procedures explain the necessary steps for effectively understanding and sampling continuous sediment cores, avoiding zones of pedogenesis and diagenesis, retrieving unlight-exposed sediments from cores (Figure 3 and Figure 4); to isolate quartz grains for OSL dating protocols to constrain the timing of sediment deposition in the past ca. 500 ka (Figure 5). The mineralogy of grains of the unprepared sample and prepared separates are assessed continuously through the preparation process to identify the contaminating mineralogy and actively assess the process of removal of unwanted minerals (Figure 6 and Figure 7). The quartz mineralogic purity is determined for subset grains (100-400) through binocular microscopic inspection (10-20x) and by Raman spectroscopy. The use of this technology and prerequisite knowledge is vital to assess and confirm the needed purity (&#62;99%) of quartz separations for OSL dating (Figure 8).
The process for quartz separation is started with the removal of organic matter with H2O2 and then the subsequent purging of Ca/MgCO3 with soaking in HCl. Subsequently, a size fraction is designated by sieving with disposable nylon mesh (e.g., 150 and 250 &#181;m), which is necessary for calculating dose rate values (in mGy/y) (Figure 6A inset). The purity of the quartz separate is enhanced by two density separations at 2.6 and 2.7 g/cc, the bounding density of quartz (Figure 6B inset). The subsequent soaking of sized grains in HF for 80 min removes non-quartz minerals. This treatment also etches the outer 10-20 &#181;m of grains to remove the alpha-dose affected area, simplifying dose rate calculations (Figure 6). The purity of the quartz separate is never assumed but assessed through binocular microscopic inspection and Raman-based measurements at the end of grain separation. Density separations and/or HF treatment can be repeated to rid the separate contaminating grains if a representative aliquot contains &#62;1% non-quartz grains, particularly feldspar minerals (Figure 7). The quartz purification procedure was repeated up to four times with quartz contents of &#60;15% to render shine down curves with a fast ratio of &#62;20, characteristic of pure quartz (Figure 8).
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/62706/62706fig01.jpg" /> 
Figure 1: Processes with OSL dating. (A) Mineral grains acquire OSL with ionizing radiation exposure. (B) Grain OSL is reset by sunlight with erosion/ transport. (C) Exposure to ionizing with burial; luminescence acquired. (D) Light exposure resets OSL with erosion/ transport. (E) Grains are re-buried, and OSL is acquired with exposure to ionizing radiation. (F) Shows sampling without light exposure. The resultant measured natural OSL is followed by a normalizing test dose (Ln/Tn) which is equated to the regenerative dose curve to yield an equivalent dose (De). This figure has been modified from Forman, S. L. et al.7. <a href="https://www.jove.com/files/ftp_upload/62706/62706fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/62706/62706fig02.jpg" /> 
Figure 2: Optical Stimulating Luminescence- Single Aliquot Regeneration (OSL-SAR) protocols for quartz grains. (A) Equivalent dose using SAR protocols; the natural OSL is Ln/Tn, and the regenerative dose is Lx/Tx; sensitivity changes are corrected by giving a test dose (e.g., 5 Gy). (B) Generalized SAR protocol. This figure has been modified from Forman, S. L. et al.7. <a href="https://www.jove.com/files/ftp_upload/62706/62706fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/62706/62706fig03.jpg" /> 
Figure 3: Flow diagram outlining the steps necessary to open, describe, and interpret a recovered sediment core. This figure shows retrieval of sediment core using percussion corer, followed by the opening, cleaning, description, and study of the core to obtain the optimal sample for OSL dating. <a href="https://www.jove.com/files/ftp_upload/62706/62706fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/62706/62706fig04.jpg" /> 
Figure 4: Example of a typical log of a core sedimentary and stratigraphic section. Units and pedosedimentary facies are defined using sedimentology, stratigraphy, pedology, granulometry, and carbonate percentage. The soils horizons found in the stratigraphic column from top to bottom are: A: Surface organic-rich horizon, B: subsoil with weak structure and color (Bw), and buried B horizon Btb with clay accumulation, Btkb with secondary calcium carbonate and clay accumulation, and Bkb with an accumulation of secondary calcium carbonate. The dominant particle size of sedimentary units is shown on the lower horizontal with medium sand (MS), fine sand (FS), very fine sand (VFS), and Silt (Si). <a href="https://www.jove.com/files/ftp_upload/62706/62706fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/62706/62706fig5v2.jpg" /> 
Figure 5: Flow diagram for the steps necessary to collect an OSL sample from a sediment core. This figure presents a flow diagram with the main steps followed to prepare a quartz separate for OSL dating. The protocols start with the extraction of a polymineral sediment from light-shielded areas of the core in the light safe OLS lab. They continue with the extraction of the mono-mineralogic fraction of quartz, comprising the removal of organic matter with peroxide, carbonates with HCl, and magnetic minerals using hand magnets. The separation of the specific fraction of sand-size sediment is done by sieving; separation of minerals less dense and heavier than quartz is done using density liquids (&#961; = 2.6 g/cc and 2.7 g/cc). The final steps of cleaning require immersion of the sediment into HF and HCl full strength to isolate quartz from any other mineral in the fraction. The purity of the separate is evaluated by binocular inspection, RAMAN spectroscopy, and further verification of IRSL (Infrared) emissions. The goal is to obtain a sample with a purity &#8805;99%. Failure to do so requires that some of the steps must be repeated.<a href="https://www.jove.com/files/ftp_upload/62706/62706fig5v2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/62706/62706fig06.jpg" /> 
Figure 6: Flow chart depicting all the steps necessary to obtain a pristine quartz separate from a sediment sample from a core. This clean quartz fraction will be used for OSL-SAR analyses for age assessment. <a href="https://www.jove.com/files/ftp_upload/62706/62706fig06large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/62706/62706fig07.jpg" /> 
Figure 7:&#160;Comparison of two samples collected in two different areas: White Sands (first row) and Mongolia (second row). Column A shows raw samples under the binocular microscope, as collected in the field. Column B shows the separate fractions for each processed samples, under the binocular microscope. Column C shows the corresponding RAMAN spectroscopy results. The sample from White Sands contains sulfates (mainly gypsum), halides, and very little quartz (column A). Correspondently, the separate fraction (63-100 &#181;m) for the processed sample in a Column B&#160;shows that it contains mostly quartz, still with some vestiges of gypsum, as shown by the RAMAN Spectroscopy in Column C. The ratio between the OSL IR and blue responses for this sample is 9%, confirming that it needs a second separation in density at 2.6 g/cc, which possibly will remove the lighter gypsum (2.36 g/cc) from heavier quartz. Contrastingly, the Mongolian sample (Column A) is initially very rich in feldspars, predominantly K-feldspar. After undergoing the cleaning procedures, shows abundant quartz isolated in the 100-150 &#181;m separate (Columns B and C), rendering a satisfactory IR/Bl ratio of 3.7%.&#160;<a href="https://www.jove.com/files/ftp_upload/62706/62706fig07large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 8" class="xfigimg" src="/files/ftp_upload/62706/62706fig08.jpg" /> 
Figure 8: Comparison of fast ratio for the natural in three samples that represent different degrees of quartz fraction purity. (A) The ideal fast ratio distribution in a pristine eolian sample from Red River, with fast ratio= 72. Contrasting figures (Figure 8B,C) have a less fast component with blue LED stimulation, which is below 20. (B) A sample with incomplete quartz and plagioclases separation. The L2 and L3 components are a significant % of the L1 component (see Equation 2). (C) A shine-down curve for feldspathic quartz, with a dominant medium component (L2). <a href="https://www.jove.com/files/ftp_upload/62706/62706fig08large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Isolation of Quartz Grains for Optically Stimulated Luminescence OSL Dating of Quaternary Sediments for Paleoenvironmental Research at JoVE.com

Isolation of Quartz Grains for Optically Stimulated Luminescence OSL Dating of Quaternary Sediments for Paleoenvironmental Research

This protocol is for the isolation of quartz grains by size for luminescence dating of sediments. Outlined are physical cleansing and chemical digestions by soaking sequentially in H2O2, HCl, HF, and HCl again to isolate quartz grains. The quartz purity is quantified with microscopic assessment, Raman spectroscopy, and IR depletion ratio.

Isolation of Quartz Grains for Optically Stimulated Luminescence (OSL) Dating of Quaternary Sediments for Paleoenvironmental Research

Optically stimulated luminescence (OSL) dating quantifies the time since mineral grains were deposited and shielded from additional light or heat ...

isolation-quartz-grains-for-optically-stimulated-luminescence-osl

Research

JoVE Journal

Environment

2.6K Views.  Baylor University. This protocol is for the isolation of quartz grains by size for luminescence dating of sediments. Outlined are physical cleansing and chemical digestions by soaking sequentially in H2O2, HCl, HF, and HCl again to isolate quartz grains. The quartz purity is quantified with microscopic assessment, Raman spectroscopy, and IR depletion ratio.

Video: Isolation of Quartz Grains for Optically Stimulated Luminescence OSL Dating of Quaternary Sediments for Paleoenvironmental Research

Determination of Microbial Extracellular Enzyme Activity in Waters, Soils, and Sediments using High Throughput Microplate Assays

Much of the nutrient cycling and carbon processing in natural environments occurs through the activity of extracellular enzymes released by microorganisms. Thus, measurement of the activity of these extracellular enzymes can give insights into the rates of ecosystem level processes, such as organic matter decomposition or nitrogen and phosphorus mineralization. Assays of extracellular enzyme activity in environmental samples typically involve exposing the samples to artificial colorimetric or fluorometric substrates and tracking the rate of substrate hydrolysis. Here we describe microplate based methods for these procedures that allow the analysis of large numbers of samples within a short time frame. Samples are allowed to react with artificial substrates within 96-well microplates or deep well microplate blocks, and enzyme activity is subsequently determined by absorption or fluorescence of the resulting end product using a typical microplate reader or fluorometer. Such high throughput procedures not only facilitate comparisons between spatially separate sites or ecosystems, but also substantially reduce the cost of such assays by reducing overall reagent volumes needed per sample.

Microplate based procedures are described for the colorimetric or fluorometric analysis of extracellular enzyme activity. These procedures allow for the rapid assay of such activity in large numbers of environmental samples within a manageable time frame.

Much of the nutrient cycling and carbon  processing in natural environments occurs through the activity of extracellular  enzymes released by ...

Laboratory-determined Phosphorus Flux from Lake Sediments as a Measure of Internal Phosphorus Loading

Eutrophication is a water quality issue in lakes worldwide, and there is a critical need to identify and control nutrient sources. Internal phosphorus (P) loading from lake sediments can account for a substantial portion of the total P load in eutrophic, and some mesotrophic, lakes. Laboratory determination of P release rates from sediment cores is one approach for determining the role of internal P loading and guiding management decisions. Two principal alternatives to experimental determination of sediment P release exist for estimating internal load: in situ measurements of changes in hypolimnetic P over time and P mass balance. The experimental approach using laboratory-based sediment incubations to quantify internal P load is a direct method, making it a valuable tool for lake management and restoration.
Laboratory incubations of sediment cores can help determine the relative importance of internal vs. external P loads, as well as be used to answer a variety of lake management and research questions. We illustrate the use of sediment core incubations to assess the effectiveness of an aluminum sulfate (alum) treatment for reducing sediment P release. Other research questions that can be investigated using this approach include the effects of sediment resuspension and bioturbation on P release.
The approach also has limitations. Assumptions must be made with respect to: extrapolating results from sediment cores to the entire lake; deciding over what time periods to measure nutrient release; and addressing possible core tube artifacts. A comprehensive dissolved oxygen monitoring strategy to assess temporal and spatial redox status in the lake provides greater confidence in annual P loads estimated from sediment core incubations.

Lake eutrophication is a water quality issue worldwide, making the need to identify and control nutrient sources critical. Laboratory determination of phosphorus release rates from sediment cores is a valuable approach for determining the role of internal phosphorus loading and guiding management decisions.

Eutrophication is a water quality issue in lakes worldwide, and there is a critical need to identify and control nutrient sources. Internal phosphorus (P) ...

A Whole Cell Bioreporter Approach to Assess Transport and Bioavailability of Organic Contaminants in Water Unsaturated Systems

Bioavailability of contaminants is a prerequisite for their effective biodegradation in soil. The average bulk concentration of a contaminant, however, is not an appropriate measure for its availability; bioavailability rather depends on the dynamic interplay of potential mass transfer (flux) of a compound to a microbial cell and the capacity of the latter to degrade the compound. In water-unsaturated parts of the soil, mycelia have been shown to overcome bioavailability limitations by actively transporting and mobilizing organic compounds over the range of centimeters. Whereas the extent of mycelia-based transport can be quantified easily by chemical means, verification of the contaminant-bioavailability to bacterial cells requires a biological method. Addressing this constraint, we chose the PAH fluorene (FLU) as a model compound and developed a water unsaturated model microcosm linking a spatially separated FLU point source and the FLU degrading bioreporter bacterium Burkholderia sartisoli RP037-mChe by a mycelial network of Pythium ultimum. Since the bioreporter expresses eGFP in response of the PAH flux to the cell, bacterial FLU exposure and degradation could be monitored directly in the microcosms via confocal laser scanning microscopy (CLSM). CLSM and image analyses revealed a significant increase of the eGFP expression in the presence of P. ultimum compared to controls without mycelia or FLU thus indicating FLU bioavailability to bacteria after mycelia-mediated transport. CLSM results were supported by chemical analyses in identical microcosms. The developed microcosm proved suitable to investigate contaminant bioavailability and to concomitantly visualize the involved bacteria-mycelial interactions.

A whole cell bioreporter assay with Burkholderia sartisoli RP037-mChe was developed to detect fractions of an organic contaminant (i.e., fluorene) available for bacterial degradation after active transport by mycelia bridging air-filled pores in a water unsaturated model system.

Bioavailability of contaminants is a prerequisite for their effective biodegradation in soil. The average bulk concentration of a contaminant, however, is ...

Development of Sulfidogenic Sludge from Marine Sediments and Trichloroethylene Reduction in an Upflow Anaerobic Sludge Blanket Reactor

The importance of microbial sulfate reduction relies on the various applications that it offers in environmental biotechnology. Engineered sulfate reduction is used in industrial wastewater treatment to remove large concentrations of sulfate along with the chemical oxygen demand (COD) and heavy metals. The most common approach to the process is with anaerobic bioreactors in which sulfidogenic sludge is obtained through adaptation of predominantly methanogenic granular sludge to sulfidogenesis. This process may take a long time and does not always eliminate the competition for substrate due to the presence of methanogens in the sludge. In this work, we propose a novel approach to obtain sulfidogenic sludge in which hydrothermal vents sediments are the original source of microorganisms. The microbial community developed in the presence of sulfate and volatile fatty acids is wide enough to sustain sulfate reduction over a long period of time without exhibiting inhibition due to sulfide. 
This protocol describes the procedure to generate the sludge from the sediments in an upflow anaerobic sludge blanket (UASB) type of reactor. Furthermore, the protocol presents the procedure to demonstrate the capability of the sludge to remove by reductive dechlorination a model of a highly toxic organic pollutant such as trichloroethylene (TCE). The protocol is divided in three stages: (1) the formation of the sludge and the determination of its sulfate reducing activity in the UASB, (2) the experiment to remove the TCE by the sludge, and (3) the identification of microorganisms in the sludge after the TCE reduction. Although in this case the sediments were taken from a site located in Mexico, the generation of a sulfidogenic sludge by using this procedure may work if a different source of sediments is taken since marine sediments are a natural pool of microorganisms that may be enriched in sulfate reducing bacteria.

Microbial sulfate reduction is a process of great importance in environmental biotechnology. The success of the sulfidogenic reactors depends among other factors on the microbial composition of the sludge. Here, we present a protocol to develop sulfidogenic sludge from hydrothermal vents sediments in a UASB reactor for reductive dechlorination purposes.

The importance of microbial sulfate reduction relies on the various applications that it offers in environmental biotechnology. Engineered sulfate ...

Administering and Detecting Protein Marks on Arthropods for Dispersal Research

Monitoring arthropod movement is often required to better understand associated population dynamics, dispersal patterns, host plant preferences, and other ecological interactions. Arthropods are usually tracked in nature by tagging them with a unique mark and then re-collecting them over time and space to determine their dispersal capabilities. In addition to actual physical tags, such as colored dust or paint, various types of proteins have proven very effective for marking arthropods for ecological research. Proteins can be administered internally and/or externally. The proteins can then be detected on recaptured arthropods with a protein-specific enzyme-linked immunosorbent assay (ELISA). Here we describe protocols for externally and internally tagging arthropods with protein. Two simple experimental examples are demonstrated: (1) an internal protein mark introduced to an insect by providing a protein-enriched diet and (2) an external protein mark topically applied to an insect using a medical nebulizer. We then relate a step-by-step guide of the sandwich and indirect ELISA methods used to detect protein marks on the insects. In this demonstration, various aspects of the acquisition and detection of protein markers on arthropods for mark-release-recapture, mark-capture, and self-mark-capture types of research are discussed, along with the various ways that the immunomarking procedure has been adapted to suit a wide variety of research objectives.

Proteins are often used to mark arthropods for dispersal research. Methods for administering internal and external protein marks on arthropods are demonstrated. Protein mark detection techniques, either through indirect or sandwich enzyme-linked immunosorbent assays (ELISAs), are also illustrated.

Monitoring arthropod movement is often required to better understand associated population dynamics, dispersal patterns, host plant preferences, and other ...

Simultaneous DNA-RNA Extraction from Coastal Sediments and Quantification of 16S rRNA Genes and Transcripts by Real-time PCR

Real Time Polymerase Chain Reaction also known as quantitative PCR (q-PCR) is a widely used tool in microbial ecology to quantify gene abundances of taxonomic and functional groups in environmental samples. Used in combination with a reverse transcriptase reaction (RT-q-PCR), it can also be employed to quantify gene transcripts. q-PCR makes use of highly sensitive fluorescent detection chemistries that allow quantification of PCR amplicons during the exponential phase of the reaction. Therefore, the biases associated with &#39;end-point&#39; PCR detected in the plateau phase of the PCR reaction are avoided. A protocol to quantify bacterial 16S rRNA genes and transcripts from coastal sediments via real-time PCR is provided. First, a method for the co-extraction of DNA and RNA from coastal sediments, including the additional steps required for the preparation of DNA-free RNA, is outlined. Second, a step-by-step guide for the quantification of 16S rRNA genes and transcripts from the extracted nucleic acids via q-PCR and RT-q-PCR is outlined. This includes details for the construction of DNA and RNA standard curves. Key considerations for the use of RT-q-PCR assays in microbial ecology are included.

A protocol to quantify bacterial 16S rRNA genes and transcripts from coastal sediments via real-time PCR is provided. The methodology includes the co-extraction of DNA and RNA; preparation of DNA-free RNA; and 16S rRNA gene and transcript quantification via RT-q-PCR, including standard curve construction.

Real Time Polymerase Chain Reaction also known as quantitative PCR (q-PCR) is a widely used tool in microbial ecology to quantify gene abundances of ...

Experimental Protocol for Biodiesel Production with Isolation of Alkenones as Coproducts from Commercial Isochrysis Algal Biomass

The need to replace petroleum fuels with alternatives from renewable and more environmentally sustainable sources is of growing importance. Biomass-derived biofuels have gained considerable attention in this regard, however first generation biofuels from edible crops like corn ethanol or soybean biodiesel have generally fallen out of favor. There is thus great interest in the development of methods for the production of liquid fuels from domestic and superior non-edible sources. Here we describe a detailed procedure for the production of a purified biodiesel from the marine microalgae Isochrysis. Additionally, a unique suite of lipids known as polyunsaturated long-chain alkenones are isolated in parallel as potentially valuable coproducts to offset the cost of biodiesel production. Multi-kilogram quantities of Isochrysis are purchased from two commercial sources, one as a wet paste (80% water) that is first dried prior to processing, and the other a dry milled powder (95% dry). Lipids are extracted with hexanes in a Soxhlet apparatus to produce an algal oil (&#34;hexane algal oil&#34;) containing both traditional fats (i.e., triglycerides, 46-60% w/w) and alkenones (16-25% w/w). Saponification of the triglycerides in the algal oil allows for separation of the resulting free fatty acids (FFAs) from alkenone-containing neutral lipids. FFAs are then converted to biodiesel (i.e., fatty acid methyl esters, FAMEs) by acid-catalyzed esterification while alkenones are isolated and purified from the neutral lipids by crystallization. We demonstrate that biodiesel from both commercial Isochrysis biomasses have similar but not identical FAME profiles, characterized by elevated polyunsaturated fatty acid contents (approximately 40% w/w). Yields of biodiesel were consistently higher when starting from the Isochrysis wet paste (12% w/w vs. 7% w/w), which can be traced to lower amounts of hexane algal oil obtained from the powdered Isochrysis product.

Experimental Protocol for Biodiesel Production with Isolation of Alkenones as Coproducts from Commercial Isochrysis Algal Biomass

Detailed methods are presented for the production of biodiesel along with the co-isolation of alkenones as valuable coproducts from commercial Isochrysis microalgae.

The need to replace petroleum fuels with alternatives from renewable and more environmentally sustainable sources is of growing importance. ...

A Flow-through Exposure System for Evaluating Suspended Sediments Effects on Aquatic Life

This paper describes the Fish Larvae and Egg Exposure System (FLEES). The flow-through exposure system is used to investigate the effects of suspended sediment on various aquatic species and life stages in the laboratory by using pumps and automating delivery of sediment and water to simulate suspension of sediment. FLEES data are used to develop exposure-response curves between the effects on aquatic organisms and suspended sediment concentrations at the desired exposure duration. The effects data are used to evaluate management practices used to reduce the interactions between aquatic organisms and anthropogenic causes of suspended sediments. The FLEES is capable of generating total suspended solids (TSS) concentrations as low as 30 to as high as 800 mg/L, making this system an ideal choice for evaluating the effects of TSS resulting from many activities including simulating low ambient levels of TSS to evaluating sources of suspended sediments from dredging operations, vessel traffic, freshets, and storms.

A robust and flexible flow-through exposure system designed to maintain sediment in suspension is presented. The system is used to investigate the effects of suspended sediment on various aquatic species and life stages in the laboratory.

This paper describes the Fish Larvae and Egg Exposure System (FLEES). The flow-through exposure system is used to investigate the effects of suspended ...

Extraction of Organochlorine Pesticides from Plastic Pellets and Plastic Type Analysis

Plastic resin pellets, categorized as microplastics (&#8804;5 mm in diameter), are small granules that can be unintentionally released to the environment during manufacturing and transport. Because of their environmental persistence, they are widely distributed in the oceans and on beaches all over the world. They can act as a vector of potentially toxic organic compounds (e.g., polychlorinated biphenyls) and might consequently negatively affect marine organisms. Their possible impacts along the food chain are not yet well understood. In order to assess the hazards associated with the occurrence of plastic pellets in the marine environment, it is necessary to develop methodologies that allow for rapid determination of associated organic contaminant levels. The present protocol describes the different steps required for sampling resin pellets, analyzing adsorbed organochlorine pesticides (OCPs) and identifying the plastic type. The focus is on the extraction of OCPs from plastic pellets by means of a pressurized fluid extractor (PFE) and on the polymer chemical analysis applying Fourier Transform-InfraRed (FT-IR) spectroscopy. The developed methodology focuses on 11 OCPs and related compounds, including dichlorodiphenyltrichloroethane (DDT) and its two main metabolites, lindane and two production isomers, as well as the two biologically active isomers of technical endosulfan. This protocol constitutes a simple and rapid alternative to existing methodology for evaluating the concentration of organic contaminants adsorbed on plastic pieces.

Microplastics act as vector of potentially toxic organic contaminants with unpredictable effects. This protocol describes an alternative methodology for assessing the levels of organochlorine pesticides adsorbed on plastic pellets and identifying the polymer chemical structure. The focus is on pressurized fluid extraction and attenuated total reflectance Fourier transform infrared spectroscopy.

Plastic resin pellets, categorized as microplastics (&#8804;5 mm in diameter), are small granules that can be unintentionally released to the environment ...

Preparation of Authigenic Pyrite from Methane-bearing Sediments for In Situ Sulfur Isotope Analysis Using SIMS

Different sulfur isotope compositions of authigenic pyrite typically result from the sulfate-driven anaerobic oxidation of methane (SO4-AOM) and organiclastic sulfate reduction (OSR) in marine sediments. However, unravelling the complex pyritization sequence is a challenge because of the coexistence of different sequentially formed pyrite phases. This manuscript describes a sample preparation procedure that enables the use of secondary ion mass spectroscopy (SIMS) to obtain in situ &#948;34S values of various pyrite generations. This allows researchers to constrain how SO4-AOM affects pyritization in methane-bearing sediments. SIMS analysis revealed an extreme range in &#948;34S values, spanning from -41.6 to +114.8&#8240;, which is much wider than the range of &#948;34S values obtained by the traditional bulk sulfur isotope analysis of the same samples. Pyrite in the shallow sediment mainly consists of 34S-depleted framboids, suggesting early diagenetic formation by OSR. Deeper in the sediment, more pyrite occurs as overgrowths and euhedral crystals, which display much higher SIMS &#948;34S values than the framboids. Such 34S-enriched pyrite is related to enhanced SO4-AOM at the sulfate-methane transition zone, postdating OSR. High-resolution in situ SIMS sulfur isotope analyses allow for the reconstruction of the pyritization processes, which cannot be resolved by bulk sulfur isotope analysis.

Preparation of Authigenic Pyrite from Methane-bearing Sediments for In Situ Sulfur Isotope Analysis Using SIMS

Analyses of the sulfur isotopic composition (&#948;34S) of pyrite from methane-bearing sediments have typically focused on bulk samples. Here, we applied secondary ion mass spectroscopy to analyze the &#948;34S values of various pyrite generations to understand the diagenetic history of pyritization.