Filtration of water samples by a 0.45-m membrane filter was done, and the heavy metals were pre-concentrated and separated from seawater samples by the ammonium pyrrolidine dithiocarbamate (APDC)/methyl isobutyl ketone (MIBK) solvent extraction technique (Eaton et al., 1995 ; Folk, 1980 ). Finally, the metals in the organic layer were extracted using 50% HNO3 and collected in a polyethylene bottle to be analyzed by atomic absorption spectrometry (FAAS PerkinElmer model A Analyst 100) for Zn2+, Fe2+, Pb2+, Co2+, Mn2+, Ni2+, and Cd2+. On the other hand, the sediments were dried for 48 h at 60 °C in a thermostatically controlled oven, homogenized with an agate pestle and mortar and sieved using a 63-μm sieve. In a dry Teflon beaker, 0.5 g of fine sediment powder was thoroughly digested at 85 °C with a mixed acid solution containing HNO3:HClO4 (3:1 v/v) according to the method described by Oregioni and Aston (1984 ). Studied metals were analyzed by FAAS (PerkinElmer model A Analyst 100), and the results were expressed as mg/kg. Each heavy metal was analyzed in three replicates, and the results were presented as mean (Chester et al., 1994 (link); Oregioni & Aston, 1984 )
Aanalyst 100
Manufactured by PerkinElmer
Sourced in United States, United Kingdom, Australia, Germany
The AAnalyst 100 is a compact, single-beam atomic absorption spectrometer designed for routine analysis of metals and metalloids in a variety of sample types. The instrument utilizes a high-intensity, long-life lamp to provide the light source for atomic absorption measurements. The AAnalyst 100 features a simple, intuitive user interface and is capable of performing flame atomic absorption spectroscopy (FAAS) analysis.
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Lab products found in correlation
Uv 1800, by Shimadzu (2 mentions)
Nitric acid, by Honeywell (1 mentions)
J2 hc, by Beckman Coulter (1 mentions)
No 42 filter paper, by Cytiva (1 mentions)
Whatman, by Cytiva (1 mentions)
Nexion 350d, by PerkinElmer (1 mentions)
Ethos e, by Milestone (1 mentions)
Mhs 10, by PerkinElmer (1 mentions)
Lambda 365, by PerkinElmer (1 mentions)
Sonoplus, by Bandelin (1 mentions)
Nitric acid, by PerkinElmer (1 mentions)
Hydrochloric acid (hcl), by PerkinElmer (1 mentions)
Hydrochloric acid (hcl), by Merck Group (1 mentions)
Pinaacle 900z, by PerkinElmer (1 mentions)
Centrifuge 5415d, by Eppendorf (1 mentions)
50 protocols using aanalyst 100
1
Quantitative Analysis of Heavy Metals in Seawater and Sediments
2
Soil Silver Concentration Analysis
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This article is protected by copyright. All rights reserved hydrochloric acid (37% p.a., Baker, Grainger, USA) and nitric acid (65% p.a., Riedel-de-Haen, Seelze, Germany) in closed Teflon® bombs heated at 140 °C for 7 h. Digests were diluted with 8 ml of deionised water and analysed for total Ag by flame atomic absorption spectroscopy (Perkin Elmer AAnalyst 100). For quality assurance purposes, a certified reference material (River Clay, WEPAL-ISE-886) and reagent blanks were also analysed. The averages of measured Ag concentrations were within 25% of the certified reference value (River Clay contains 2.8 ± 0.4 mg Ag kg -1 , silver concentrations in the certified reference material ranged between 2.6 and 3.9 mg Ag kg -1 dry weight).
No Ag was detected in the blank samples. The experimental detection limit for the Ag measurements was 0.3 mg kg -1 .
Soil pore water samples were also collected for Ag analysis from separately prepared replicate soil samples by centrifugation (J2-HC, Beckman Coulter, California) for all four Ag treatments assessed. A 50 g batch of spiked soils was taken from these four replicates and saturated with 13.5 ml of Milli-Q water and centrifuged for 60 min at 4000 xg [7] . The samples (supernatants) were then acidified and analysed for total Ag concentration by flame atomic absorption spectroscopy (Perkin Elmer AAnalyst 100).
No Ag was detected in the blank samples. The experimental detection limit for the Ag measurements was 0.3 mg kg -1 .
Soil pore water samples were also collected for Ag analysis from separately prepared replicate soil samples by centrifugation (J2-HC, Beckman Coulter, California) for all four Ag treatments assessed. A 50 g batch of spiked soils was taken from these four replicates and saturated with 13.5 ml of Milli-Q water and centrifuged for 60 min at 4000 xg [7] . The samples (supernatants) were then acidified and analysed for total Ag concentration by flame atomic absorption spectroscopy (Perkin Elmer AAnalyst 100).
3
Quantifying Cadmium in Soil and Plants
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For determination of Cd in soil, 10 g air-dried soil subsamples were placed in 20 mL of DTPA solution (0.05 mol L−1 at pH 7.3) and shaken on a reciprocal shaker at 200 rpm for 15 min [94 (link)]. Cadmium concentration in soil samples was then measured directly from soil extracts after filtration by AAS. For determination of Cd concentration in plants tissues, samples were dried in an oven at 70 °C for 24 h and then ground, weighed, and digested with 2 mL nitric acid (HNO3) and 1 mL per chloric acid (HClO4) (2:1 ratio v/v). Afterward, samples were heated on a hot plate at 350 °C until dense white fumes appeared. The contents of these flasks were cooled, filtered, and stored in plastic bottles for further determinations by AAS (Perkin Elmer Aanalyst-100, Waltham, USA) [95 (link)].
4
Arsenic Accumulation and Export Assay
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Determination of cellular arsenic accumulation and export was modified from a previous report [35 (link)]. Cells were grown in arsenic-free medium to approximately 70% confluence. The medium was replaced with either fresh medium or medium containing 10 μM sodium arsenite. After 24 h, to determine arsenic accumulation, cells were washed three times with PBS, harvested with trypsin, counted, and digested overnight with 50% perchloric acid : nitric acid (2 : 1) at 70°C; to determine arsenic export, PBS-washed cells were incubated in fresh arsenic-free medium for another 24 h. Then, medium was collected, and cells were trypsinized and counted. Arsenic concentration in the cell or medium was determined by graphite furnace atomic absorption spectrophotometry using a Perkin Elmer AAnalyst 100. Arsenic concentrations were normalized to cell number.
5
Copper Toxicity in Larval Development
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Groups of 300 newly hatched larvae (within 8 hr) in the 20th generation (F
20) were fed on diets containing Cu
2+at concentrations of 0, 100, and 800 µg/g. Each group was reared in a glass bottle, and each concentration was replicated three times. After four days of treatment, 30 larvae from each group were picked randomly, washed with distilled water, and starved for 24 hr. They were then dried on paper towels and weighed on an electronic scale (AB204-E, Mettler Toledo,
www.mt.com ). Each treatment was divided into four groups with one group of larvae treated with xylene:ethanol (1:1) solution to measure larval body length by using a vernier caliper after stretching. The second group of larvae was used to determine tissue metal content by using an atomic absorption spectrophotometer (AAnalyst100, Perkin Elmer,
www.perkinelmer.com ) after digestion with 1 mL mixed acid (HClO
4:HNO
3=1:5 v/v) (
Wu et al
.2007
). The third group was used to determine enzyme activity, and the remaining larvae in the fourth group were allowed to pupate, emerge, and mate to determine egg production by individual females.
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Groups of 300 newly hatched larvae (within 8 hr) in the 20th generation (F
20) were fed on diets containing Cu
2+at concentrations of 0, 100, and 800 µg/g. Each group was reared in a glass bottle, and each concentration was replicated three times. After four days of treatment, 30 larvae from each group were picked randomly, washed with distilled water, and starved for 24 hr. They were then dried on paper towels and weighed on an electronic scale (AB204-E, Mettler Toledo,
4:HNO
3=1:5 v/v) (
.2007
6
Determination of Cadmium in Soil and Plant Samples
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For determining DTPA extractable Cd content in soils receiving different treatments, 10 g air dried soil sample was suspended in 20 mL DTPA extractant solution, suspension was shaken for 15 min at 200 rpm on reciprocal shaker [45 ]. Samples were filtered through Whatman No. 42 filter paper. Cadmium in extract was directly measured using atomic absorption spectrometer (PerkinElmer, AAnalyst 100, Waltham, USA). The shoot and root samples were dried in an oven at 65°C to a constant weight, ground in Wiley mill and 0.5 g of ground material was digested using HNO3: HCLO4 (3:1) mixture. After digestion, the contents in flasks were cooled, filtered properly and stored in washed plastic bottles for subsequent analysis of Cd on atomic absorption spectrometer following the method described by Yong et al. [46 ].
7
Waste PCB Characterization and Bioleaching
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The waste PCBs used in this study were obtained from Comimtel Recycling, Peru. The preparation of waste PCBs was carried out as follows: (i) manually separated electronic components (for example, capacitors, cards, batteries, resistors, among others) were size reduced by using metal cutting scissors and a portable powder crusher (Keene Engineering, Chatsworth, CA, USA), until obtaining a fine powder with a particle size ≤ 300 µm [32 (link),33 (link)]; (ii) powder samples were then washed with a saturated solution of NaCl 35% (w/v), at a ratio of 10 g/100 mL, and were dried in an oven at 60 °C for 24 h (for the elimination of plastic particles potentially toxic to bacterial metabolism) [5 (link),34 (link)].
For the characterization of waste PCBs metal content, the powder obtained was digested with an acid mixture of HNO3, HCl, HF and HClO4 in a ratio of 5:1:2:2, respectively [35 (link)]. The aforementioned digested solution was analyzed by Atomic Absorption Spectrophotometry (AAS) (AAnalyst100-Perkinelmer, Waltham, MA, USA), while the bioleaching solutions were analyzed by Inductively Coupled Plasma Optical Emission Spectrophotometry (ICP-OES) (Nexion350D-Perkinelmer, Waltham, MA, USA).
For the characterization of waste PCBs metal content, the powder obtained was digested with an acid mixture of HNO3, HCl, HF and HClO4 in a ratio of 5:1:2:2, respectively [35 (link)]. The aforementioned digested solution was analyzed by Atomic Absorption Spectrophotometry (AAS) (AAnalyst100-Perkinelmer, Waltham, MA, USA), while the bioleaching solutions were analyzed by Inductively Coupled Plasma Optical Emission Spectrophotometry (ICP-OES) (Nexion350D-Perkinelmer, Waltham, MA, USA).
8
Measurement of Iron and Zinc in Sera
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To avoid contamination, all the reagents were carefully handled, and polyethylene disposables were thoroughly washed with HCl 1 N under a fume hood. All the reagents were from Merck (Darmstadt, Germany); the acids were of Suprapur grade. Samples of sera were diluted 1:10 with bidistilled water and were directly analyzed using a flame atomic spectrophotometer (AAnalyst 100, PerkinElmer, Waltham, MA, USA). The accuracy of the method was evaluated by analyzing an international standard (ERM®-BB422 fish muscle). The concentrations found with the method used in this study fell into the certified uncertainty interval given by the ERM, corresponding to a 95% confidence level. The detection limits were 0.04 μg mL−1 and 0.01 μg mL−1 for iron and zinc, respectively. Element concentrations were reported as μg mL−1.
9
Zn Content Quantification in Cell Media
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During the NRU assay, the Zn content of the cell medium was quantified to check the level of ZnO dissolution in the cell media. In order to assess the release of Zn within the first half of the 7-day exposure, the cell medium was sampled after 3 days of cell exposure, then the cell medium was replaced with a fresh one and again sampled after another 4 days (at the end of the exposure). The selection of the sampling time was in line with the cell viability assessment (NRU assay). Prior to the analysis, 0.75 mL of concentrated nitric acid (65 % HNO3, pro analysis, Carlo Erba Reagenti, Arese, Italy) was added to 0.75 mL of the cell medium from each of the test vessels. The samples were digested in the Milestone Ethos E (Bergamo, Italy) microwave lab station equipped with an SK-10 high-pressure segmented rotor and 3 mL quartz microsampling inserts. The digestion was conducted at 180 °C and 600 W power, with step 1 (heating) lasting 15 min, step 2 (constant temperature) lasting 10 min, and cooling to 60 °C lasting 45 min. The total Zn concentrations in the samples were measured by flame atomic absorption spectroscopy (AAS; Perkin-Elmer AAnalyst 100, Waltham, MA, USA).
10
Measuring Leaf Zinc Concentrations
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Whole-leaf zinc concentrations were measured as described in [13 (link)]. Briefly, fresh leaf material was oven-dried, digested in concentrated nitric acid, diluted 10-fold with ultrapure water and filtered. Metal concentrations were measured in an air–acetylene flame by atomic absorption spectrophotometry, using a double-beam optical system with deuterium arc background correction (AAnalyst 100; Perkin-Elmer, UK). Two independent, replicate experiments were performed, in which the zinc concentration in plants of the four populations, each grown on each of the four metal regimes for eight weeks, was measured. For each treatment/population combination, three biological replicates, each consisting of pooled leaf tissue from 5 to 10 plants, were used; for each of these biological replicates, six technical replicates were measured.
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