The Dame Was Loaded | |
---|---|
Developer(s) | Beam Software |
Publisher(s) | Philips Interactive Media |
Director(s) | Cameron Brown |
Designer(s) | David Giles |
Writer(s) | Mark Morrison |
Platform(s) | MS-DOS, Macintosh |
Release | 1996 |
Genre(s) | Adventure game |
Mode(s) | Single player |
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The Dame Was Loaded is a first-person point-and-click adventure game for the PC platform created by Australian developer Beam Software (now Krome Studios Melbourne) first published in 1996 by Philips Interactive Media.
Gameplay[edit]
The game is a detective film noir set in the 1940s and combined live action cinematic's with pre-rendered point-and-click gameplay much like previous games in the genre like Under a Killing Moon and Myst.[1] The live action was produced by Vixen Films, director Jo Lane, and was at the time the largest multimedia production ever made in Australia.
18 Games like The Dame Was Loaded for Playstation 4, daily generated comparing over 40 000 video games across all platforms. This suggestion collection includes real-time adventure games. The order in this selection is not absolute, but the best games tends to be up in the list. The bombed-out World War II relic was deliberately sunk - or scuttled - in 1951, loaded with 55-gallon drums of radioactive waste, and was recently rediscovered using sonar equipment. The Dame Was Loaded is a first person, point and click adventure game for the PC platform created by Australian developer Beam Software (now Melbourne House) first published in 1995 by Philips Interactive Media domestically then in 1996 world wide. To do this in The Dame, you have to return to your office all the time which takes too much valuable time. If your libido wins from your P.I. Gut feelings, at least you'll know what the title means; this dame is loaded. And so are you. With some bullets! So click on the car door and tell her to leave. She wants to have a last smoke but. Yesterday, S. Leblond et al. Published the paper First Observation of 20B and 21B in Phys. 121 (2018) 262502, describing the discovery of the neutron-unbound isotopes 20B and 21B. A summary of all isotope discoveries until the end of 2015 are compiled in my book The Discovery of Isotopes.
The game is nonlinear, having nine possible endings featuring over thirty fully acted and voiced characters to interact with and over one hour of fully performed cinematics.
Critical reception[edit]
A reviewer for Next Generation panned the game, citing the use of still shots instead of FMV for most of the character interactions, the low difficulty, and the 'cheesy, ersatz Maltese Falcon story'. He scored it one out of five stars.[2]
Quandary wrote 'In the end, the real time nature of the game prevented me from enjoying this outing...Timed puzzles are anathema to many adventure players'.[3]
External links[edit]
- The Dame Was Loaded on IMDb
- The Dame Was Loaded at MobyGames
References[edit]
- ^Hall, Mike (10 September 1996). Deckert, Rod (ed.). 'Extra goodies make Dame a challenge'. Albuquerque Journal. 116 (254). Albuquerque, New Mexico: Journal Publishing Co. p. B1,B3 – via Newspapers.com.
- ^'The Dame Was Loaded'. Next Generation. No. 19. Imagine Media. July 1996. p. 87.
- ^Ramsey, Steve (March 2003). 'The Dame Was Loaded'. Quandary. North Fremantle, Western Australia: Quandary Computer Game Reviews. Archived from the original on 27 May 2008.
Retrieved from 'https://en.wikipedia.org/w/index.php?title=The_Dame_Was_Loaded&oldid=900336653'
Abstract
Lewisite (2-chlorovinyldichloroarsine) is a chemical warfare agent developed during World War I. A quantitative method using solid phase extraction (SPE) followed by dual column liquid chromatography (LC)—isotope dilution tandem mass spectrometry (MS-MS) was developed for the determination of (2-chlorovinyl)arsonic acid (CVAOA), a metabolite of Lewisite, in human urine. The sample was treated with hydrogen peroxide to oxidize any (2-chlorovinyl)arsonous acid (CVAA) that remained in the trivalent arsenic oxidation state. There was 1.19% (arsenic purity) of bis-(2-chlorovinyl)arsinic acid (BCVAOA), a minor Lewisite metabolite, in the stock CVAA material. The high-throughput method qualitatively assessed BCVAOA simultaneously utilizing normal-phase silica SPE followed by reversed-phase C18 LC for an orthogonal separation. The chromatographic method results in a 5.8-min cycle time with adequate retention (k′ = 2.4) of CVAOA. The mass spectrometer was operated in positive electrospray ionization mode with quantitative m/z 186.9→61.0 and confirmation 186.9→91.0 mass transitions. This selective method demonstrated linearity, accuracy and reproducibility for the clinically relevant calibration range (25–3,200 µg/L as CVAA). The method detection limit was 3.3 µg/L as CVAA from a 10 µL injection. This LC–MS-MS emergency response method has a throughput of >240 samples (2.5 extracted 96-well plates) per day.
Introduction
The Organisation for the Prohibition of Chemical Weapons (OPCW), Chemical Weapons Convention (CWC) listed Lewisite (L) as a Schedule I chemical warfare agent (1). Lewisite was mass produced by several countries during World War I (2) and the destruction of stockpiles has been overseen by the OPCW. However, a few countries have not ratified the CWC and their stockpiles remain unaccounted for. Lewisite is a highly toxic vesicant (3, 4) that can be relatively easily synthesized by the catalytic reaction of arsenic trichloride and acetylene (5, 6). The relative ease of synthesis and availability of stockpiled agent makes Lewisite a chemical threat that could be used by terrorist organizations to endanger public health.
In the early 1990s, Lewisite ordnance rounds were unearthed in the Spring Valley neighborhood in Washington, DC, former site of American University Experimental Station (7, 8). The extent to which people were exposed to chemical warfare agent(s) or their breakdown products was not fully known. Although during soil excavations, some individuals reported burning eyes and respiratory irritation, which would be consistent with Lewisite exposure from the chemical munitions found. Clinical testing was limited to inorganic arsenic and total arsenic in urine and hair, and tested 40 individuals with arsenic levels within the normal reference range. Koryagina et al. (9) demonstrated the time-dependent urinary excretion profile of rats subcutaneously injected with Lewisite, with and without antidotal therapy. Lewisite urinary biomarkers persist in the body for a few days after an exposure; however, the highest concentrations are detected within the first 24 hours.
Lewisite metabolite detection is complex, because the catalytic reaction forms multiple products; (2-chlorovinyl)dichloroarsine (L1, Cl2AsC2H2Cl), bis(2-chlorovinyl)chloroarsine (L2, ClAs(C2H2Cl)2), tris(2-chlorovinyl)arsine (L3, As(C2H2Cl)3), byproducts and isomers. Weapons-grade Lewisite munitions contain, on average, 64.0% (wt%) L1, 28.5% L2 and <0.2% L3 with the remainder as impurities such as unreacted arsenic trichloride (10). The trans geometric isomer was predicted by ab initio calculations to dominate (11), and the geminal isomer (i.e., 1-chlorovinyl) was identified as a minor product (12). The primary metabolic pathway for Lewisite is hydrolysis; L1 is hydrolyzed to (2-chlorovinyl)arsonous acid (CVAA, (OH)2AsC2H2Cl), and L2 is hydrolyzed to bis(2-chlorovinyl)arsinous acid (BCVAA, (OH)As(C2H2Cl)2) with the native agent rarely found in nature (13, 14). The trivalent arsenic metabolites can be oxidized to the pentavalent arsenic metabolites; (2-chlorovinyl)arsonic acid (CVAOA, (OH)2As(=O)C2H2Cl) and bis(2-chlorovinyl)arsinic acid (BCVAOA, (OH)As(=O)(C2H2Cl)2), which can also be formed directly with reaction of hydrogen peroxide with the native agent (15). When a single mouse (16) and multiple rats (17) were exposed to trivalent CVAA, the major metabolite found was pentavalent CVAOA. Consequently, analytical methods must be able to detect the pentavalent arsenic metabolites.
Emergency response analytical methods must have short cycle times to allow timely analysis of several hundred to thousands of patient specimens. Gas chromatographic (GC) methods (9, 18–24) for the detection of Lewisite exposure do not meet requirements for high-throughput emergency response due to the lengthy chromatographic run-times and the long dithiol-derivatization incubation period needed to form volatile, thermally stable products amenable to GC analysis. However, GC coupled to electron-impact ionization mass spectrometry does yield valuable structural information that adds specificity to the detection. Liquid chromatography (LC) has been coupled to a harder-ionization technique, inductively coupled plasma (ICP), using arsenic-specific mass spectroscopic detection of Lewisite metabolites (16, 25, 26). Ion-pairing LC has been used to retain CVAA and CVAOA and separate these from other common arsenic compounds that may be found in urine (26), or oxidize the sample to measure only the pentavalent arsenic metabolites (25). LC has also been coupled to a softer-ionization technique, electrospray ionization (ESI), with tandem mass spectroscopic detection (17, 20, 27). Although CVAA is retained in this method, the pentavalent CVAOA metabolite is not well retained by reverse-phased C18 chromatography and requires long run-times to elute hydrophobic urinary compounds. Additionally, CVAOA is more easily ionized than CVAA by positive-ion ESI, leading to increased sensitivity of detection. The LC–ESI–MS-MS methods have more specificity with an additional confirmation ion, compared to mono-isotopic ICP–MS detection. However, the chromatographic method lacks the resolution to separate the CVAOA metabolite from other urinary interferences that are not retained. The method also lacks an isotopically labeled internal standard to properly compensate for solid phase extraction (SPE) recoveries and matrix effects/ion suppression. Phenylarsonic acid, pentavalent arsenic, is used as a surrogate internal standard (17), but does not monitor AsIII→AsV oxidation. Phenylarsine oxide, trivalent arsenic, rapidly oxidizes to phenylarsonic acid in the presence of water. We describe an isotope dilution SPE–LC–ESI–MS-MS method using dual column reverse-phased chromatography, with a 5.8-min cycle time, that retains the CVAOA and BCVAOA metabolites.
Experimental
Materials
Inorganic- and organic-free 18 MΩ cm water was purified by EMD Millipore (Billerica, MA) Super-Q plus water purification system equipped with a 0.22 µm filter. Methanol (Optima™ grade), acetonitrile (HPLC grade), formic acid (99% pure), 30% hydrogen peroxide (ACS reagent grade), nitric acid (Optima™ grade) and L-cysteine (>98% pure) were purchased from Thermo Fisher Scientific (Fair Lawn, NJ).
The US Army Medical Research Institute of Chemical Defense (Aberdeen Proving Ground, MD) provided semi-purified hydrolyzed Lewisite to the Centers for Disease Control and Prevention (CDC) National Center for Environmental Health (NCEH). An aqueous stock CVAA standard was provided to the Florida Department of Health, Bureau of Public Health Laboratories, a member of the Laboratory Response Network—Chemical (LRN-C). The concentration was determined by diluting (1:200) the stock material and measuring total arsenic against a National Institute of Standards and Technology traceable standard purchased from High Purity Standards (Charleston, SC). An additional 1:20 dilution was applied with ICP–MS diluent (2% nitric acid; 10 µg/ L iridium, m/z 193 as an internal standard) to be within the 5–4,000 µg/L calibration range. Total arsenic was measured by ICP–MS at m/z 91 using a PerkinElmer (Shelton, CT) DRCII with 1.2 mL/min oxygen (research grade >99.999% Airgas, Radnor, PA) in the dynamic reaction cell (RPa = 0, RPq = 0.75).
A 100 µL aliquot of the 1:200 diluted stock standard was further diluted with 300 µL of mobile phase (11.5 mM tetrabutylammonium hydroxide, 5 mM succinic acid, 2% isopropyl alcohol and pH 5.5) and 100 µL of water and analyzed by an Agilent Technologies (Santa Clara, CA) 1200 series LC and 8800 ICP–MS-MS using conditions described (25) with and without the addition of hydrogen peroxide. A 4.6 × 150-mm, 5-µm RP-amide column from Sigma-Aldrich (Milwaukee, WI) at 1.0 mL/min was used for better resolution of L1 metabolites, and a 32-min run-time was used to elute the L2 metabolite(s). The unoxidized sample was used to determine the extent of CVAA oxidation to CVAOA. The stock standard was determined to be predominately CVAA, gem-CVAOA was not detected. The oxidized sample was used to determine the arsenic purity: 91.38% CVAA (+CVAOA), 2.97% gem-CVAA and 1.19% BCVAA (+BCVAOA), with the remainder as inorganic arsenic, to correct for the stock standard concentration.
The concentration was also corrected for the molecular/elemental arsenic ratio (i.e., CVAA, 170.4/74.9). The calculated concentrations were 2.132, 0.0693 and 0.0350 mg/mL for CVAA, gem-CVAA and BCVAA, respectively. Analytical standards for CVAA (25, 50, 100, 200, 400, 800, 1,600 and 3,200 µg/L) and quality controls (QC) for CVAA (75, 300 and 1,200 µg/L) were prepared in base urine. After volumetric preparation of matrix-matched calibrators and QC, they were subsequently stored at ≤–70°C in cryogenic vials until use. The base urine is defined as urine from laboratory volunteers collected with informed consent. The collection protocol was described in detail elsewhere (25) and approved by the Florida Department of Health Institutional Review Board (IRB). Isotopically labeled CVAA-13C2D2 was synthesized by Los Alamos National Laboratory (Los Alamos, NM). The concentration was determined at 1.023 mg/mL of CVAA-13C2D2 (99.17% pure, as arsenic) by LC–ICP–MS-MS, analogous to CVAA as described above. The gem-CVAA-13C2D2 was not detected, and only trace (<0.1%) levels isotopically labeled BCVAA (+BCVAOA) was detectable in the stock standard.
Individual de-identified human urine specimens were collected in coordination with the CDC NCEH for the purpose of screening current analytical methods for interferences. Urine specimens were collected from volunteers of participating LRN-C laboratories with informed consent. Specimens were screened for interferences by the described method. Since specimens tested were de-identified the research was determined exempt from IRB review by the Florida Department of Health Human Research Protection Program (DOH IRB Number: H112320).
Sample preparation
All urine samples were thawed to room temperature prior to analysis. A 125 µL aliquot of urine was vigorously mixed with 20 µL of internal standard (5.0 mg/L of CVAA-13C2D2 in water) and 20 µL of 30% hydrogen peroxide into a 2-mL 96-well plate on an IKA® Works (Wilmington, NC) micro-titer shaker for 5 min. Hydrogen peroxide was used in excess to oxidize AsIII to AsV species. The hydrogen peroxide oxidation reaction is considered instantaneous, with respect to time of detection. When a drop of 30% H2O2 was added to an aqueous CVAA solution and immediately injected and detected by LC–ICP–MS, complete conversion of CVAA to CVAOA was observed in <2 min. Then, 1.4 mL of acetonitrile was added and gently mixed for an additional 5 min.
For post-extraction addition, the CVAA material was oxidized with hydrogen peroxide prior to addition to produce CVAOA at three different levels; 75, 300 and 1,200 µg/L in urine. The internal standard was not added, but the 20 µL of 30% hydrogen peroxide was added to oxidize the urine. Water was substituted for the 20 µL spikes or 125 µL of urine when appropriate. The percent recovery was determined by the pre-extraction spike divided by the post-extraction spike. The relative ionization efficiency was determined by the post-extraction spike in matrix divided by the post-extraction spike in water. The process efficiency was determined by the pre-extraction spike in matrix divided by the post-extraction spike in water.
The 2-mL 96-well plate was placed on the PerkinElmer Zephyr liquid handler for automated SPE. The SPE was based on structural similarity of alkyl-arsonic acids with alkyl-phosphonic acids (28) with an additional conditioning and washing step added. The Phenomenex (Torrance, CA) Silica Si-1 (100 mg per well) was conditioned in three steps: 1 mL of 75% acetonitrile/25% water, 1 mL of 80% acetonitrile/20% methanol, followed by 1 mL of 100% acetonitrile. A 1.5-mL extract of sample was loaded onto the extraction plate. This was followed by a three-step wash in 1 mL 100% acetonitrile, 1 mL 80% acetonitrile/20% methanol and 1 mL 90% acetonitrile/10% water. The analytes were eluted with 1 mL 75% acetonitrile/25% water into a clean 2 mL 96-well plate.
The samples were evaporated to dryness with the Biotage (Charlotte, NC) TurboVap® 96 evaporator. The eluent was concentrated for 15 min, or until half the sample volume remained, with 35 standard cubic feet per hour (SCFH) nitrogen at 70°C. The samples were gently mixed on the micro-titer shaker for 1 min. Then, the samples in the 2 mL 96-well elution plate were evaporated to dryness (typically another 45–60 min) with 70 SCFH nitrogen at 70°C. The samples were reconstituted with 100 µL water, and then mixed vigorously for 5 min. The reconstituted samples were transferred to a 200 µL 96-well plate, and heat foil sealed with a Thermo Fisher Scientific ALPS™ 25 manual heat sealer.
Liquid chromatography
An Agilent Technologies 1200 series LC was used consisting of two binary solvent pumps with a degasser for each pump, a temperature-controlled autosampler and a temperature-controlled column compartment with an integrated 2-position 10-port switching valve. Separation was achieved with Waters (Milford, MA) SunFire C18 reversed-phase dual columns (2.1 × 10-mm, 2.5-µm guard column and 2.1 × 30-mm, 2.5-µm analytical column) and configured to the switching valve as shown in Figure 1. The columns were maintained at 30 ± 1°C in the column compartment. The mobile phase consisted of 20 mM formic acid in water (A) and 20 mM formic acid in methanol (B) and used unfiltered. The primary and secondary binary pumps flow rates were 400 µL/min with the time table defined with %B composition and switching valve position in Table I. The 200 µL 96-well plate was maintained at 4 ± 1°C from which 10 µL was injected on the column.
Dual columns allowed for high-throughput analysis of Lewisite metabolites.
Dual columns allowed for high-throughput analysis of Lewisite metabolites.
Time table used for chromatography