The forgotten experiment
T
he brown cardboard box was taped up and weathered with age. On it, someone had scribbled “Electric Discharge Samples” and “1953⫺1954.” The old box sat, unnoticed, in a laboratory, giving no hint of its extraordinary contents. For several years, Jeffrey Bada at the Scripps Institution of Oceanography had no idea the box was even in his possession, but then a chance remark by a friend drew Bada’s attention to it. When he and his colleagues opened up the box, they discovered a scientific treasure trove. For in it were vials containing the syrupy, brown residues produced by the famous Miller⫺Urey experiment and its derivationsOthe first attempts to re-create the conditions under which primordial organic compounds might have first appeared on Earth. Bada and colleagues applied modern analytical techniques to these residues to see if they still held clues to the mystery of the origins of life. And, much to the researchers’ surprise, they did (Science 2008, DOI 10.1126/ science.1161527).
How Bada got the box When Stanley Miller was a graduate student in the laboratory of Harold Urey at the University of Chicago, he carried out a series of experiments in an attempt to re-create the conditions of early Earth. Miller sealed a reducing atmosphere of water, methane, ammonia, and hydrogen into a sterile flask with a pair of electrodes that provided sparks to emulate lightning. After a week of heating, cooling, and sparking, Miller tested the residue in the flask and found that amino acids had formed (Science 1953, DOI 10.1126/ science.117.3046.528). The experiment ultimately became known as the 6
ANALYTICAL CHEMISTRY /
JANUARY 1, 2009
Miller⫺Urey experiment (Origins Life Evol. Biosph. 2003, 33, 235⫺242). As Miller progressed through his career as a faculty member at the University of California San Diego (UCSD), he focused on origin-of-life research and developed elaborations of his original work. Bada was his second graduate student. After suffering a stroke in 1999, Miller asked Bada to take custody of his laboratory. When Miller’s laboratory finally closed down a few years later, Bada moved most of the contents to his own laboratory and, after some basic organization of the material, left it to collect dust. Shortly before Miller’s death in 2007, Bada met up with Antonio Lazcano of the National Autonomous University of Mexico, who was also a longtime friend of Miller’s. Bada recounts a conversation with Lazcano: “Antonio mentioned in passing, ‘You know, Jeff, one time I was in Stanley’s office, [and] he pointed up to a cardboard box and said, “I saved some of my residues from my original experiments.” ’ ” Bada was dumbstruck. There were still residues left from that famous experiment? “I had known Stanley for 40 years, and he never told me that,” he says. “I realized I must have the box.”
The lesser-known Miller experiment Bada and his colleagues hunted through the Miller hand-me-downs and soon found the cardboard box. When they opened it up, they found smaller white boxes. The smaller boxes contained 3 mL glass vials with labels taped on them and the dried, brown residues inside (Figure 1). “We could tell from the labels on the boxes that they came from the original experiments,” says Bada.
COURTESY OF ADAM JOHNSON
A variation on the famous Miller-Urey experiment emerges more than 50 years later to reveal possible details of the origins of life.
Figure 1. Miller took extraordinary care to catalog and document his experiments. “If you want to have a wonderful lesson for graduate students on why it’s so important to take really good notes in your notebook, this is it,” says Bada.
Miller had been meticulous. The vials were clearly labeled with an experiment number and a page number from his laboratory notebooks in which he had described the experiments. Miller’s notebooks were in the archives at UCSD, so Adam Johnson of Indiana University, who was doing an internship in Bada’s laboratory, went to the university and tracked down the information from Miller’s notes. Bada says, “We knew right then we had extracts from all those original 1953⫺1954 experiments.” The apparatus used in the classical experiment has been immortalized in textbooks, but Miller had actually tested two other setups at the same time. One setup was simply a variation on how the electric discharge was generated. The other setup caught Bada’s eye. A small nozzle on top of the flask allowed steam into the flask as the spark was generated (Figure 2). Bada remembered that Miller had published a single paper on this experimental
10.1021/AC8023922 2009 AMERICAN CHEMICAL SOCIETY
Published on Web 12/09/2008
acids and other organic compounds. Upon finding the vials, Bada says he realized that “here, we had that experiment right in front of us.”
Modern analytical chemistry on old samples
Figure 2. The volcanic spark discharge apparatus that Miller used. (Adapted from J. Am. Chem. Soc. 1955, 77, 2351-2361.)
setup in 1955 (J. Am. Chem. Soc. 1955, 77, 2351⫺2361). The classical experiment had been criticized because many scientists thought that the global atmosphere of early Earth couldn’t be as reducing as the experimental conditions that Miller had used. But adding steam represented a scenario that Bada had been considering in his own line of research. He wondered whether the apparatus could mimic a more localized synthesis of primordial organic molecules around a volcanic eruption where, he says, “we know today, there is intense lightning.” Bada says it’s conceivable that even if the global atmosphere wasn’t reducing, volcanoes could have spewed out gases, creating local regions of a reducing atmosphere. With the fierce lightning around volcanoes, these gases undergo reactions to locally synthesize amino
The investigators decided to go through all the extracts from the three sets of apparatus with a particular focus on the spark-and-steam setup, which they named the “volcanic spark discharge apparatus”. When Miller did his experiments, he used a combination of paper and ion exchange chromatography followed by a ninhydrin treatment to detect the amino acids. But Bada and colleagues took advantage of the leaps made in quantitative analytical chemistry in the past 50 years and carried out HPLC and LC/TOFMS on the residues. The investigators detected 22 amino acids and 5 amines in the samples from the volcanic spark discharge apparatus, several of which Miller himself had identified. Compared with the other two setups, residues from the volcanic spark discharge apparatus had a greater diversity and yield of amino acids. The experiment “confirmed the idea that with these little volcanic-island-type systemsOlike the volcanoes of Hawaii dotting the surface of the oceanOyou could have had a whole string of these tiny localized chemical factories, rather than a global factory,” says Bada. The investigators found hydroxylated amino acids, which have never been documented before. Bada speculates that as steam was injected into the spark, it dissociated to form hydroxyl radicals.
These radicals then attacked either the precursors of the amino acids or the amino acids themselves to generate the hydroxyl-substituted amino acids. “This is the first time we have seen these compounds,” says Bada. “Now, as far as whether or not they might be important in early prebiotic chemistry, we don’t know yet because no one has ever thought of them.” Bada says concerns about the 55year-old residues being contaminated are unjustified. Amino acids that are created in abiotic environments, like the experimental setup Miller worked with, are racemic. Contaminating amino acids would be exclusively L-isomers, which would have thrown off the enantiomeric balance; Bada and colleagues were unable to find evidence of this.
More than 50 years later, the experiments are still relevant Bada now thinks the volcanic spark discharge experiment can compete with other theories of how organic material appeared on Earth. One theory postulates that meteorites brought amino acids from outer space, but Miller’s experiment also suggests that “localized syntheses... could have been an effective contributor to the organic material on the early Earth,” says Bada. What would his old mentor think of the attention his neglected experiment is now receiving? Bada says, “I think he’d be pleased that there is still a lot of respect for this experiment. It still is very relevant to our understanding of the processes that took place on the early Earth that led to the origin of life.” —Rajendrani Mukhopadhyay
JANUARY 1, 2009 / ANALYTICAL CHEMISTRY
7
