Nucleic acid analogue

From Wikipedia, the free encyclopedia

https://en.wikipedia.org/wiki/Nucleic_acid_analogue 

RNA with its nucleobases to the left and DNA to the right.

Nucleic acid analogues are compounds which are analogous (structurally similar) to naturally occurring RNA and DNA, used in medicine and in molecular biology research. Nucleic acids are chains of nucleotides, which are composed of three parts: a phosphate backbone, a pentose sugar, either ribose or deoxyribose, and one of four nucleobases. An analogue may have any of these altered. Typically the analogue nucleobases confer, among other things, different base pairing and base stacking properties. Examples include universal bases, which can pair with all four canonical bases, and phosphate-sugar backbone analogues such as PNA, which affect the properties of the chain (PNA can even form a triple helix). Nucleic acid analogues are also called Xeno Nucleic Acid and represent one of the main pillars of xenobiology, the design of new-to-nature forms of life based on alternative biochemistries.

Artificial nucleic acids include peptide nucleic acid (PNA), Morpholino and locked nucleic acid (LNA), as well as glycol nucleic acid (GNA), threose nucleic acid (TNA) and hexitol nucleic acids (HNA). Each of these is distinguished from naturally occurring DNA or RNA by changes to the backbone of the molecule.

In May 2014, researchers announced that they had successfully introduced two new artificial nucleotides into bacterial DNA, and by including individual artificial nucleotides in the culture media, were able to passage the bacteria 24 times; they did not create mRNA or proteins able to use the artificial nucleotides. The artificial nucleotides featured 2 fused aromatic rings.

Medicine

Main article: Nucleoside analogues

Several nucleoside analogues are used as antiviral or anticancer agents. The viral polymerase incorporates these compounds with non-canonical bases. These compounds are activated in the cells by being converted into nucleotides, they are administered as nucleosides since charged nucleotides cannot easily cross cell membranes.

Molecular biology

Common changes in nucleotide analogues

Nucleic acid analogues are used in molecular biology for several purposes: Investigation of possible scenarios of the origin of life: By testing different analogs, researchers try to answer the question of whether life's use of DNA and RNA was selected over time due to its advantages, or if they were chosen by arbitrary chance; As a tool to detect particular sequences: XNA can be used to tag and identify a wide range of DNA and RNA components with high specificity and accuracy; As an enzyme acting on DNA, RNA and XNA substrates - XNA has been shown to have the ability to cleave and ligate DNA, RNA and other XNA molecules similar to the actions of RNA ribozymes; As a tool with resistance to RNA hydrolysis; Investigation of the mechanisms used by enzyme; Investigation of the structural features of nucleic acids.

Backbone analogues

Hydrolysis resistant RNA-analogues

Chemical structure of Morpholino

To overcome the fact that ribose's 2' hydroxy group that reacts with the phosphate linked 3' hydroxy group (RNA is too unstable to be used or synthesized reliably), a ribose analogue is used. The most common RNA analogues are 2'-O-methyl-substituted RNA, locked nucleic acid (LNA) or bridged nucleic acid (BNA), morpholino, and peptide nucleic acid (PNA). Although these oligonucleotides have a different backbone sugar or, in the case of PNA, an amino acid residue in place of the ribose phosphate, they still bind to RNA or DNA according to Watson and Crick pairing, but are immune to nuclease activity. They cannot be synthesized enzymatically and can only be obtained synthetically using phosphoramidite strategy or, for PNA, methods of peptide synthesis.

Other notable analogues used as tools

Dideoxynucleotides are used in sequencing . These nucleoside triphosphates possess a non-canonical sugar, dideoxyribose, which lacks the 3' hydroxyl group normally present in DNA and therefore cannot bond with the next base. The lack of the 3' hydroxyl group terminates the chain reaction as the DNA polymerases mistake it for a regular deoxyribonucleotide. Another chain-terminating analogue that lacks a 3' hydroxyl and mimics adenosine is called cordycepin. Cordycepin is an anticancer drug that targets RNA replication. Another analogue in sequencing is a nucleobase analogue, 7-deaza-GTP and is used to sequence CG rich regions, instead 7-deaza-ATP is called tubercidin, an antibiotic.

Precursors to the RNA world

Main article: RNA world hypothesis

RNA may be too complex to be the first nucleic acid, so before the RNA world several simpler nucleic acids that differ in the backbone, such as TNA and GNA and PNA, have been offered as candidates for the first nucleic acids.

Base analogues

Nucleobase structure and nomenclature

Purine

 

Pyrimidine

 

Main article: Nucleobase

 

For structures of the analogues that may be mentioned in the literature, see Simple aromatic ring.

Naturally occurring bases can be divided into two classes according to their structure:

• pyrimidines are six-membered heterocyclic with nitrogen atoms in position 1 and 3.
• purines are bicyclic, consisting of a pyrimidine fused to an imidazole ring.

Artificial nucleotides (Unnatural Base Pairs (UBPs) named d5SICS UBP and dNaM UBP) have been inserted into bacterial DNA but these genes did not template mRNA or induce protein synthesis. The artificial nucleotides featured two fused aromatic rings which formed a (d5SICS–dNaM) complex mimicking the natural (dG–dC) base pair.

Mutagens

One of the most common base analogs is 5-bromouracil (5BU), the abnormal base found in the mutagenic nucleotide analog BrdU. When a nucleotide containing 5-bromouracil is incorporated into the DNA, it is most likely to pair with adenine; however, it can spontaneously shift into another isomer which pairs with a different nucleobase, guanine. If this happens during DNA replication, a guanine will be inserted as the opposite base analog, and in the next DNA replication, that guanine will pair with a cytosine. This results in a change in one base pair of DNA, specifically a transition mutation.

Additionally, HNO2, or nitrous acid is a potent mutagen that acts on replicating and non-replicating DNA. It can cause deamination of the amino groups of Adenine, Guanine and Cytosine. Adenine is deaminated to hypoxanthine, which base pairs to cytosine instead of thymine. Cytosine is deaminated to uracil, which base pairs with Adenine instead of Guanine. Deamination of Guanine is not mutagenic. Nitrous acid-induced mutations also are induced to mutate back to wild-type using nitrous acid.

Fluorophores

Main article: Fluorophore

 

Structure of aminoallyl-uridine

Commonly fluorophores (such as rhodamine or fluorescein) are linked to the ring linked to the sugar (in para) via a flexible arm, presumably extruding from the major groove of the helix. Due to low processivity of the nucleotides linked to bulky adducts such as florophores by taq polymerases, the sequence is typically copied using a nucleotide with an arm and later coupled with a reactive fluorophore (indirect labelling):

• amine reactive: Aminoallyl nucleotide contain a primary amine group on a linker that reacts with the amino-reactive dye such as a cyanine or Alexa Fluor dyes, which contain a reactive leaving group, such as a succinimidyl ester (NHS). (base-pairing amino groups are not affected).
• thiol reactive: thiol containing nucleotides reacts with the fluorophore linked to a reactive leaving group, such as a maleimide.
• biotin linked nucleotides rely on the same indirect labelling principle (+ fluorescent streptavidin) and are used in Affymetrix DNAchips.

Fluorophores find a variety of uses in medicine and biochemistry.

Fluorescent base analogues

The most commonly used and commercially available fluorescent base analogue, 2-aminopurine (2-AP), has a high-fluorescence quantum yield free in solution (0.68) that is considerably reduced (appr. 100 times but highly dependent on base sequence) when incorporated into nucleic acids. The emission sensitivity of 2-AP to immediate surroundings is shared by other promising and useful fluorescent base analogues like 3-MI, 6-MI, 6-MAP, pyrrolo-dC (also commercially available), modified and improved derivatives of pyrrolo-dC, furan-modified bases and many other ones (see recent reviews). This sensitivity to the microenvironment has been utilized in studies of e.g. structure and dynamics within both DNA and RNA, dynamics and kinetics of DNA-protein interaction and electron transfer within DNA.

A newly developed and very interesting group of fluorescent base analogues that has a fluorescence quantum yield that is nearly insensitive to their immediate surroundings is the tricyclic cytosine family. 1,3-Diaza-2-oxophenothiazine, tC, has a fluorescence quantum yield of approximately 0.2 both in single- and in double-strands irrespective of surrounding bases. Also the oxo-homologue of tC called tC^O (both commercially available), 1,3-diaza-2-oxophenoxazine, has a quantum yield of 0.2 in double-stranded systems. However, it is somewhat sensitive to surrounding bases in single-strands (quantum yields of 0.14–0.41). The high and stable quantum yields of these base analogues make them very bright, and, in combination with their good base analogue properties (leaves DNA structure and stability next to unperturbed), they are especially useful in fluorescence anisotropy and FRET measurements, areas where other fluorescent base analogues are less accurate. Also, in the same family of cytosine analogues, a FRET-acceptor base analogue, tC_nitro, has been developed. Together with tC^O as a FRET-donor this constitutes the first nucleic acid base analogue FRET-pair ever developed. The tC-family has, for example, been used in studies related to polymerase DNA-binding and DNA-polymerization mechanisms.

Natural non-canonical bases

In a cell, there are several non-canonical bases present: CpG islands in DNA (are often methylated), all eukaryotic mRNA (capped with a methyl-7-guanosine), and several bases of rRNAs (are methylated). Often, tRNAs are heavily modified postranscriptionally in order to improve their conformation or base pairing, in particular in/near the anticodon: inosine can base pair with C, U, and even with A, whereas thiouridine (with A) is more specific than uracil (with a purine). Other common tRNA base modifications are pseudouridine (which gives its name to the TΨC loop), dihydrouridine (which does not stack as it is not aromatic), queuosine, wyosine, and so forth. Nevertheless, these are all modifications to normal bases and are not placed by a polymerase. 

Base-pairing

Canonical bases may have either a carbonyl or an amine group on the carbons surrounding the nitrogen atom furthest away from the glycosidic bond, which allows them to base pair (Watson-Crick base pairing) via hydrogen bonds (amine with ketone, purine with pyrimidine). Adenine and 2-aminoadenine have one/two amine group(s), whereas thymine has two carbonyl groups, and cytosine and guanine are mixed amine and carbonyl (inverted in respect to each other).

Natural basepairs
A GC basepair: purine carbonyl/amine forms three intermolecular hydrogen bonds with pyrimidine amine/carbonyl	An AT basepair: purine amine/- forms two intermolecular hydrogen bonds with pyrimidine carbonyl/carbonyl

The precise reason why there are only four nucleotides is debated, but there are several unused possibilities. Furthermore, adenine is not the most stable choice for base pairing: in Cyanophage S-2L diaminopurine (DAP) is used instead of adenine (host evasion). Diaminopurine basepairs perfectly with thymine as it is identical to adenine but has an amine group at position 2 forming 3 intramolecular hydrogen bonds, eliminating the major difference between the two types of basepairs (Weak:A-T and Strong:C-G). This improved stability affects protein-binding interactions that rely on those differences. Other combination include,

• isoguanine and isocytosine, which have their amine and ketone inverted compared to standard guanine and cytosine, (not used probably as tautomers are problematic for base pairing, but isoC and isoG can be amplified correctly with PCR even in the presence of the 4 canonical bases)
• diaminopyrimidine and a xanthine, which bind like 2-aminoadenine and thymine but with inverted structures (not used as xanthine is a deamination product)

Unused basepair arrangements
A DAP-T base: purine amine/amine forms three intermolecular hydrogen bonds with pyrimidine ketone/ketone	An X-DAP base: purine ketone/ketone forms three intermolecular hydrogen bonds with pyrimidine amine/amine	A iG-iC base: purine amine/ketone forms three intermolecular hydrogen bonds with pyrimidine ketone/amine

However, correct DNA structure can form even when the bases are not paired via hydrogen bonding; that is, the bases pair thanks to hydrophobicity, as studies have shown using DNA isosteres (analogues with same number of atoms), such as the thymine analogue 2,4-difluorotoluene (F) or the adenine analogue 4-methylbenzimidazole (Z). An alternative hydrophobic pair could be isoquinoline, and the pyrrolo[2,3-b]pyridine

Other noteworthy basepairs:

• Several fluorescent bases have also been made, such as the 2-amino-6-(2-thienyl)purine and pyrrole-2-carbaldehyde base pair.
• Metal-coordinated bases, such as pairing between a pyridine-2,6-dicarboxylate (tridentate ligand) and a pyridine (monodentate ligand) through square planar coordination to a central copper ion.
• Universal bases may pair indiscriminately with any other base, but, in general, lower the melting temperature of the sequence considerably; examples include 2'-deoxyinosine (hypoxanthine deoxynucleotide) derivatives, nitroazole analogues, and hydrophobic aromatic non-hydrogen-bonding bases (strong stacking effects). These are used as proof of concept and, in general, are not utilised in degenerate primers (which are a mixture of primers).
• The numbers of possible base pairs is doubled when xDNA is considered. xDNA contains expanded bases, in which a benzene ring has been added, which may pair with canonical bases, resulting in four possible base-pairs (8 bases:xA-T,xT-A,xC-G,xG-C, 16 bases if the unused arrangements are used). Another form of benzene added bases is yDNA, in which the base is widened by the benzene.

Novel basepairs with special properties
A F-Z base: methylbenzimidazole does not form intermolecular hydrogen bonds with toluene F/F	An S-Pa base: purine thienyl/amine forms three intermolecular hydrogen bonds with pyrrole -/carbaldehyde	An xA-T base: same bonding as A-T

Metal base-pairs

In metal base-pairing, the Watson-Crick hydrogen bonds are replaced by the interaction between a metal ion with nucleosides acting as ligands. The possible geometries of the metal that would allow for duplex formation with two bidentate nucleosides around a central metal atom are: tetrahedral, dodecahedral, and square planar. Metal-complexing with DNA can occur by the formation of non-canonical base pairs from natural nucleobases with participation by metal ions and also by the exchanging the hydrogen atoms that are part of the Watson-Crick base pairing by metal ions. Introduction of metal ions into a DNA duplex has shown to have potential magnetic, conducting properties, as well as increased stability.

Metal complexing has been shown to occur between natural nucleobases. A well-documented example is the formation of T-Hg-T, which involves two deprotonated thymine nucleobases that are brought together by Hg²⁺ and forms a connected metal-base pair. This motif does not accommodate stacked Hg²⁺ in a duplex due to an intrastrand hairpin formation process that is favored over duplex formation. Two thymines across from each other in a duplex do not form a Watson-Crick base pair in a duplex; this is an example where a Watson-Crick basepair mismatch is stabilized by the formation of the metal-base pair. Another example of a metal complexing to natural nucleobases is the formation of A-Zn-T and G-Zn-C at high pH; Co⁺² and Ni⁺² also form these complexes. These are Watson-Crick base pairs where the divalent cation in coordinated to the nucleobases. The exact binding is debated.

A large variety of artificial nucleobases have been developed for use as metal base pairs. These modified nucleobases exhibit tunable electronic properties, sizes, and binding affinities that can be optimized for a specific metal. For, example a nucleoside modified with a pyridine-2,6-dicarboxylate has shown to bind tightly to Cu²⁺, whereas other divalent ions are only loosely bound. The tridentate character contributes to this selectivity. The fourth coordination site on the copper is saturated by an oppositely arranged pyridine nucleobase. The asymmetric metal base pairing system is orthogonal to the Watson-Crick base pairs. Another example of an artificial nucleobase is that with hydroxypyridone nucleobases, which are able to bind Cu²⁺ inside the DNA duplex. Five consecutive copper-hydroxypyridone base pairs were incorporated into a double strand, which were flanked by only one natural nucleobase on both ends. EPR data showed that the distance between copper centers was estimated to be 3.7 ± 0.1 Å, while a natural B-type DNA duplex is only slightly larger (3.4 Å). The appeal for stacking metal ions inside a DNA duplex is the hope to obtain nanoscopic self-assembling metal wires, though this has not been realized yet.

Unnatural base pair (UBP)

Main article: Unnatural base pair

An unnatural base pair (UBP) is a designed subunit (or nucleobase) of DNA which is created in a laboratory and does not occur in nature. In 2012, a group of American scientists led by Floyd Romesberg, a chemical biologist at the Scripps Research Institute in San Diego, California, published that his team designed an unnatural base pair (UBP). The two new artificial nucleotides or Unnatural Base Pair (UBP) were named d5SICS and dNaM. More technically, these artificial nucleotides bearing hydrophobic nucleobases, feature two fused aromatic rings that form a (d5SICS–dNaM) complex or base pair in DNA. In 2014 the same team from the Scripps Research Institute reported that they synthesized a stretch of circular DNA known as a plasmid containing natural T-A and C-G base pairs along with the best-performing UBP Romesberg's laboratory had designed, and inserted it into cells of the common bacterium E. coli that successfully replicated the unnatural base pairs through multiple generations. This is the first known example of a living organism passing along an expanded genetic code to subsequent generations. This was in part achieved by the addition of a supportive algal gene that expresses a nucleotide triphosphate transporter which efficiently imports the triphosphates of both d5SICSTP and dNaMTP into E. coli bacteria. Then, the natural bacterial replication pathways use them to accurately replicate the plasmid containing d5SICS–dNaM.

The successful incorporation of a third base pair is a significant breakthrough toward the goal of greatly expanding the number of amino acids which can be encoded by DNA, from the existing 20 amino acids to a theoretically possible 172, thereby expanding the potential for living organisms to produce novel proteins. Earlier, the artificial strings of DNA did not encode for anything, but scientists speculated they could be designed to manufacture new proteins which could have industrial or pharmaceutical uses. Transcription of DNA containing unnatural base pair and translation of corresponding mRNA were actually achieved recently. In November 2017, the same team at the Scripps Research Institute that first introduced two extra nucleobases into bacterial DNA, reported having constructed a semi-synthetic E. coli bacteria able to make proteins using such DNA. Its DNA contained six different nucleobases: four canonical and two artificially added, dNaM and dTPT3 (these two form a pair). Also, this bacteria had two corresponding additional RNA bases included in two new codons, additional tRNAs recognizing these new codons (these tRNAs also contained two new RNA bases within their anticodons) and additional amino acids, making the bacteria able to synthesize "unnatural" proteins.

Another demonstration of UBPs were achieved by Ichiro Hirao's group at RIKEN institute in Japan. In 2002, they developed an unnatural base pair between 2-amino-8-(2-thienyl)purine (s) and pyridine-2-one (y) that functions in vitro in transcription and translation, for the site-specific incorporation of non-standard amino acids into proteins. In 2006, they created 7-(2-thienyl)imidazo[4,5-b]pyridine (Ds) and pyrrole-2-carbaldehyde (Pa) as a third base pair for replication and transcription. Afterward, Ds and 4-[3-(6-aminohexanamido)-1-propynyl]-2-nitropyrrole (Px) was discovered as a high fidelity pair in PCR amplification. In 2013, they applied the Ds-Px pair to DNA aptamer generation by in vitro selection (SELEX) and demonstrated the genetic alphabet expansion significantly augment DNA aptamer affinities to target proteins.

Orthogonal system

The possibility has been proposed and studied, both theoretically and experimentally, of implementing an orthogonal system inside cells independent of the cellular genetic material in order to make a completely safe system, with the possible increase in encoding potentials. Several groups have focused on different aspects:

• novel backbones and base pairs as discussed above
• XNA (Xeno Nucleic Acid) artificial replication/transcription polymerases starting generally from T7 RNA polymerase
• ribosomes (16S sequences with altered anti Shine-Dalgarno sequence allowing the translation of only orthogonal mRNA with a matching altered Shine-Dalgarno sequence)
• novel tRNA encoding non-natural aminoacids. See Expanded genetic code

Proton-exchange membrane fuel cell

From Wikipedia, the free encyclopedia

https://en.wikipedia.org/wiki/Proton-exchange_membrane_fuel_cell 

 

Diagram of a PEM fuel cell

Proton-exchange membrane fuel cells (PEMFC), also known as polymer electrolyte membrane (PEM) fuel cells, are a type of fuel cell being developed mainly for transport applications, as well as for stationary fuel-cell applications and portable fuel-cell applications. Their distinguishing features include lower temperature/pressure ranges (50 to 100 °C) and a special proton-conducting polymer electrolyte membrane. PEMFCs generate electricity and operate on the opposite principle to PEM electrolysis, which consumes electricity. They are a leading candidate to replace the aging alkaline fuel-cell technology, which was used in the Space Shuttle.

Science

PEMFCs are built out of membrane electrode assemblies (MEA) which include the electrodes, electrolyte, catalyst, and gas diffusion layers. An ink of catalyst, carbon, and electrode are sprayed or painted onto the solid electrolyte and carbon paper is hot pressed on either side to protect the inside of the cell and also act as electrodes. The pivotal part of the cell is the triple phase boundary (TPB) where the electrolyte, catalyst, and reactants mix and thus where the cell reactions actually occur. Importantly, the membrane must not be electrically conductive so the half reactions do not mix. Operating temperatures above 100 °C are desired so the water byproduct becomes steam and water management becomes less critical in cell design.

Reactions

Further information: Fuel cell

A proton exchange membrane fuel cell transforms the chemical energy liberated during the electrochemical reaction of hydrogen and oxygen to electrical energy, as opposed to the direct combustion of hydrogen and oxygen gases to produce thermal energy.

A stream of hydrogen is delivered to the anode side of the MEA. At the anode side it is catalytically split into protons and electrons. This oxidation half-cell reaction or hydrogen oxidation reaction (HOR) is represented by:

At the anode:

${\displaystyle \mathrm {H} _{2}\rightarrow \mathrm {2H} ^{+}+\mathrm {2e} ^{-}}$

${\displaystyle E^{\circ }=0\,\mathrm {V} \,\,\,{\frac {\mathrm {d} E^{\circ }}{\mathrm {d} T}}=0\,\mathrm {mV\,K^{-1}} }$

${\displaystyle \left(1\right)}$

The newly formed protons permeate through the polymer electrolyte membrane to the cathode side. The electrons travel along an external load circuit to the cathode side of the MEA, thus creating the current output of the fuel cell. Meanwhile, a stream of oxygen is delivered to the cathode side of the MEA. At the cathode side oxygen molecules react with the protons permeating through the polymer electrolyte membrane and the electrons arriving through the external circuit to form water molecules. This reduction half-cell reaction or oxygen reduction reaction (ORR) is represented by:

At the cathode:

${\displaystyle {\frac {1}{2}}\mathrm {O} _{2}+{2\mathrm {H} }^{+}+\mathrm {2e} ^{-}\rightarrow \mathrm {H} _{2}\mathrm {O} }$

${\displaystyle E^{\circ }=1.2291\,\mathrm {V} \,\,\,{\frac {\mathrm {d} E^{\circ }}{\mathrm {d} T}}=-0.8456\,\mathrm {mV\,K^{-1}} }$

${\displaystyle \left(2\right)}$

Overall reaction:

${\displaystyle \mathrm {H} _{2}+{\frac {1}{2}}\mathrm {O} _{2}\rightarrow \mathrm {H} _{2}\mathrm {O} }$

${\displaystyle E^{\circ }=1.2291\,\mathrm {V} \,\,\,{\frac {\mathrm {d} E^{\circ }}{\mathrm {d} T}}=-0.8456\,\mathrm {mV\,K^{-1}} }$

${\displaystyle \left(3\right)}$

The reversible reaction is expressed in the equation and shows the reincorporation of the hydrogen protons and electrons together with the oxygen molecule and the formation of one water molecule. The potentials in each case are given with respect to the standard hydrogen electrode.

Polymer electrolyte membrane

Main article: Polymer electrolyte membrane

SEM micrograph of a PEMFC MEA cross-section with a non-precious metal catalyst cathode and Pt/C anode. False colors applied for clarity.

 

MEA fabrication methods for PEMFC

To function, the membrane must conduct hydrogen ions (protons) but not electrons as this would in effect "short circuit" the fuel cell. The membrane must also not allow either gas to pass to the other side of the cell, a problem known as gas crossover. Finally, the membrane must be resistant to the reducing environment at the cathode as well as the harsh oxidative environment at the anode.

Splitting of the hydrogen molecule is relatively easy by using a platinum catalyst. Unfortunately however, splitting the oxygen molecule is more difficult, and this causes significant electric losses. An appropriate catalyst material for this process has not been discovered, and platinum is the best option.

Strengths

The PEMFC is a prime candidate for vehicle and other mobile applications of all sizes down to mobile phones, because of its compactness.

Weaknesses

Fuel Cells based on PEM still have many issues:

1. Water management

Water management is crucial to performance: if water is evaporated too slowly, it will flood the membrane and the accumulation of water inside of field flow plate will impede the flow of oxygen into the fuel cell, but if water evaporates too fast, the membrane will dry and the resistance across it increases. Both cases will cause damage to stability and power output. Water management is a very difficult subject in PEM systems, primarily because water in the membrane is attracted toward the cathode of the cell through polarization.

A wide variety of solutions for managing the water exist including integration of an electroosmotic pump.

Another innovative method to resolve the water recirculation problem is the 3D fine mesh flow field design used in the Toyota Mirai, 2014. Conventional design of FC stack recirculates water from the air outlet to the air inlet through a humidifier with a straight channel and porous metal flow fields. The flow field is a structure made up of a rib and channels. However, the rib partially covers the gas diffusion layer (GDL) and the resultant gas-transport distance is longer than the inter-channel distance. Furthermore, the contact pressure between the GDL and the rib also compresses the GDL, making its thickness non-uniform across the rib and channel. The large width and non-uniform thickness of the rib will increase potential for water vapor to accumulate and the oxygen will be compromised. As a result, oxygen will be impeded to diffuse into catalyst layer, leading to nonuniform power generation in the FC.

This new design enabled the first FC stack functions without a humidifying system meanwhile overcoming water recirculation issues and achieving high power output stability. The 3D micro lattice allows more pathways for gas flow; therefore, it promotes airflow toward membrane electrode and gas diffusion layer assembly (MEGA) and promotes O2 diffusion to the catalyst layer. Unlike conventional flow fields, the 3D micro-lattices in the complex field, which act as baffles and induce frequent micro-scale interfacial flux between the GDL and flow-fields^[53]. Due to this repeating micro-scale convective flow, oxygen transport to catalyst layer (CL) and liquid water removal from GDL is significantly enhanced. The generated water is quickly drawn out through the flow field, preventing accumulation within the pores. As a result, the power generation from this flow field is uniform across the cross-section and self-humidification is enabled.

2. Vulnerability of the Catalyst

The platinum catalyst on the membrane is easily poisoned by carbon monoxide, which is often present in product gases formed by methane reforming (no more than one part per million is usually acceptable). This generally necessitates the use of the water gas shift reaction to eliminate CO from product gases and form more hydrogen. Additionally, the membrane is sensitive to the presences of metal ions, which may impair proton conduction mechanisms and can be introduced by corrosion of metallic bipolar plates, metallic components in the fuel cell system or from contaminants in the fuel/oxidant.

PEM systems that use reformed methanol were proposed, as in Daimler Chrysler Necar 5; reforming methanol, i.e. making it react to obtain hydrogen, is however a very complicated process, that also requires purification from the carbon monoxide the reaction produces. A platinum-ruthenium catalyst is necessary as some carbon monoxide will unavoidably reach the membrane. The level should not exceed 10 parts per million. Furthermore, the start-up times of such a reformer reactor are of about half an hour. Alternatively, methanol, and some other biofuels can be fed to a PEM fuel cell directly without being reformed, thus making a direct methanol fuel cell (DMFC). These devices operate with limited success.

3. Limitation of Operating Temperature

The most commonly used membrane is Nafion by Chemours, which relies on liquid water humidification of the membrane to transport protons. This implies that it is not feasible to use temperatures above 80 to 90 °C, since the membrane would dry. Other, more recent membrane types, based on polybenzimidazole (PBI) or phosphoric acid, can reach up to 220 °C without using any water management (see also High Temperature Proton Exchange Membrane fuel cell, HT-PEMFC): higher temperature allow for better efficiencies, power densities, ease of cooling (because of larger allowable temperature differences), reduced sensitivity to carbon monoxide poisoning and better controllability (because of absence of water management issues in the membrane); however, these recent types are not as common. PBI can be doped with phosphoric or sulfuric acid and the conductivity scales with amount of doping and temperature. At high temperatures, it is difficult to keep Nafion hydrated, but this acid doped material does not use water as a medium for proton conduction. It also exhibits better mechanical properties, higher strength, than Nafion and is cheaper. However, acid leaching is a considerable issue and processing, mixing with catalyst to form ink, has proved tricky. Aromatic polymers, such as PEEK, are far cheaper than Teflon (PTFE and backbone of Nafion) and their polar character leads to hydration that is less temperature dependent than Nafion. However, PEEK is far less ionically conductive than Nafion and thus is a less favorable electrolyte choice. Recently, protic ionic liquids and protic organic ionic plastic crystals have been shown as promising alternative electrolyte materials for high temperature (100–200 °C) PEMFCs.

Electrodes

An electrode typically consists of carbon support, Pt particles, Nafion ionomer, and/or Teflon binder. The carbon support functions as an electrical conductor; the Pt particles are reaction sites; the ionomer provides paths for proton conduction, and the Teflon binder increases the hydrophobicity of the electrode to minimize potential flooding. In order to enable the electrochemical reactions at the electrodes, protons, electrons and the reactant gases (hydrogen or oxygen) must gain access to the surface of the catalyst in the electrodes, while the product water, which can be in either liquid or gaseous phase, or both phases, must be able to permeate from the catalyst to the gas outlet. These properties are typically realized by porous composites of polymer electrolyte binder (ionomer) and catalyst nanoparticles supported on carbon particles. Typically platinum is used as the catalyst for the electrochemical reactions at the anode and cathode, while nanoparticles realize high surface to weight ratios (as further described below) reducing the amount of the costly platinum. The polymer electrolyte binder provides the ionic conductivity, while the carbon support of the catalyst improves the electric conductivity and enables low platinum metal loading. The electric conductivity in the composite electrodes is typically more than 40 times higher as the proton conductivity.

Gas diffusion layer

The GDL electrically connects the catalyst and current collector. It must be porous, electrically conductive, and thin. The reactants must be able to reach the catalyst, but conductivity and porosity can act as opposing forces. Optimally, the GDL should be composed of about one third Nafion or 15% PTFE. The carbon particles used in the GDL can be larger than those employed in the catalyst because surface area is not the most important variable in this layer. GDL should be around 15–35 µm thick to balance needed porosity with mechanical strength. Often, an intermediate porous layer is added between the GDL and catalyst layer to ease the transitions between the large pores in the GDL and small porosity in the catalyst layer. Since a primary function of the GDL is to help remove water, a product, flooding can occur when water effectively blocks the GDL. This limits the reactants ability to access the catalyst and significantly decreases performance. Teflon can be coated onto the GDL to limit the possibility of flooding. Several microscopic variables are analyzed in the GDLS such as: porosity, tortuosity and permeability. These variables have incidence over the behavior of the fuel cells.

Efficiency

The maximal theoretical efficiency applying the Gibbs free energy equation ΔG = −237.13 kJ/mol and using the heating value of Hydrogen (ΔH = −285.84 kJ/mol) is 83% at 298 K.

${\displaystyle \eta ={\frac {\Delta G}{\Delta H}}=1-{\frac {T\Delta S}{\Delta H}}}$

The practical efficiency of a PEMs is in the range of 50–60% . Main factors that create losses are:

• Activation losses
• Ohmic losses
• Mass transport losses

Metal-organic frameworks

Metal-organic frameworks (MOFs) are a relatively new class of porous, highly crystalline materials that consist of metal nodes connected by organic linkers. Due to the simplicity of manipulating or substituting the metal centers and ligands, there are a virtually limitless number of possible combinations, which is attractive from a design standpoint. MOFs exhibit many unique properties due to their tunable pore sizes, thermal stability, high volume capacities, large surface areas, and desirable electrochemical characteristics. Among their many diverse uses, MOFs are promising candidates for clean energy applications such as hydrogen storage, gas separations, supercapacitors, Li-ion batteries, solar cells, and fuel cells. Within the field of fuel cell research, MOFs are being studied as potential electrolyte materials and electrode catalysts that could someday replace traditional polymer membranes and Pt catalysts, respectively.

As electrolyte materials, the inclusion of MOFs seems at first counter-intuitive. Fuel cell membranes generally have low porosity to prevent fuel crossover and loss of voltage between the anode and cathode. Additionally, membranes tend to have low crystallinity because the transport of ions is more favorable in disordered materials. On the other hand, pores can be filled with additional ion carriers that ultimately enhance the ionic conductivity of the system and high crystallinity makes the design process less complex.

The general requirements of a good electrolyte for PEMFCs are: high proton conductivity (>10⁻² S/cm for practical applications) to enable proton transport between electrodes, good chemical and thermal stability under fuel cell operating conditions (environmental humidity, variable temperatures, resistance to poisonous species, etc.), low cost, ability to be processed into thin-films, and overall compatibility with other cell components. While polymeric materials are currently the preferred choice of proton-conducting membrane, they require humidification for adequate performance and can sometimes physically degrade due to hydrations effects, thereby causing losses of efficiency. As mentioned, Nafion is also limited by a dehydration temperature of < 100 °C, which can lead to slower reaction kinetics, poor cost efficiency, and CO poisoning of Pt electrode catalysts. Conversely, MOFs have shown encouraging proton conductivities in both low and high temperature regimes as well as over a wide range of humidity conditions. Below 100 °C and under hydration, the presence of hydrogen bonding and solvent water molecules aid in proton transport, whereas anhydrous conditions are suitable for temperatures above 100 °C. MOFs also have the distinct advantage of exhibiting proton conductivity by the framework itself in addition to the inclusion of charge carries (i.e., water, acids, etc.) into their pores.

A low temperature example is work by Kitagawa, et al. who used a two-dimensional oxalate-bridged anionic layer framework as the host and introduced ammonium cations and adipic acid molecules into the pores to increase proton concentration. The result was one of the first instances of a MOF showing “superprotonic” conductivity (8 × 10⁻³ S/cm) at 25 °C and 98% relative humidity (RH). They later found that increasing the hydrophilic nature of the cations introduced into the pores could enhance proton conductivity even more. In this low temperature regime that is dependent on degree of hydration, it has also been shown that proton conductivity is heavily dependent on humidity levels.

A high temperature anhydrous example is PCMOF2, which consists of sodium ions coordinated to a trisulfonated benzene derivative. To improve performance and allow for higher operating temperatures, water can be replaced as the proton carrier by less volatile imidazole or triazole molecules within the pores. The maximum temperature achieved was 150 °C with an optimum conductivity of 5 × 10⁻⁴ S/cm, which is lower than other current electrolyte membranes. However, this model holds promise for its temperature regime, anhydrous conditions, and ability to control the quantity of guest molecules within the pores, all of which allowed for the tunability of proton conductivity. Additionally, the triazole-loaded PCMOF2 was incorporated into a H₂/air membrane-electrode assembly and achieved an open circuit voltage of 1.18 V at 100 °C that was stable for 72 hours and managed to remain gas tight throughout testing. This was the first instance that proved MOFs could actually be implemented into functioning fuel cells, and the moderate potential difference showed that fuel crossover due to porosity was not an issue.

To date, the highest proton conductivity achieved for a MOF electrolyte is 4.2 × 10⁻² S/cm at 25 °C under humid conditions (98% RH), which is competitive with Nafion. Some recent experiments have even successfully produced thin-film MOF membranes instead of the traditional bulk samples or single crystals, which is crucial for their industrial applicability. Once MOFs are able to consistently achieve sufficient conductivity levels, mechanical strength, water stability, and simple processing, they have the potential to play an important role in PEMFCs in the near future.

MOFs have also been targeted as potential replacements of platinum group metal (PGM) materials for electrode catalysts, although this research is still in the early stages of development. In PEMFCs, the oxygen reduction reaction (ORR) at the Pt cathode is significantly slower than the fuel oxidation reaction at the anode, and thus non-PGM and metal-free catalysts are being investigated as alternatives. The high volumetric density, large pore surface areas, and openness of metal-ion sites in MOFs make them ideal candidates for catalyst precursors. Despite promising catalytic abilities, the durability of these proposed MOF-based catalysts is currently less than desirable and the ORR mechanism in this context is still not completely understood.

Catalyst research

Much of the current research on catalysts for PEM fuel cells can be classified as having one of the following main objectives:

1. to obtain higher catalytic activity than the standard carbon-supported platinum particle catalysts used in current PEM fuel cells
2. to reduce the poisoning of PEM fuel cell catalysts by impurity gases
3. to reduce the cost of the fuel cell due to use of platinum-based catalysts
4. to enhance the ORR activity of platinum group metal-free electrocatalysts

Examples of these approaches are given in the following sections.

Increasing catalytic activity

As mentioned above, platinum is by far the most effective element used for PEM fuel cell catalysts, and nearly all current PEM fuel cells use platinum particles on porous carbon supports to catalyze both hydrogen oxidation and oxygen reduction. However, due to their high cost, current Pt/C catalysts are not feasible for commercialization. The U.S. Department of Energy estimates that platinum-based catalysts will need to use roughly four times less platinum than is used in current PEM fuel cell designs in order to represent a realistic alternative to internal combustion engines. Consequently, one main goal of catalyst design for PEM fuel cells is to increase the catalytic activity of platinum by a factor of four so that only one-fourth as much of the precious metal is necessary to achieve similar performance.

One method of increasing the performance of platinum catalysts is to optimize the size and shape of the platinum particles. Decreasing the particles’ size alone increases the total surface area of catalyst available to participate in reactions per volume of platinum used, but recent studies have demonstrated additional ways to make further improvements to catalytic performance. For example, one study reports that high-index facets of platinum nanoparticles (that is Miller indexes with large integers, such as Pt (730)) provide a greater density of reactive sites for oxygen reduction than typical platinum nanoparticles.

Since the most common and effective catalyst, platinum, is extremely expensive, alternative processing is necessary to maximize surface area and minimize loading. Deposition of nanosized Pt particles onto carbon powder (Pt/C) provides a large Pt surface area while the carbon allows for electrical connection between the catalyst and the rest of the cell. Platinum is so effective because it has high activity and bonds to the hydrogen just strongly enough to facilitate electron transfer but not inhibit the hydrogen from continuing to move around the cell. However, platinum is less active in the cathode oxygen reduction reaction. This necessitates the use of more platinum, increasing the cell's expense and thus feasibility. Many potential catalyst choices are ruled out because of the extreme acidity of the cell.

The most effective ways of achieving the nanoscale Pt on carbon powder, which is currently the best option, are through vacuum deposition, sputtering, and electrodeposition. The platinum particles are deposited onto carbon paper that is permeated with PTFE. However, there is an optimal thinness to this catalyst layer, which limits the lower cost limit. Below 4 nm, Pt will form islands on the paper, limiting its activity. Above this thickness, the Pt will coat the carbon and be an effective catalyst. To further complicate things, Nafion cannot be infiltrated beyond 10 um, so using more Pt than this is an unnecessary expense. Thus the amount and shape of the catalyst is limited by the constraints of other materials.

A second method of increasing the catalytic activity of platinum is to alloy it with other metals. For example, it was recently shown that the Pt₃Ni(111) surface has a higher oxygen reduction activity than pure Pt(111) by a factor of ten. The authors attribute this dramatic performance increase to modifications to the electronic structure of the surface, reducing its tendency to bond to oxygen-containing ionic species present in PEM fuel cells and hence increasing the number of available sites for oxygen adsorption and reduction.

Further efficiencies can be realized using an Ultrasonic nozzle to apply the platinum catalyst to the electrolyte layer or to carbon paper under atmospheric conditions resulting in high efficiency spray. Studies have shown that due to the uniform size of the droplets created by this type of spray, due to the high transfer efficiency of the technology, due to the non-clogging nature of the nozzle and finally due to the fact that the ultrasonic energy de-agglomerates the suspension just before atomization, fuel cells MEA's manufactured this way have a greater homogeneity in the final MEA, and the gas flow through the cell is more uniform, maximizing the efficiency of the platinum in the MEA. Recent studies using inkjet printing to deposit the catalyst over the membrane have also shown high catalyst utilization due to the reduced thickness of the deposited catalyst layers.

Very recently, a new class of ORR electrocatalysts have been introduced in the case of Pt-M (M-Fe and Co) systems with an ordered intermetallic core encapsulated within a Pt-rich shell. These intermetallic core-shell (IMCS) nanocatalysts were found to exhibit an enhanced activity and most importantly, an extended durability compared to many previous designs. While the observed enhancement in the activities is ascribed to a strained lattice, the authors report that their findings on the degradation kinetics establish that the extended catalytic durability is attributable to a sustained atomic order.

Reducing poisoning

The other popular approach to improving catalyst performance is to reduce its sensitivity to impurities in the fuel source, especially carbon monoxide (CO). Presently, pure hydrogen gas is becoming economical to mass-produce by electrolysis. However, at the moment hydrogen gas is produced by steam reforming light hydrocarbons, a process which produces a mixture of gases that also contains CO (1–3%), CO₂ (19–25%), and N₂ (25%). Even tens of parts per million of CO can poison a pure platinum catalyst, so increasing platinum's resistance to CO is an active area of research.

For example, one study reported that cube-shaped platinum nanoparticles with (100) facets displayed a fourfold increase in oxygen reduction activity compared to randomly faceted platinum nanoparticles of similar size. The authors concluded that the (111) facets of the randomly shaped nanoparticles bonded more strongly to sulfate ions than the (100) facets, reducing the number of catalytic sites open to oxygen molecules. The nanocubes they synthesized, in contrast, had almost exclusively (100) facets, which are known to interact with sulfate more weakly. As a result, a greater fraction of the surface area of those particles was available for the reduction of oxygen, boosting the catalyst's oxygen reduction activity.

In addition, researchers have been investigating ways of reducing the CO content of hydrogen fuel before it enters a fuel cell as a possible way to avoid poisoning the catalysts. One recent study revealed that ruthenium-platinum core–shell nanoparticles are particularly effective at oxidizing CO to form CO₂, a much less harmful fuel contaminant. The mechanism that produces this effect is conceptually similar to that described for Pt₃Ni above: the ruthenium core of the particle alters the electronic structure of the platinum surface, rendering it better able to catalyze the oxidation of CO.

Lowering cost

The challenge for the viability of PEM fuel cells today still remains in their cost and stability. The high cost can in large part be attributed to the use of the precious metal of platinum in the catalyst layer of PEM cells. The electrocatalyst currently accounts for nearly half of the fuel cell stack cost. Although the Pt loading of PEM fuel cells has been reduced by two orders of magnitude over the past decade, further reduction is necessary to make the technology economically viable for commercialization. Whereas some research efforts aim to address this issue by improving the electrocatalytic activity of Pt-based catalysts, an alternative is to eliminate the use of Pt altogether by developing a non-platinum-group-metal (non-PGM) cathode catalyst whose performance rivals that of Pt-based technologies. The U.S. Department of Energy has been setting milestones for the development of fuel cells, targeting a durability of 5000 hours and a non-PGM catalyst ORR volumetric activity of 300 A cm⁻³.

Promising alternatives to Pt-based catalysts are Metal/Nitrogen/ Carbon-catalysts (M/N/C-catalysts). To achieve high power density, or output of power over surface area of the cell, a volumetric activity of at least 1/10 that of Pt-based catalysts must be met, along with good mass transport properties. While M/N/C-catalysts still demonstrate poorer volumetric activities than Pt-based catalysts, the reduced costs of such catalysts allows for greater loading to compensate. However, increasing the loading of M/N/C-catalysts also renders the catalytic layer thicker, impairing its mass transport properties. In other words, H₂, O₂, protons, and electrons have greater difficulty in migrating through the catalytic layer, decreasing the voltage output of the cell. While high microporosity of the M/N/C catalytic network results in high volumetric activity, improved mass transport properties are instead associated to macroporosity of the network. These M/N/C materials are synthesized using high temperature pyrolysis and other high temperature treatments of precursors containing the metal, nitrogen, and carbon.

Recently, researchers have developed a Fe/N/C catalyst derived from iron (II) acetate (FeAc), phenanthroline (Phen), and a metal-organic-framework (MOF) host. The MOF is a Zn(II) zeolitic imidazolate framework (ZIF) called ZIF-8, which demonstrates a high microporous surface area and high nitrogen content conducive to ORR activity. The power density of the FeAc/Phen/ZIF-8-catalyst was found to be 0.75 W cm⁻² at 0.6 V. This value is a significant improvement over the maximal 0.37 W cm⁻² power density of previous M/N/C-catalysts and is much closer to matching the typical value of 1.0–1.2 W cm⁻² for Pt-based catalysts with a Pt loading of 0.3 mg cm⁻². The catalyst also demonstrated a volumetric activity of 230 A·cm⁻³, the highest value for non-PGM catalysts to date, approaching the U.S. Department of Energy milestone.

While the power density achieved by the novel FeAc/Phen/ZIF-8-catalyst is promising, its durability remains inadequate for commercial application. It is reported that the best durability exhibited by this catalyst still had a 15% drop in current density over 100 hours in H₂/air. Hence while the Fe-based non-PGM catalysts rival Pt-based catalysts in their electrocatalytic activity, there is still much work to be done in understanding their degradation mechanisms and improving their durability.

Applications

The major application of PEM fuel cells focuses on transportation primarily because of their potential impact on the environment, e.g. the control of emission of the green house gases (GHG). Other applications include distributed/stationary and portable power generation. Most major motor companies work solely on PEM fuel cells due to their high power density and excellent dynamic characteristics as compared with other types of fuel cells. Due to their light weight, PEMFCs are most suited for transportation applications. PEMFCs for buses, which use compressed hydrogen for fuel, can operate at up to 40% efficiency. Generally PEMFCs are implemented on buses over smaller cars because of the available volume to house the system and store the fuel. Technical issues for transportation involve incorporation of PEMs into current vehicle technology and updating energy systems. Full fuel cell vehicles are not advantageous if hydrogen is sourced from fossil fuels; however, they become beneficial when implemented as hybrids. There is potential for PEMFCs to be used for stationary power generation, where they provide 5 kW at 30% efficiency; however, they run into competition with other types of fuel cells, mainly SOFCs and MCFCs. Whereas PEMFCs generally require high purity hydrogen for operation, other fuel cell types can run on methane and are thus more flexible systems. Therefore, PEMFCs are best for small scale systems until economically scalable pure hydrogen is available. Furthermore, PEMFCs have the possibility of replacing batteries for portable electronics, though integration of the hydrogen supply is a technical challenge particularly without a convenient location to store it within the device.

History

Before the invention of PEM fuel cells, existing fuel cell types such as solid-oxide fuel cells were only applied in extreme conditions. Such fuel cells also required very expensive materials and could only be used for stationary applications due to their size. These issues were addressed by the PEM fuel cell. The PEM fuel cell was invented in the early 1960s by Willard Thomas Grubb and Leonard Niedrach of General Electric. Initially, sulfonated polystyrene membranes were used for electrolytes, but they were replaced in 1966 by Nafion ionomer, which proved to be superior in performance and durability to sulfonated polystyrene.

PEM fuel cells were used in the NASA Gemini series of spacecraft, but they were replaced by Alkaline fuel cells in the Apollo program and in the Space shuttle. General Electric continued working on PEM cells and in the mid-1970s developed PEM water electrolysis technology for undersea life support, leading to the US Navy Oxygen Generating Plant. The British Royal Navy adopted this technology in early 1980s for their submarine fleet. In the late 1980s and early 1990s, Los Alamos National Lab and Texas A&M University experimented with ways to reduce the amount of platinum required for PEM cells.

Parallel with Pratt and Whitney Aircraft, General Electric developed the first proton exchange membrane fuel cells (PEMFCs) for the Gemini space missions in the early 1960s. The first mission to use PEMFCs was Gemini V. However, the Apollo space missions and subsequent Apollo-Soyuz, Skylab and Space Shuttle missions used fuel cells based on Bacon's design, developed by Pratt and Whitney Aircraft.

Extremely expensive materials were used and the fuel cells required very pure hydrogen and oxygen. Early fuel cells tended to require inconveniently high operating temperatures that were a problem in many applications. However, fuel cells were seen to be desirable due to the large amounts of fuel available (hydrogen and oxygen).

Despite their success in space programs, fuel cell systems were limited to space missions and other special applications, where high cost could be tolerated. It was not until the late 1980s and early 1990s that fuel cells became a real option for wider application base. Several pivotal innovations, such as low platinum catalyst loading and thin film electrodes, drove the cost of fuel cells down, making development of PEMFC systems more realistic. However, there is significant debate as to whether hydrogen fuel cells will be a realistic technology for use in automobiles or other vehicles. (See hydrogen economy.) A large part of PEMFC production is for the Toyota Mirai. The US Department of Energy estimates a 2016 price at $53/kW if 500,000 units per year were made.