MSc Chemistry
Molecular Sciences
Literature Thesis
Biologically relevant alkyl-phosphates
A review of the applications, synthesis and market of organophosphates
by
Marit Beerse
11249358
December 2020
6 ECs
Period 2 – October 2020 - January 2021
Supervisor/Examiner:
Examiner:
Contents
Abstract ... 3 Abbreviations ... 4 1 Introduction ... 6 2 Mononucleotides ... 8 2.1 Adenosine-5’-monophosphate ... 82.2 Inosinic acid and guanosine-5’-monophosphate ... 10
2.3 Synthesis of monophosphates ... 12
3 Dinucleotides ... 14
3.1 Synthesis of NAD+ and NADP+ ... 15
4 Phospholipids ... 16 4.1 Liposomes ... 17 4.2 Phosphatidylserine ... 17 4.3 Phosphatidylcholine ... 18 4.4 Synthesis of phospholipids ... 18 4.5 Glycerophosphate ... 19 5 Terpenes ... 20 6 Vitamins ... 21 6.1 Vitamin B6 ... 21 6.2 Vitamin B12 ... 22 7 Discussion ... 25 8 Conclusion ... 28
Acknowledgements ... Fout! Bladwijzer niet gedefinieerd. References ... 30
Abstract
Phosphate plays many important roles globally, as a building block for cells, DNA, and vitamins. Additionally, it is used a lot in fertiliser and for the manufacture of pharmaceuticals. However, inadequate control of phosphorus pollution causes eutrophication and the current fossil phosphorus sources will become depleted. Struvite from wastewater is a good contestant for a renewable source of phosphates, creating a circular phosphate economy. Most artificial phosphate-containing products, including alkyl-phosphates, are currently made using fossil white phosphate in a very energy demanding process. Transforming this synthesis to struvite as a starting material could solve both eutrophication and depletion of mines by creating a circular phosphate economy. This review focusses on a small part of phosphates’ possibilities in a circular economy: biologically relevant alkyl-phosphates, for example, phospholipids, adenosine-5’-monophospate and vitamin B12. The biologically relevant phosphate esters are the organophosphates with applications that differ from the regular ones inside the human body. Examples of those applications are the use of guanosine-5’-monophosphate for enhancing mushroom flavour or using phospholipids with antibodies and phosphatidylserine as HIV-1 removal agents. This review will discuss the regular application, the different applications, the possible synthesis, its greenness and struvite-including possibilities, and lastly, the price and commercially available quantities of the organophosphates. Vitamin B12 appears to be the best alkyl-phosphate, based on struvite inclusion, greenness of the synthesis, importance of the application and market-indication.
Abbreviations
ADP/AMP/ATP Adenosine-5’-di/mono/tri-phosphate Adenylic acid Adenosine-5’-monophosphate
BV Benzylviologen
CDP/CMP/CTP Cytosine-5’-di/mono/tri-phosphate
CDT 1,1’-Carbonyl-di-(1,2,4-triazole)
Cytidylic acid Cytosine-5’-monophosphate
D Diaphorase
DCC Dicyclohexylcarbodiimide
DCM Dichloromethane
DIPEA N,N-Diisopropylethylamine
DMAP 4-dimethylaminepyridine
DMAP2 Dimethylallyl monophosphate
DMAPP Dimethylallyl diphosphate
DMF Dimethylformamide
DMSO Dimethyl sulfoxide
DNA Deoxyribonucleic acid
GDP/GMP/GTP Guanosine-5’-di/mono/tri-phosphate Guanylic acid Guanosine-5’-monophosphate IMP/Inosinic acid Inosine-5’-monophosphate
IP Isopentenyl monophosphate
IPP Isopentenyl diphosphate
NAD+/NADH Nicotinamide adenine dinucleotide
NADP+/NADPH Nicotinamide adenine dinucleotide phosphate
NAD-PP NAD pyro-phosphorylase
NDC N1-(2,4-dinitrophenyl)-3-carbamoylpyridinium chloride
NMN Nicotinamide mononucleotide
PPase Pyrophosphatase
Pi Orthophosphate
PC Phosphatidylcholine
PEP-K Phosphoenolpyruvic acid monopotassium salt
PLP Pyridoxal phosphate
POS Phosphosulphate
POSOP Diphosphosulphate
PPi Pyrophosphate
RNA Ribonucleic acid
ScCK Choline kinase from Saccharomyces cerevisiae
TBAHS Tetrabutylammonium hydrogen sulphate
TMP/Thymidylic acid Thymidine-5’-monophosphate UDP/UMP/UTP Uridine-5’-di/mono/tri-phosphate Uridylic acid Uridine-5’-monophosphate
1 Introduction
Phosphorus is one of the main building blocks in life and abundant all around us.1 It plays a role
in the backbone of deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), is a part of cell membranes as phospholipids, aids metabolism and energy transfer as adenosine triphosphate (ATP) or similar coenzymes and is a part of some vitamins. Phosphorus is present in living forms and can be found in cosmetics, food additives, flame retardants, fertilisers and many more applications.2 Most of
the current artificial phosphorus products are synthesised from fossil phosphates extracted from mines.3
These fossil phosphates consist mostly of apatite, Ca10(PO4)6(X)2, where X is F−, OH− or Cl−. Apatite
can easily be converted into fertiliser by treating it with acid and heat. This fertiliser has enabled the world’s food production to increase massively and has allowed the human population to grow to eight billion today. However, the fertiliser has also resulted in diffuse source contamination and eutrophication of water bodies because of agricultural land run-off.4 The removal of the phosphorus
from the water bodies is essential in the diminishing of eutrophication. This can be achieved using geo-engineering with P-absorbent salts or clays.5
Not only lake and river water bodies contain phosphorus, but municipal and industrial wastewater as well. This contaminated water eventually ends up in a wastewater treatment plant, where the phosphorus must be removed to reduce the eutrophication further down the line. 6,7 In many regions the
removal of phosphorus is required by law (e.g., in the EU). The biological nutrient removal process that removes nutrients such as nitrogen and phosphorus from the water can result in a large concentration of phosphorus in the wastewater sludge. The phosphorus can subsequently be precipitated with MgCl2 as
the mineral struvite (Eq. 1).
Mg2+ + NH
4+ + PO43- + 6H2O à MgNH4PO4 6(H2O) (1)
Struvite is a mineral composed of NH4MgPO4 and is easily precipitated by a high pH, saturation
and temperature, and with the aid of present precipitation nuclei.6 This precipitation can enable the
removal of phosphorus, not only from sludges but also from (swine) wastewater, animal manure and landfill leachate. Additionally, the formation of struvite can be used in the removal of phosphorus from natural water bodies. The formed struvite can then be used again as an additive of fertiliser, but it can also be converted to other chemicals. Both options will aid the circular phosphorus economy, as long as the phosphorus is re-used.
The current linear route from source to waste has caused a lot of problems such as climate change, loss of biodiversity and food or water shortages.8 Additionally, the fossil phosphorus sources will
become depleted in the future.2 A change to a circular phosphorus economy is therefore required, if
mankind wants to continue using phosphorus. In this circular economy, the twelve principles of circular chemistry need to be pursued.8 These are: 1) Collect and use waste, 2) maximise atom circulation, 3)
optimise resource, 4) energy, and 5) process efficiency, 6) no release of toxic chemicals, 7) optimal design, 8) sustainability, 9) circularity, 10) service instead of products, 11) allow innovations and 12) unify industry and policy.
A small fraction of the possible future circular phosphorus economy will be discussed, namely the possible biologically relevant phosphate esters that could be produced from struvite. These molecules could be used as food additives, pharmaceuticals, or ion detection in water and are now still manufactured in a linear fashion.9–11 The ultimate goal is to use struvite as feedstock for the production
of all sorts of organophosphates. This is still very much a work in process, however, as the use of struvite as feedstock is still under investigation. This review will focus on the synthesis of the different organophosphates from ordinary starting materials. It will shortly describe the natural applications of the alkyl-phosphates in the body, after which possible alternative applications are described. Only biologically relevant phosphates with such alternative applications are discussed. Subsequently, a possible synthesis of the alkyl-phosphates is reviewed. Lastly, a quick and by all means not complete overview of the current market of the biologically relevant phosphate esters is mentioned by looking at the prices per gram and the commercially maximum available quantity of a fine chemicals’ supplier. This review aims to find which organophosphates are worth trying to be produced from struvite in respect to their current markets. The conclusion will be based on whether the synthesis can include struvite, the greenness of the synthesis based on the twelve principles of circular chemistry, the importance of the alternative application and the market-indication given by the price per gram and the maximum available quantity.
The phosphate esters are either divided by their general structures, such as mononucleotides, dinucleotides, phospholipids, and terpenes, or by their use, vitamins. Phosphorylated proteins and DNA/RNA strands are left out of this review because of the complex structures and complicated control in the synthesis.
2 Mononucleotides
Mononucleotides are the single monomers of the polymers DNA and RNA and are built from three main building blocks, 1) a phosphate group, 2) a sugar (ribose or deoxyribose) and 3) a purine or pyridine.12 An example of a mononucleotide, ATP, is shown in Figure 1. The possible purines and
pyridines vary between RNA and DNA, with DNA and RNA both containing adenine, guanine and cytosine, and DNA containing thymine, while RNA contains uracil.1 Of all possible mononucleotides
(GTP, UTP, GDP, etc.), only adenylic acid/adenosine monophosphate (AMP), guanylic acid/guanosine monophosphate (GMP) and inosinic acid/inosine monophosphate will be discussed in this review because of their possible applications outside of the body.
2.1 Adenosine-5’-monophosphate
Adenosine-5’-monophosphate (AMP), or adenylic acid is the hydrolysis product of adenosine triphosphate (ATP), a source of energy in organisms.12 The energy is generated from the two
phosphoanhydride bonds in the triphosphate. When the triphosphate is hydrolysed, either adenosine diphosphate (ADP) and orthophosphate (Pi) or adenosine monophosphate (AMP) and pyrophosphate
(PPi) are formed. The latter reaction is shown in Figure 1. The reaction to ADP releases about 30.5
kJ/mol energy and the reaction to AMP generates about 45.6 kJ/mol.12 This release depends on a lot of
factors and can differ from one system to another. After the energy release, ATP is again created from ADP and Pi in an oxidising reaction in chemotrophs or phototrophs.
AMP can increase the oxidation of glucose in an insulin-like manner in an adipose tissue, as stated by Dole et al.13 It accelerates the oxidation of glucose and increases the fatty acid synthesis from acetate
in the presence of glucose. Additionally, AMP can suppress the breakdown of triglycerides of fat tissue in an incubation medium with growth hormone, adrenocorticotropic hormone and adrenaline. The AMP probably has a negative effect on fat metabolism and a positive effect on fat synthesis, which gives it its insulin-like effect. However, further research needs to be done to enable adenylic acid as a possible substitution for insulin.
Adenylic acid is synthesised with yeast, a nutrient solution and adenine, described by Kerr et al.14
The yeast was shaken for 16 hours in a nutrient solution with adenine to promote the growth of the yeast. Afterwards, the yeast mixture was centrifuged and washed three times with iced trichloroacetic acid (5%) to remove any remaining adenine. The residue was subsequently washed once with ethanol and three times with a hot 3:1 ethanol-ether mixture to remove lipids. The final wash was done with ether, followed by air drying. The dried residue was incubated for 18 hours at 37 ℃ in 0.3M NaOH to split the RNA nucleotides. The basic solution was then reduced to a pH of 5, resulting in the precipitation of proteins. Again, the mixture was washed by centrifugation. The supernatant and washings were put in an anion exchange resin and washed with water overnight. The resulting solution contained a mixture of 10:14:10:16 5’-adenylic acid, 5’-guanylic acid, 5’-cytidylic acid and 5’-uridylic acid (Figure 2). These mononucleotides were subsequently separated from each other using a column and HCl of increasing concentrations (0.002 M for cytidylic and adenylic acid, 0.003 M for uridylic acid, 0.005 M for guanylic acid). This method results in a 75% yield for adenylic acid and less than 50% for guanylic acid. The synthesis is relatively green because there is not a lot of waste generated, and because little toxins are used (only trichloroacetic acid has an environmental hazard). The only drawback is the limited use of struvite in this method. Perhaps the yeast can use a struvite based nutrient solution to grow.
All five mononucleotides (thymidylic acid for DNA) are commercially available either in the form of their disodium salts, pure or as hydrates for prices that range from €8 to €449 per gram and quantities up to 100g, as shown in Table 1.15 AMP is the cheapest of them all with a price of €8 and
available up to 100 grams.
2.2 Inosinic acid and guanosine-5’-monophosphate
When the free amine group of the adenine in AMP is transformed into a carbonyl (Figure 2), the acid becomes inosinic acid (IMP). It plays a role in the biosynthesis of nucleotide tri- and pyrophosphates and is used in metabolism inside of the body.1
Inosinic acid together with several other chemicals, such as monosodium glutamate, glutamic acid and creatine creatinine, plays a role in meat flavouring.16 Meat with the highest IMP concentration
is usually considered to be tasting better than meat with a lower inosinic acid concentration. The concentration increases a few hours to a few days after slaughter by the breakdown of ATP to ADP, ADP to AMP, and AMP to eventually inosinic acid. The timespan of the breakdown differs from animal to animal and is influenced by the method of slaughter. Because of changes in pH and lactic acid concentration in a strained muscle, the maximum concentration of IMP is found only a few hours after slaughter when the animal has died in agony, instead of a few days. When the meat is stored too long, the decomposition products of IMP are formed: inosine and hypoxanthine, of which the first has no taste, the latter has a bitter taste, and neither contain phosphate.
Additionally, both IMP and monosodium glutamate, together with guanylic acid (GMP), are used in enhancing mushroom flavour in food and beverages because of their umami taste.9 The disodium
salts of IMP and GMP are used as food additives with E-numbers E630 and E627, respectively.17 For
GMP, its dipotassium (E628) and calcium (E629) salts are also used.
IMP and GMP can be extracted from meat, fish (sardines), or bacteria, or are synthesised in the same way as AMP, using yeasts. GMP can also be synthesised from 2’,3’-O-isopropylidene-guanosine stirred in phosphorus oxychloride at 0 to 5 ℃ as described in the patent of Gaines et al. and shown in Figure 3.18 Pyro-phosphoryl chloride was added to the slurry. Subsequently, the cooling bath was
removed. The stirring was continued for another 45 minutes, followed by the evaporation of the solvent by a vacuum below 35 ℃. The remaining oil was put into cold water with LA-1 Amberlite liquid anion exchange resin and Esso solvent WS 4215 and shaken for 2 minutes. The aqueous layer was removed from the organic layer and diluted with dioxane. It was then left for 12 hours at 5 ℃ to form a precipitate. The precipitate was washed twice with dioxane and dried at 45 ℃. It resulted in a guanylic acid
dioxanate (GMP, dioxane and water) crystalline complex that is easily transformed into the GMP disodium salt according to Vitali et al.19 The GMP dioxanate was dissolved in water and neutralised to
a pH of 8 with sodium hydroxide. 95% ethanol was then added to precipitate the product. The disodium salt was filtered and washed first with 1:1 ethanol-water and second with pure ethanol. It was dried in vacuo at 40 ℃ overnight and then exposed to the atmosphere to settle. The yield of the first steps is not mentioned, but the last step from the GMP dioxanate to the GMP disodium salt has a yield of 92.3%. The phosphorus oxychloride is very toxic, therefore the synthesis can be viewed as not green, even though little to no waste is generated and it is all performed at a low temperature. If struvite can be converted into phosphorus oxychloride or pyro-phosphoryl chloride it could be included in the synthesis. IMP is relatively cheap with about €8 per gram, while GMP costs about €50 per gram and both are available up to an quantity of 100 grams (Table 1).15 Not for all mononucleotides in Table 1 an
alternative application has been found, but for comparison all mononucleotides are mentioned. The bulk prices are mentioned as well for the largest available quantities.
Table 1 The prices and commercially available quantities of the monophosphates of all five different mononucleotides.15
Mononucleotide Available as Price (€ per
gram) Maximum quantity (in grams) Minimum price/gram for largest quantity (€)
Adenylic acid Monohydrate 8 100 5.64
Cytidylic acid Disodium salt 62 5 40.40
Guanylic acid Disodium salt
hydrate
50 100 8.09
Uridylic acid Disodium salt 44 10 30
Inosinic acid Disodium salt
hydrate
8 100 4.91
Thymidylic acid Disodium salt
hydrate
2.3 Synthesis of monophosphates
All monophosphates mentioned above and below can probably also be synthesised in a manner without yeast, using their alcohol counterparts, nucleosides, in the case of mononucleotides. Simple monophosphates are readily catalytically synthesised, as described by Domon et al., using a phosphoenolpyruvic acid monopotassium salt (PEP-K) and tetrabutylammonium hydrogen sulphate (TBAHS) system in dimethylformamide (DMF) at 100 ℃, see Figure 4.20 PEP-K acts as a phosphoryl
donor and simultaneously can be protonated by TBAHS to form PEP-H, which reacts with sulphate to phosphosulphate (POS), which itself acts as a sulphuryl donor but in the presence of PEP-K reacts with another PEP-H to form POSOP, which is again a phosphoryl donor. The phosphorylation of an alcohol by POSOP follows an SN1 reaction mechanism and recreates POS. The article does not discuss the exact
phosphorylation of nucleosides, but most yields mentioned by Domon et al. are about 70-80% with a few outliers to 20%. The synthesis is really green, because of the catalytic use of the POS and POSOP, no waste generation and no use of toxins. If struvite can be converted into PEP-K, it could be included in the synthesis and make it even greener.
This method has a high functional group tolerance and is a relatively easy single-step phosphorylation. Therefore, it might be able to produce mononucleotides from their nucleoside counterparts. This method can probably also be used in the synthesis of monophosphates mentioned further on in this review.
The nucleosides (cytidine, guanosine, thymidine, uridine, adenosine and inosine) are readily commercially available for a price ranging from €15 to €57 per gram and quantities up to 25 to 500 grams, shown in Table 2.15
Table 2 The prices and quantities of commercially available nucleosides.15
Nucleoside Price (€ per gram) Maximum quantity (in
grams)
Minimum price per gram for largest quantity (€) Adenosine 15 100 2.69 Cytidine 16 10 5.50 Guanosine 54 500 1.94 Inosine 23 100 3.71 Uridine 26 100 4.66 Thymidine 57 25 3.11
3 Dinucleotides
Nicotinamide adenine dinucleotide (NAD+/NADH) and nicotinamide adenine dinucleotide
phosphate (NADP+/NADPH) are, just like ATP, an energy carrier for the metabolism in the body.1 Their
structures are shown in Figure 5. Instead of transporting whole groups as ATP does, NADH and NADPH transfer electrons, and that is the exact reason why these dinucleotides can be used in biofuel cells.21
Biofuel cells generate electrical energy by fuel oxidation at the anode, transferring charge and reducing a second compound at the cathode. Different kinds of fuels can be used, but only a methanol/oxygen biofuel cell will be considered, shown in Figure 6. NAD+-dependent enzymes (ADH, AldDH and FDH)
are used to oxidise methanol to carbon dioxide at the anode. In this reaction, three equivalents of NAD+
are needed for every equivalent methanol. NADH is formed, and NAD+ should be regenerated for the
reaction to be catalytic. The regeneration of NAD+ can be achieved using benzyl viologen (BV) and
diaphorase (D). The current flow then reduces the oxygen at the cathode and to close the circuit a
a) b)
Figure 5 a) The structures of NAD+ and NADP+, b) NADH and NADPH.
positive charge is transferred over a Nafion membrane. The methanol/oxygen biofuel cell makes a relatively good cell with a Vcell, max of 0.8 V, and a power density of 0.68 mW/cm2 at 0.49 V.
3.1 Synthesis of NAD
+and NADP
+Walt et al. have suggested a synthesis of NADP+ from AMP, shown in Figure 7.22 First, AMP is
hydrolysed by acid-catalysis to ribose 5’-phosphate. The ribose phosphate is then treated with anhydrous ammonia in dry ethanediol at 0 ℃ for 1 hour to aminate one alcohol group. After a week of storage at 4 ℃ excess ammonia was removed using a rotary evaporator and a vacuum pump. The product was subsequently stirred with N1-(2,4-dinitrophenyl)-3-carbamoylpyridinium chloride (NDC) in the dark at
room temperature for 18 hours to form nicotinamide mononucleotide (NMN). Water was added to precipitate 2,4-dinitroaniline, which was removed by filtration. The excess NDC was removed with activated charcoal and filtration. After that, NMN reacted with ATP, catalysed by NAD pyro-phosphorylase (NAD-PP) immobilised in a Polyacrylonitrile gel at a pH of 7.2. Magnesium chloride, 1,3-dimercapto-2-propanol and diammonium acetyl phosphate solution (to keep ATP concentration above Km for NAD-PP) were added, and the mixture was put under argon. For ten days, additional NMN
and AMP were added. The equilibrium reaction was driven to completion by hydrolysis of the pyrophosphate product using pyrophosphatase (PPase). The NAD+ solution could be used directly,
without further purification for cofactor enzymatic syntheses. NADP+ could be synthesised from the
NAD+ by NADK, AcK and the transformation from AcP to Ac-. The synthesis has a 25% yield from the
ribose-5’-phosphate to NMN and from there a 90-97% yield to NAD+. The synthesis is not that great
because 2,4-dinitroaniline is also synthesised, which is explosive, toxic and has health and environmental hazards. The use of ethanediol is also not that great, as it is moderately toxic. Additionally, the synthesis is not able to use any struvite-based product other than AMP, and because the inclusion of struvite in AMP synthesis is limited, the NAD+ synthesis is not green at all.
NAD+ is readily commercially available for about €70 per gram and quantities up to 25 grams,
NADP+ is available for €835 per gram up to an quantity of 1 gram.15
4 Phospholipids
Phospholipids are the main components of the cell membranes, play a major role in transporting signals across these membranes, and are used as an energy source.1,23 They consist of long hydrophobic
hydrocarbon tails that can either be saturated or unsaturated, a glycerol backbone and a polar headgroup containing the phosphate group and possible additions; an example is shown in Figure 8. Some of the possible polar headgroups are choline, inositol, ethanolamine, or serine. Phospholipids are used a lot for new material synthesis, both in vivo and in vitro.
Their ability to self-assemble, compartmentalise and template gives phospholipids the ability to form the basis for several micro- and nanostructures.23 In liquids, phospholipids immediately
self-assemble into micelles, reversed micelles, mono- and bilayers, or hexagonal phases. The lipid nanostructures that are formed are determined by lipid polarity, chain length, the location of unsaturation and branching, the extent of the two, the size, charge and headgroup polarity, the concentration and the temperature.
4.1 Liposomes
The most important nanostructure both in vivo and in vitro are closed vesicles of bilayers called liposomes. They can entrap hydrophilic molecules in their inner compartment and hydrophobic ones in their membranes. Because of this ability, they are currently being tested as drug delivery vehicles, for example, in anticancer medicine.24,25 The ideal use of liposomes would enable a high drug concentration
within the liposome, a high biological, physical, chemical stability and the selectivity to release their contents at the desired place.11 The size of the liposomes needs to be controlled, however, to make sure
they are not too large to be able to move into tissues, but that they are also not readily filtered out by the kidney because they are too small.26 The liposomes can be fabricated by natural phospholipids, such as
egg lecithin, or synthetically produced. Lecithin can easily be extracted from eggs with the use of hexane and ethanol.27 The downside of using lipids from a natural origin is the limited control over the lipid
structure. Therefore, synthetic lipids are often used, such as 1,2-dioleoyl-sn-glycero-3-phosphocholine and 1,2-dimyristoyl-sn-glycero-3-phosphocholine, which are commercially available for about €465 or €903 per gram, respectively, and quantities up to 1 gram.15,28
4.2 Phosphatidylserine
Phosphatidylserine (PS) is a specific phospholipid with a serine headgroup that can mimic apoptotic cells and stimulates phagocytosis by macrophages.29 Petazzi et al. have created an
asymmetrical immunoliposome with PS on the inner sphere and antibodies attached to phospholipids on the outer sphere that can recognise HIV-1 virus-like particles.30 The idea is schematically shown in
Figure 9. The antibodies will bind HIV-1, the PS will move to the outer sphere in the span of a few hours by a spontaneous flip-flop. Flip-flop occurs in asymmetric liposomes with a different inner and outer sphere and reduces the asymmetry. Subsequently, the whole thing is removed from the system by uptake of the mononuclear phagocytose system. This might reduce the effect HIV-1 has on the patient’s immune system or could be a step towards the cure for HIV-1. However, more research is needed, also on the possibility of this immunoliposome for the cure for SARS-CoV-2. Phosphatidylserine is commercially available for about €4810 per gram, up to an quantity of 100 mg.15
4.3 Phosphatidylcholine
Phosphatidylcholine (PC) (Figure 8) is a phospholipid with anti-inflammatory properties that can help people with ulcerative colitis, a chronic inflammatory colon disorder.31 Ulcerative colitis is
characterised by a reduced amount of PC in the colon mucus and a reduced hydrophobic protective layer. The addition of extra PC reduces the inflammatory response of the colon and can reduce ulcerative colitis symptoms. The phospholipid can substitute the less preferable steroid or immunosuppressive therapies. It is commercially available for about €454 to €924 per gram and up to a quantity of 1 gram for the pure compound.15 PC is also available as a less pure mixture up to 1 kg for €203 per kilogram.
4.4 Synthesis of phospholipids
Phospholipids can be synthesised from 3-O-benzylglycerol, 4-dimethylaminepyridine (DMAP) and a carboxylic acid with the preferred chain length, shown in Figure 10.32 In the general synthesis of
1,2-diacyl-O-benzylglycerol described by Martin et al. dicyclohexylcarbodiimide (DCC) in dichloromethane (DCM) was dropwise added to a solution of the aforementioned chemicals in dry DCM at 0 ℃ or 20 ℃ over different timespans and in different concentrations, depending on the desired product. The two carbon chains can be the same length or varying lengths, depending on the conditions and the step repetition. The product that is formed was filtered through Celite and dried under vacuum.
The product was purified by either Kugelrohr distillation, flash chromatography, or HPLC. After purification, the benzyl group was removed in a 10:1 (volume) ethanol:glacial acetic acid solution containing 10% Pd/C under H2 at room temperature. After completion of the reaction, the solution was
diluted with DCM and vacuum filtrated with Celite. The storage of the 1,2-O-diacylglycerols needs to be carefully tuned per diacylglycerol because 2,3-acyl migration occurs under the wrong conditions to form 1,3-O-diacylglycerols. A phosphite (methyl-dichloro phosphite, phenyl-dichlorophosphite or 2-(trimethylsilyl)ethyl phosphite) acts as a coupling agent in dialkylglycerols phosphorylation. The phosphite reacts with an alcohol that has a protecting group (e.g., bromoethanol or HO(CH2)2NHBoc)
in the presence of N,N-Diisopropylethylamine (DIPEA). The alcohol present in the 1,2-O-diacylglycerols to form an intermediate phosphite triester, and becomes, after oxidation and deprotection by hydrogen peroxide, the phospholipid. The polar head groups can be attached in a later step. For partly unsaturated phospholipids, first, a saturated carboxylic acid is attached to a glycerol. A protected phosphate is then added, and only after that, the unsaturated carboxylic acid is attached to the glycerol. This is probably done because otherwise, the double bond will react with the remaining alcohol group. The yield of the 1,2-O-diacylglycerols, no matter the carbon chain lengths, was always over 55%. For phosphatidylcholine, the synthesis from the 1,2-O-diacylglycerol to the phospholipid with the bromine headgroup is 71% and from there to the end product is 83%. The total synthesis yield is 32%. The yield of phosphatidylserine in a slightly different synthesis is 71%. The use of DCC, DMAP, DIPEA and bromoethanol makes the synthesis not that green, as they are toxic, and bromine is generated as waste. However, struvite can be incorporated into the synthesis if it can be transformed into a phosphite.
4.5 Glycerophosphate
A single component of phospholipids, the glycerophosphate, plays a role in drug administration.33
Chitosan (a polysaccharide) is already used as a mucoadhesive but its low mechanical strength decreases the drug residence time. A mucoadhesive adheres to mucus, or wet substances in the body – e.g., saliva or snot. The addition of β-glycerophosphate to the chitosan increases the mechanical strength, mucoadhesiveness and lowers the cytotoxicity. The chitosan and β-glycerophosphate mixture can be used in local drug delivery almost everywhere in the body, sometimes as a nanoparticle loaded hydrogel with liposomes encompassing drugs.34 Another application of the hydrogel is tissue engineering, though
usually, some other chemical needs to be added to finetune the hydrogel characteristics.
β-glycerophosphate is readily commercially available as its disodium hydrate for about €5 per gram and quantities up to 500g.15
5 Terpenes
Terpenes are compounds built from isoprene (2-methyl-1,3-butadiene) monomers. They smell pleasant, taste spicy, exhibit certain pharmacological activities, and act as pheromones.36 They are most
often found in plants and insects and fulfil all different kinds of roles, such as plants attracting insects by smell, or insects warning other insects in alarm pheromones. Terpenes that are more interesting to humans are carotene and natural rubber. Carotene is used as a colourant and is present in carrots, and natural rubber is a polyisoprene used for the rubber production.
Terpenes generally do not have phosphates attached to them; however, phosphates play a role in the terpene biosynthesis, using enzymes (prenyltransferases: tPT, cPT), shown in Figure 11.1,36,37 First
the formation of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) takes place, then the condensation of IPP onto DMAPP to form oligoprenyl diphosphate occurs. Another IPP can condensate on the oligoprenyl diphosphate to lengthen the chain. Eventually, the pyrophosphate is removed and transformed into another group to form natural rubber or other terpenes. The stereochemistry of the isoprene units determines the characteristics of the substance and whether rubber or gutta-percha is synthesised. Gutta-percha is a more rigid thermoplastic latex that is used in dentistry and for electrical insulation, and that has only trans-polyisoprene units.38
The rubber market has grown since the coronavirus outbreak because more rubber is needed for medical gloves.39 As of 9 December 2020, it is sold at about €0.002 per gram (€2 per kg).
IPP and DMAPP can easily be synthesised using prenol or isoprenol as starting materials described by Chatzivasileiou et al.40 A choline kinase from Saccharomyces cerevisiae (ScCK)(a yeast)
and ATP as cofactor can synthesise isopentenyl monophosphate (IP) and dimethylallyl monophosphate (DMAP2). Isopentenyl phosphate kinase from Arabidopsis thaliana (a plant) can then be added to the
ScCK to catalyse the second phosphorylation to IPP and DMAPP, although ScCK is also capable to perform the second phosphorylation on its own. The yield is not mentioned by Chatzivasileiou et al. The incorporation of struvite is limited, but perhaps the yeast and plant can be grown on struvite-based nutrients. The synthesis is quite green.
IPP is commercially available for about €31,900 per gram, up to an quantity of 50 mg, DMAPP is available for a price of €33,900 per gram, also up to an quantity of 50 mg.15
6 Vitamins
Vitamin A is a part of the beforementioned terpenes and might be produced with phosphates within the body; however, it has no phosphate groups and will not be further analysed.
6.1 Vitamin B6
Pyridoxal phosphate (PLP) is one of the active components of vitamin B6 and acts as a coenzyme in transamination, decarboxylation, and racemisation reactions in the body.1 Additionally, it is an
essential cofactor for amino acid metabolism. Outside of the body, pyridoxal phosphate can be used as a detection substance for metal ions in aqueous environments, with a limit of detection close to the maximum permitted amount in drinking water.10 In this way, it can help determine the nature of the
contamination in water bodies that need to be as clean as possible. Because of its conjugated system, PLP shows fluorescent properties, and interaction with different metals transforms PLP’s colour. In the presence of Fe3+, PLP changes from colourless to yellow. While, in the presence of both Fe3+ and Al3+
the fluorescence is quenched and blue-shifted. When gold nanoparticles are crowned with PLPs, they turn from red to purple/blue in the presence of Al3+, Cd2+ and Pb2+ ions. These gold nanoparticles can
easily be added to a paper strip, which allows for easy detection methods. If the PLP is condensed with 2-amino thiophenol it can also detect F- and AcO-, which may be useful detecting cytoplasmic fluoride.41
remaining suspensions were added to 2 M NaOH and extracted twice with diethyl ether. 2 M hydrochloric acid was then added to the water layer. The solution was cooled to 0-5 ℃ overnight. Pure PLP could easily be filtrated out of the solution in its crystalline form. Iwanami et al. do not elaborate on the selectivity of the phosphorylation. The yield ranges from 40% for a chlorinated phenyl R-group to 92% for an ethoxy phenyl R-group. The synthesis is relatively green. Struvite can be included if transformed into phosphorus pentoxide or phosphorus acid.
Pyridoxal phosphate is commercially available in its hydrate form for about €37 per gram and up to an quantity of 25 grams.15
6.2 Vitamin B12
Vitamin B12 (Figure 13a) is not produced in human bodies and needs to be present in the food consumed.43 It plays a role in amino acid and nucleotide metabolism and is essential for life. Vitamin
B12 derivatives, where the R group is transformed, can be used for all sorts of different applications, such as diagnosis or therapy drugs.
When the R group in vitamin B12 is substituted with cyanide-bridged cisplatin (Figure 13b) it can be used as an anticancer drug.44 Cisplatin itself is a good anticancer drug. Still, to be effective, it has
to be administered in large doses, and its toxicity to healthy cells as well as cancer cells causes severe side effects. However, the fast-multiplying cancer cells need a lot of vitamin B12, so when the cisplatin is connected to vitamin B12, it can selectively reach cancer cells and destroy them. The cisplatin is easily added to the vitamin B12 by adding a cis-[PtCl(NH3)2(OH2)]+-complex to the vitamin in water
and stirring it for 16 hours at 50 ℃. The yield is 72% and the synthesis is green.
Figure 13 Structure of vitamin B12 and its derivatives: a) regular vitamin B12, b) Cisplatin B12 from Mundwiler et
al.44, c) Rhenium complex B12 from Zobi et al.43,45
a)
b)
When the R group in vitamin B12 is substituted by a cyanide-bridged cis-trans-[ReII(CO) 2Br2]0
-complex, its solubility in water is increased, and the complex can act as a CO-releasing molecule, see Figure 13c.43,45 Carbon monoxide can, in turn, act as an anti-inflammatory agent, a blood vessel widener
in cardiovascular diseases, or play a role in organ preservation and transplantation. The use of an M-CO complex is much safer and easier than using gaseous CO. The Rhenium complex can be connected to the vitamin B12 by adding [Et4N]2[ReIIBr4(CO)2] in methanol to the vitamin under a nitrogen
atmosphere and stirring it for 1 hour while letting the temperature drop from 50 to 40 ℃. The mixture was then stirred for another additional half-hour at 40 ℃. Methanol was then removed under reduced pressure, followed by washing of the red powder multiple times with DCM and acetone until the liquids were clear. The yield is 98% and the synthesis is relatively green.
Vitamin B12 can also be used as a fluorescent biomarker for certain organs in the body with a high vitamin B12 concentration.43 It can, for example, help during cancer cell removal operations, as
they contain a lot of vitamin B12. Vitamin B12 can become fluorescent when the ‘5-OH in the deoxyribose is changed into an ester with attached a fluorescent group. An example from Lee et al. is further elaborated here.46
The synthesis of this structure is a little more intricate than the synthesis of the beforementioned structures, see Figure 14.46 Vitamin B12 was dissolved in anhydrous dimethyl sulfoxide (DMSO), and
1,1’-Carbonyl-di-(1,2,4-triazole) (CDT) was added. The solution was stirred for 30 minutes, and subsequently slowly added to a rapidly stirring 1:1 ether:chloroform mixture. The precipitate that formed was collected by vacuum filtration and washed with acetone. The product was dissolved again in DMSO and dropwise added to a trans-1,4-diaminocyclohexane solution in DMSO. The mixture was stirred for 1 hour and 20 minutes and again added to a rapidly stirring ether chloroform mixture, vacuum filtrated and washed with acetone.
Similarly, the product was dissolved in DMSO and 5(6)-fluorescein NHS ester and DIPEA were added. The mixture was stirred for 4 hours and subsequently added to a rapidly stirring 1:1 DCM:ether solution. The final red product was retrieved by vacuum filtration and dried over phosphorus pentoxide in a vacuum. The first step of the synthesis, the addition of half of the CDT has a 75% yield, the second 60% and the overall yield is only 22%. The use of DIPEA is not preferred because of its toxicity. Furthermore, quite a lot of waste is generated, such as 1,2,4-triazole and pyrrolidine-2,5-dione. Struvite cannot play the role of phosphate source in this synthesis, as no phosphate is used.
Bacteria commercially produce vitamin B12 because the total chemical synthesis consists of about 70 steps and is very complicated due to the many chiral centres.47 A particular strain of bacteria
from the pseudomonas denitrificans is primarily used for the synthesis. The bacteria are fermented for 2 to 3 days at 30 ℃ and a slightly neutral-acidic pH. Usually, cobalt ions and 5,6-dimethylbenzimidazole are added to the bacteria. After the fermentation, the whole broth is heated to 80 to 120 ℃ for 10 to 30 minutes at a pH of 6.5 to 8.5 for the vitamin B12 extraction. The conversion to cyanocobalamin is achieved by adding cyanide or thiocyanate to the suspension. After filtration of the suspension, the vitamin B12 in the filtrate is precipitated by tannic acid or cresol. The product can be further purified and crystallised, depending on the intended uses for the vitamin. The synthesis is quite green because of the use of bacteria. The addition of cyanide or thiocyanate is not preferred because of their toxicity, however, for the alternative applications of vitamin B12, this addition is not necessary. Struvite could play a role as nutrient source for the bacteria. The article does not state the yield of the reaction.
7 Discussion
This review aims to find which organophosphates are worth trying to be produced from struvite in respect to their current markets. For this analysis the synthesis will be reviewed whether it can include struvite in some way, either as inorganic struvite or in an organic/altered form, the greenness of the synthesis based on the twelve principles of circular chemistry, additionally, the importance of the alternative application and the market-indication given by the price per gram and the maximum available quantity will be analysed.
The yields of the syntheses have a large range (Table 3), and a lot of articles do not mention the yield of the whole synthesis but only fer certain steps. For GMP the yeast-based synthesis is not referred to because of the lower than 50% yield. The synthesis for the NAD+, GMP and the phospholipids is not
green because of the low yields, the use of toxic chemicals and the bad atom economies. Additionally, the inclusion of struvite into the NAD+ synthesis is not possible because AMP is used as phosphate
source (AMP can be synthesised with the use of struvite). The other syntheses can include struvite in some way, the product in which struvite should be converted or used as are stated in the table.
Table 3 Synthesis analysis of the biologically relevant phosphate esters.
Biologically relevant phosphate ester Overall yield of synthesis Greenness of synthesis Inclusion of struvite
AMP 75% Relatively green Nutrient source
IMP Not mentioned Relatively green Nutrient source
GMP Not mentioned Not green Phosphorus oxychloride
or pyro-phosphoryl chloride
NAD+ Not mentioned (first
step is 25%)
Not green No
Phospholipids Not mentioned Not green Phosphite
PS 71% Not green Phosphite
PC 32% Not green Phosphite
Glycerophosphate 37.4% Relatively green Ammonium dihydrogen
The alternative applications range from anti-inflammatory to food additives to ion detection and are summarised in Table 4. The most promising new applications are the use of PS immunoliposomes for HIV-1 phagocytosis, PC for the therapy of ulcerative colitis, PLP for ion detection in water and vitamin B12 as anticancer drug. The alternative applications of beforementioned phosphate esters will be put into perspective with the synthesis below.
Table 4 A quick overview of the applications of the biologically relevant alkyl-phosphates that differ from their regular applications.
Biologically relevant phosphate ester Application
AMP An effect similar to that of insulin
IMP Flavouring of meat/umami taste
GMP Induce mushroom flavour/umami taste
NAD+ Biofuel cell
Phospholipids Drug delivery liposomes
PS HIV-1 phagocytosis
PC Therapy for ulcerative colitis
Glycerophosphate Drug administration hydrogels
DMAPP/IPP Synthesis of terpenes/polyprenols
PLP Ion detection in water
Vitamin B12 Anticancer drug, CO releasing molecule, biomarker
The PS synthesis is relatively difficult and does not meet the circular chemistry goals. However, when struvite can sustainably be transformed into a phosphite, its synthesis could become a little greener. The synthesis of immunoliposomes is not described because it lies outside this review’s scope. However, the synthesis seems complicated because of the selectivity for the PS to be on the inner side of the sphere. Additionally, antibody addition is probably not as straightforward as it seems, because of their intricate structures. Nevertheless, the promise of becoming a therapy for HIV-1 patients could be worth all the trouble.
The PC synthesis is almost the same as that for PS, but since PC could be a less side-effect-inducing therapy than the current steroids or immunosuppressive therapies for ulcerative colitis patients, it could also be worth trying.
The synthesis of PLP is relatively green. It can include struvite when struvite can be transformed to phosphorus pentoxide. The synthesis might even be possible with simple pyrophosphate, but no information on that possibility could be found. Additionally, ion detection in water could be very useful for drinking water quality control and for the treatment of wastewater.
Lastly, the vitamin B12 synthesis with a cyanide-bridged cisplatin is relatively green and sustainable, because bacteria are used for the synthesis of vitamin B12 and the cisplatin addition occurs in mild conditions without waste generation. The bacteria need some sort of phosphate source, which
could be a renewable source based on struvite. Because of these sustainable synthesis processes, using Vitamin B12 as anticancer drug could become competitive with the current radiation therapies and has the potential of saving many lives.
All of the analysed biologically suitable phosphate esters (excluding the immunoliposomes) are commercially available at Sigma-Aldrich (Merck). Their prices differ significantly between the different compounds and range from €8 to €33,900 per gram, as shown in Table 5. The market favourability depends on many factors, such as the production costs, the prices determined by supply and demand, and whether the synthesis can be done with a renewable phosphorus source like struvite. For a more elaborate market research, more retailers and price ranges per product should be investigated. Additionally, the demand should be added to the research, as well as the production costs.
Table 5 Overview of the prices and quantities of the biologically relevant alkyl-phosphates reviewed in this thesis.
Biologically relevant phosphate ester Price (€ per gram) Maximum available
quantity (gram) AMP 8 100 IMP 8 100 GMP 50 100 NAD+ 70 25 1,2-dioleoyl-sn-glycero-3-phosphocholine 465 1 1,2-dimyristoyl-sn-glycero-3-phosphocholine 903 1 PS 4810 0.100 PC 400-900 1a Glycerophosphate 5 500 DMAPP/IPP 33,900/31,900 0.050 PLP 37 25 Vitamin B12 139 25
a for pure compound, 1 kg for less pure mixtures.
The glycerophosphate market could be the most favourable as the maximum available quantity is largest of them all and the production is relatively easy. However, the price is relatively low per gram,
relatively easy. If struvite could be used as renewable phosphorus source for the bacteria growth and IPP/DMAPP production, the process will be even better.
On the other hand, the phosphatidylserine production is quite complicated, but if the production of struvite to phosphite is possible it could become profitable. The commercially available quantities for DMAPP/IPP and phosphatidylserine are not large, but this may change if better synthesis methods are developed or more alternative applications are found to increase the demand. These alternative applications could be a synthetic rubber synthesis using IPP/DMAPP or multiple purposes for PS containing immunoliposomes to stimulate phagocytosis.
Vitamin B12, PLP, GMP and NAD+ all are available for a moderate price and up to a moderate
quantity. These markets could thus also be promising for a more moderate market revenue.
Further research can be done on the possible products from struvite, or other renewable phosphorus sources. The synthesis of the organophosphates could be improved or altered to include these renewable products. Additionally, the up-scalability of the synthesis should be looked at before upscaling, as these were all lab-scale syntheses. The market research could be done more extensively, including the worldwide supply, demand and prices. And, finally, ever more research can be done on more biologically relevant alkyl-phosphates because more alternative applications can possibly be found.
8 Conclusion
Phosphorus is one of the main building blocks in life, present in DNA, cell membranes, cofactors and vitamins. Its use as fertiliser causes problems, including eutrophication of natural water bodies. The mineral struvite can solve these problems, as it precipitates easily from aqueous environments and its formation is not difficult. Struvite can aid the circular phosphorus economy where phosphorus is constantly reused. One of the possible applications of phosphorus where struvite could be implemented is the synthesis of biologically relevant alkyl-phosphates, which have been reviewed in this thesis. The biologically relevant alkyl-phosphates are analysed on multiple fronts: the greenness of the synthesis and whether it can include struvite in some way, the importance of the alternative application and the market-indication given by the price per gram and the maximum available quantity. The two biologically relevant alkyl-phosphates that were most promising on all fronts, were PLP and vitamin B12, with a green and possible struvite-including synthesis, a relatively important application and a moderate market pricewise and quantity-wise. However, the anticancer application of vitamin B12 overrules the ion detection of PLP and therefore vitamin B12 is the most promising biologically relevant alkyl-phosphate of them all. However, more research is necessary to be able to draw a better supported conclusion. The possibilities for sustainably produced biologically relevant alkyl-phosphates from struvite are almost endless and more research is needed on closing the phosphorus circle.
Acknowledgements
Firstly, I’d like to thank Steven Beijer and Chris Slootweg, who, despite their already incredibly busy schedules, agreed to help me with this literature study. I hope I have delivered what you asked for and have not disappointed you. Secondly, I want to thank Francesco Mutti for agreeing to being my second examiner, I hope I have understood all the biological parts correctly. Thirdly, I want to thank Davita van Raamsdonk for studying together and putting up with my never-ending fun facts to distract her from her own literature study. Lastly, I want to thank Tori Gijzen for helping me find some sources, proofreading the final version and helping me remembering how to write chemistry theses.
References
(1) Corbrigde, D. E. C. Phosphorus - Chemistry, Biochemistry and Technology, 6th ed.; 2013. (2) Reijnders, L. Phosphorus Resources, Their Depletion and Conservation, a Review. Resour.
Conserv. Recycl. 2014, 93, 32–49. https://doi.org/10.1016/j.resconrec.2014.09.006.
(3) Samreen, S.; Kausar, S. Phosphorus Fertilizer: The Original and Commercial Sources; 2016. (4) Daniel, T. C.; Sharpley, A. N.; Lemunyon, J. L. Agricultural Phosphorus and Eutrophication: A
Symposium Overview. J. Environ. Qual. 1998, 27 (2), 251–257. https://doi.org/10.2134/jeq1998.00472425002700020002x.
(5) Spears, B. M.; Maberly, S. C.; Pan, G.; Mackay, E.; Bruere, A.; Corker, N.; Douglas, G.; Egemose, S.; Hamilton, D.; Hatton-Ellis, T.; et al. Geo-Engineering in Lakes: A Crisis of Confidence? Environ. Sci. Technol. 2014, 48 (17), 9977–9979. https://doi.org/10.1021/es5036267.
(6) Parsons, S. A.; Doyle, J. D. Struvite Formation, Control and Recovery. Water Res. 2002, 36 (16), 3925–3940.
(7) European Commission. Urban waste water directive
https://ec.europa.eu/environment/water/water-urbanwaste/legislation/directive_en.htm (accessed Dec 2, 2020).
(8) Keijer, T.; Bakker, V.; Slootweg, J. C. Circular Chemistry to Enable a Circular Economy. Nat.
Chem. 2019, 11 (3), 190–195. https://doi.org/10.1038/s41557-019-0226-9.
(9) Matsui, M.; Sano, H.; Ota, M. Composition for Enhancing Mushroom Flavor, 2019.
(10) Bothra, S.; Upadhyay, Y.; Kumar, R.; Sahoo, S. K. Applications of Vitamin B6 Cofactor Pyridoxal 5′-Phosphate and Pyridoxal 5′-Phosphate Crowned Gold Nanoparticles for Optical Sensing of Metal Ions. Spectrochim. Acta - Part A Mol. Biomol. Spectrosc. 2017, 174, 1–6. https://doi.org/10.1016/j.saa.2016.11.014.
(11) El-Hammadi, M. M.; Arias, J. L. An Update on Liposomes in Drug Delivery: A Patent Review (2014-2018). Expert Opin. Ther. Pat. 2019, 29 (11), 891–907. https://doi.org/10.1080/13543776.2019.1679767.
(12) Berg, J. .; Tymoczko, J. L.; Gatto, G. J.; Stryer, L. Biochemistry, 8th ed.; W.H. Freeman and Company, 2015.
(13) DOLE, V. P. Insulin-like Actions of Ribonucleic Acid, Adenylic Acid, and Adenosine. J. Biol.
Chem. 1962, 237 (9), 2758–2762.
(14) Kerr, S.; Seraidarian, K.; Bosworth Brown, G. On the Utilization of Purines and Their Ribose Derivatives by Yeast. J. Biol. Chem. 1950, 188, 207–217.
(15) Merck. Merck Database https://www.sigmaaldrich.com/nederland.html (accessed Dec 2, 2020). (16) Terasaki, M.; Kajikawa, M.; Fujita, E.; Ishii, K. Studies on the Flavor of Meats. Agric. Biol.
(17) European Commission. Food additive Database https://ec.europa.eu/food/safety/food_improvement_agents/additives/database_en (accessed Dec 4, 2020).
(18) Gaines, W. A.; Reinhold, D. F. Production of Guanylic Acid. 3,448,098, 1969.
(19) Vitali, R. .; Inamine, E. .; Rothrock, J. .; Jacob, T. . Guanosine-’5-Monophosphate Dioxanate: Preparation, Characterization and Application. 1965, 112, 313–321.
(20) Domon, K.; Puripat, M.; Fujiyoshi, K.; Hatanaka, M.; Kawashima, S. A.; Yamatsugu, K.; Kanai, M. Catalytic Chemoselective O ‑ Phosphorylation of Alcohols. 2020.
https://doi.org/10.1021/acscentsci.9b01272.
(21) Palmore, G. T. R.; Bertschy, H.; Bergens, S. H.; Whitesides, G. M. A Methanol/Dioxygen Biofuel Cell That Uses NAD+-Dependent Dehydrogenases as Catalysts: Application of an Electro-Enzymatic Method to Regenerate Nicotinamide Adenine Dinucleotide at Low Overpotentials. J. Electroanal. Chem. 1998, 443 (1), 155–161. https://doi.org/10.1016/S0022-0728(97)00393-8.
(22) Walt, D. R.; Rios-Mercadillo, V. M.; Auge, J.; Whitesides, G. M. Synthesis of Nicotinamide Adenine Dinucleotide (NAD) from Adenosine Monophosphate (AMP). J. Am. Chem. Soc. 1980,
102 (26), 7805–7806. https://doi.org/10.1021/ja00546a041.
(23) Collier, J. H.; Messersmith, P. B. Phospholipid Strategies in Biomineralization and Biomaterials Research. Annu. Rev. Mater. Res. 2001, 31, 237–263.
(24) Gregoriadis, G. Engineering Liposomes for Drug Delivery: Progress and Problems. Trends
Biotechnol. 1995, 13 (12), 527–537. https://doi.org/10.1016/S0167-7799(00)89017-4.
(25) Federman, N.; Denny, C. T. Targeting Liposomes toward Novel Pediatric Anticancer
Therapeutics. Pediatr. Res. 2010, 67 (5), 514–519.
https://doi.org/10.1203/PDR.0b013e3181d601c5.
(26) Alexis, F.; Pridgen, E.; Molnar, L. K.; Farokhzad, O. C. Factors Affecting the Clearance and Biodistribution of Polymeric Nanoparticles. Mol. Pharm. 2008, 5 (4), 505–515. https://doi.org/10.1021/mp800051m.
(27) Palacios, L. E.; Wang, T. Egg-Yolk Lipid Fractionation and Lecithin Characterization. JAOCS,
J. Am. Oil Chem. Soc. 2005, 82 (8), 571–578. https://doi.org/10.1007/s11746-005-1111-4.
(28) Zylberberg, C.; Matosevic, S. Pharmaceutical Liposomal Drug Delivery: A Review of New Delivery Systems and a Look at the Regulatory Landscape. Drug Deliv. 2016, 23 (9), 3319–
Virus-like Particles. Nanomedicine Nanotechnology, Biol. Med. 2015, 11 (8), 1985–1992. https://doi.org/10.1016/j.nano.2015.06.004.
(31) Stremmel, W.; Merle, U.; Zahn, A.; Autschbach, F.; Hinz, U.; Ehehalt, R. Retarded Release Phosphatidylcholine Benefits Patients with Chronic Active Ulcerative Colitis. Gut 2005, 54 (7), 966–971. https://doi.org/10.1136/gut.2004.052316.
(32) Martin, S. F.; Josey, J. A.; Wong, Y. L.; Dean, D. W. General Method for the Synthesis of Phospholipid Derivatives of 1,2-O-Diacyl-Sn-Glycerols. J. Org. Chem. 1994, 59 (17), 4805– 4820. https://doi.org/10.1021/jo00096a023.
(33) Szymańska, E.; Sosnowska, K.; Miltyk, W.; Rusak, M.; Basa, A.; Winnicka, K. The Effect of β-Glycerophosphate Crosslinking on Chitosan Cytotoxicity and Properties of Hydrogels for Vaginal Application. Polymers (Basel). 2015, 7 (11), 2223–2244. https://doi.org/10.3390/polym7111510.
(34) Zhou, H. Y.; Jiang, L. J.; Cao, P. P.; Li, J. B.; Chen, X. G. Glycerophosphate-Based Chitosan Thermosensitive Hydrogels and Their Biomedical Applications. Carbohydr. Polym. 2015, 117, 524–536. https://doi.org/10.1016/j.carbpol.2014.09.094.
(35) Epps, D. E.; Nooner, D. W.; Eichberg, J.; Sherwood, E.; Oró, J. Cyanamide Mediated Synthesis under Plausible Primitive Earth Conditions - VI. The Synthesis of Glycerol and Glycerophosphates. J. Mol. Evol. 1979, 14 (4), 235–241. https://doi.org/10.1007/BF01732490. (36) Breitmaier, E. Terpenes: Flavours, Fragrances, Pharmaca, Pheromones; Wiley-VCH:
Wenheim, 2006.
(37) Yamashita, S.; Takahashi, S. Molecular Mechanisms of Natural Rubber Biosynthesis. Annu. Rev.
Biochem. 2020, 89, 821–851. https://doi.org/10.1146/annurev-biochem-013118-111107.
(38) Lexico. Gutta-Percha.
(39) Trading Economics. Rubber price https://tradingeconomics.com/commodity/rubber (accessed Dec 9, 2020).
(40) Chatzivasileiou, A. O.; Ward, V.; Edgar, S. M. B.; Stephanopoulos, G. Two-Step Pathway for Isoprenoid Synthesis. Proc. Natl. Acad. Sci. U. S. A. 2019, 116 (2), 506–511. https://doi.org/10.1073/pnas.1812935116.
(41) Sharma, D.; Moirangthem, A.; Roy, S. M.; Kumar, A. S. K.; Nandre, J. P.; Patil, U. D.; Basu, A.; Sahoo, S. K. Bioimaging Application of a Novel Anion Selective Chemosensor Derived from Vitamin B6 Cofactor. J. Photochem. Photobiol. B Biol. 2015, 148, 37–42. https://doi.org/10.1016/j.jphotobiol.2015.03.021.
(42) Iwanami, M.; Numata, T.; MurakamiM. A New Synthesis for Pyridoxal-5’-Phosphate. Bull.
Chem. Soc. Jpn. 1968, 41, 161–165.
(43) Zelder, F. Recent Trends in the Development of Vitamin B12 Derivatives for Medicinal Applications. Chem. Commun. 2015, 51 (74), 14004–14017. https://doi.org/10.1039/c5cc04843e.
(44) Mundwiler, S.; Spingler, B.; Kurz, P.; Kunze, S.; Alberto, R. Cyanide-Bridged Vitamin B12-Cisplatin Conjugates. Chem. - A Eur. J. 2005, 11 (14), 4089–4095. https://doi.org/10.1002/chem.200500117.
(45) Zobi, F.; Blacque, O.; Jacobs, R. A.; Schaub, M. C.; Bogdanova, A. Y. 17e Rhenium Dicarbonyl CO-Releasing Molecules on a Cobalamin Scaffold for Biological Application. Dalt. Trans. 2012,
41 (2), 370–378. https://doi.org/10.1039/c1dt10649j.
(46) Lee, M.; Grissom, C. B. Design, Synthesis, and Characterization of Fluorescent Cobalamin Analogues with High Quantum Efficiencies. Org. Lett. 2009, 11 (12), 2499–2502. https://doi.org/10.1021/ol900401z.
(47) Martens, J. H.; Barg, H.; Warren, M.; Jahn, D. Microbial Production of Vitamin B12. Appl.
