Current Process Chemistry: Improved Isolation from Enzyme in Enzyme Catalyzed Organic Process Steps
kilomentor | 10 July, 2012 11:50
A topic that was much discussed at the recent Current Progress in Process
Chemistry meeting in Princeton June 13-14th was enzyme catalysis of
pharmaceutical process steps. Kilomentor has written about this subject before:
The essential message of this earlier blog article of mine was
that for some common chemical transformations there are now so many different
enzyme catalyst variants available that it is an odds-on bet that the chemist
will be able to find one that can do such steps. These transformation comprise such
things as ketone reductions, hydrogenation, P-450 type oxidations, dehydrations,
esterifications, amidations, epoxidations, andnitrile hydrolysis. This list is not exhaustive. At the June
Current Process chemistry meeting, Jacob
Janey PhD, a Senior Research Chemist with Bristol Myers Squibb, gave a
presentation titled Biocatalysts: Removal and Tracking during API Production. This
talk addressed the disadvantage that, even though these reactions proceeded
usefully, at the end of the reaction period when the protein catalyzing a
reaction in water/ 60% organic solvent gets denatured, the mixture often turned
the consistency of Jello making the isolation from the biocatalyst residue
tedious. Dr. Janey reviewed the accepted methods for these isolations:
centrifugation, evaporation followed by solid extraction, filtration,
extraction and immobilization of the catalyst. He then listed the disadvantages
of each method.
Centrifugation is typically only appropriate for small scale work because of
the equipment. Also it is slow and decanting the top phase can be tedious.
The evaporation followed by solid extraction is performed by diluting the
mixture of water and organic solvent with either acetonitrile or IPA and
repeated evaporating to remove all the water as an azeotrope, then
concentrating down to an organic slurry, filtering, and passing the organic
layer through silica gel. The disadvantages here are that once again it is
limited to small scale, it can be slow since the processing is long, and the
filtration can still be a problem.
The filtration method is applicable to reaction
mixtures with 2-50% organic cosolvent with water. First 200-400% Solka-Floc® (powdered
highly purified functional cellulose ) is added, then the mixture is acidify to
pH below 3, then agitated and heated to about 50 C to denature the enzyme. The
resultant is an aqueous precipitate that
can be filtered through cloth and rinsed through with 0.1 N HCl. This method
has the disadvantage that it only works with aqueous acid soluble substrates
(amines). It can also be a very slow filtration and the enzyme residue may
break through into the filtrate to some extent. It is not be general for all
enzymes. The advantage is that it is applicable on scale when it is applicable
However, this talk was about the advantages of the extraction procedure and how
it could be usefully adapted.
The method applies to an enzymatic transformation that has been conducted in
water with 2-50% of an organic cosolvent such as methanol, DMSO or isopropyl
alcohol (IPA). First, the pH is adjusted (if necessary) so that the substrate will
be extractable into an essentially organic layer when it is introduced. Then, 0.5
relative volumes of alcohol (IPA or MeOH) is added. Then 0.5 relative volumes
of isopropyl acetate or methyl t-butyl ether is added. The order of addition of
these liquids is noted to be crucial. Then moderate agitation is employed to
cause phase mixing to even out the concentration of liquids in between the physical layers and then allow phase
separation. The goal here is to achieve a 1:1:1 v/v/v mixture of water, alcohol
and aprotic organic before the two layers separate. The bottom, more
essentially organic, layer is then backwashed with 0.5 relative volumes of 30:70
v/v alcohol/aprotic organic. The upper, more
essentially organic wash is combined with the first upper, essentially organic
layer and after being combined together they are in turn washed with water. The
product is to be found in this wet combined alcohol/aprotic organic layer. The
advantages of this method claimed are that it is scaleable, gives reliable
easily visible quick separating cuts, and is reliable and adjustable to the
particular process by adding more or less alcohol as needed to give best
separation of the layers. I would judge this makes it the most general and scalable
isolation procedure available. The admitted disadvantages are that significant product
losses arise for relatively more polar substrate molecules. These are held more
strongly in the water rich layer and so are lost to a greater degree. The
method is of necessity more volume intensive and can be more lengthy since more
additions are made to achieve superior layer separation.
The method seems to this author to be based on a method of separating an
organic from an aqueous layer in the presence of insoluble materials that was
patented by Reuel Shinnar and Roberto Mauri for The Research Foundation of the
City University of New York in US 5,628,906. The patent authors say that “[T]he
inherent advantage of this method is that it works effectively even in the
presence of substances (solid or dispersed) that cause the formation of
emulsions or stable dispersions. Though the effect of adding a modifier on the
solubility of a solvent mixture is known, that such addition can result in
rapid phase separation even in the presence of emulsion forming impurities has
not heretofore been observed in connection with an extraction process”. I provide
this reference to fully inform my readers. I have no way of knowing whether Dr.
Janey knew of this intellectual property
or whether Bristol Myers Squib consider it relevant to this method for
separating denatured enzyme from the reaction mixture.
Current Process Chemistry Conference emphasizes Fit-for-Purpose Scaleup
kilomentor | 25 June, 2012 13:51
June 13 and
14th just passed I was
generously provided a Press Pass to attend the Cambridge Healthtech
Institute’s 6th International
Current Process Chemistry Conference in Princeton New Jersey.
This meeting is attended predominantly by very experienced scale up scientists
who usually are the team leaders of a group or the directors of a department
specializing in process scale up. The content is focused at a level suitable for persons who could write this
blog rather than those who are most likely to benefit from reading it!
Nevertheless, the general nature of the subject matter being discussed helps me
see the trends of the research big pharma and contract research organizations
are doing today and the types of background preparation upcoming process
development chemists and chemical engineers are going to need.
Most speakers provided a narrative description of the particular issues that
arose in the scale up for preparing a particular structure. The emphasis here
is on rapid, fit-for-purpose, scale up. This means that the processes were not really
optimized in the sense of using a multivariable statistic design to find the
overall best combination of all the variables.
Rather, in the typical instance, a processes was modified by
eliminating potentially serious risks
such as the potential difficult in removing metal impurities or immediately
vetoing potentially dangerous methodologies or making appropriate provisions to
deal with questions about late stage genotoxic impurities. (Issues relating to
genotoxic impurities came up regularly throughout the meeting. There are
certain functional groups which increase the likelihood that the intermediate
or reagent containing them will be found
genotoxic. Genotoxic substances cause
changes in the transmission of the genome and affect genetic inheritance and
must be held below very stringent impurity levels. They do not follow a dose
response relationship so there is no completely safe dose. Just developing
analytic methods showing that a process controls genotoxic impurities below
these levels is difficult. As a result scale up chemists often simply avoid
routes that use these late stage intermediates or reagents.)
The main concern of these specialists was as I said fit-for-purpose scale up. The
dominate concern was to provide the correct number of kilograms of product, of
the required purity, on the required date. The only long range requirement of
the method was that there should be no reason why the method could not be run
larger and more intensively to provide commercial quantities.
There wa very little mention of minimizing cost at this stage, for these
people, except the minimization of solvent usage where it reflects the
greenness of the process. Minimization of solvent use of course also helps
increase throughput and influences the time needed to deliver their kilograms
of drug candidate. It seems that the
cost reduction strategies that grind down the costs of API to the lowest level
possible are found more among the generic drug substance chemical community.
Another keen interest of these fit-for-purpose process chemists was identifying
and demonstrating the effect of critical process variables. It appears still very important for
developing process chemists to have an understanding of what constitute critic
process parameters. This is part of process validation. Kilomentor has written
an article about process validation from a process chemist’s perspective which
the emphasis of this conference seems to further recommend. http://kilomentor.chemicalblogs.com/55_kilomentor/archive/1375_the_importance_of_understanding_process_validation_for_pharmaceutical_process_chemists.html
Government regulators are interested in guaranteeing the public’s safety when
they take the actual medicine. Therefore, they are only interested that
manufacturers control variables which can deleteriously affect the quality
of the final drug substance. The companies that make drug substances, on the
other hand, are also very interested in
parameters that have an impact upon the cost of the drug substance. Although
the percentage yield of a drug
intermediate is may have no effect on the quality of the final product, a low
yield can and most likely will make the ultimate process uneconomic. Thus the
companies are very concerned about these parameters which affect the economics.
Parameters that do not affect the final product quality but do affect the
economics are called key parameters as opposed to critical parameters. Incidentally
regulators are interested in knowing about varying yields, even if the yield of a particular
intermediate is only a key parameter because it shows that there is at least
one significant uncontrolled variable in the process. Unidentified sources of
process variation are signs of inadequate process knowledge and potential
product difficulties in the future.
Separation of Diethyl Malonate and Monosubstituted Diethyl Malonate: Another Phase Switching Hdrolysis
kilomentor | 29 April, 2012 20:04
Synthesis Coll. Vol. II pg 279 there is a procedure for monomethylation of ethyl
malonate (diethyl methylmalonate). The crude, neat diester product is “shaken
for exactly one minute with a cold solution of 10 g. of sodium hydroxide in 30
cc. of water.” The corresponding note
reports that “[t]he ester is treated in this manner to remove any
unchanged ethyl malonate [diethyl malonate]. Michael1 has shown that this
treatment will completely remove unchanged ethyl malonate [diethyl malonate]
while hardly attacking ethyl methylmalonate. No ethyl dimethyl-malonate is
formed when methyl bromide is used as the methylating agent….A separation of
the desired product from traces of unchanged starting material and from ethyl
dimethylmalonate cannot be accomplished by distillation as the boiling points
of the three esters lie within three and one-half degrees of one another.” The
notes contain this additional information. “Michael found that unchanged
malonic ester can be removed completely by taking advantage of the greater ease
with which it is hydrolyzed by alkali ….”The references are Michael, J. Prakt.
Chem. (2) 72, 537 (1905) and Fieser and Novello, J. Am. Chem. Soc. 62, 1856
This appears to be an extraordinary
example of separation by competitive reaction using heterogeneous media. For
this to work the rate of hydrolysis of the starting material under these
conditions must be at least one hundred times faster than that for the
monomethyl product. Why would this be and how can other separations take advantage
of this? It might be that the deprotonation and extraction into the aqueous
basic phase of ethyl malonate in a vigorously agitated two phase mixture is
much faster for an unsubstituted ethyl malonate than for one that is
substituted, even with a very small group like methyl. It is very much less likely
to relate to a difference in solubility between the ethyl and ethyl methyl
malonates or even of their respective monoanions. Once ethyl malonate gets deprotonated and
carried into the aqueous phase it is probably quickly further hydrolyzed to
give carboxylates which keep it there. Kilomentor has given other examples of
such ‘phase switching hydrolysis’ separating esters where there is a free
phenol in only one component of a binary mixture and calls the method. http://kilomentor.chemicalblogs.com/55_kilomentor/archive/1011_extractive_and_phase_switching_hydrolysis_in_chemical_process_development.html
If the hypothesized explanation is correct this technique would be generally
applicable in any monoalkylation of ethyl malonate with a hydrophobic side
chain. It would mean that an excess of the inexpensive ethyl malonate could be
used in such alkylation to speed up the overall rate in the knowledge that the
excess ethyl malonate could be easily removed by alkaline hydrolysis and a
water wash. This would also allow the amount of dialkylation to be reduced to
About Extracurricular Training
kilomentor | 16 April, 2012 07:40
Vis-a-vis my last
post, I am responding to Bender’s questions. Whether one gets an interview for
a job depends both upon the degree of urgency that the employer faces and the
quality of the competition. Does reading kilomentor prepare one for a process
chem job better than actual experience in process chem? No. Definitely no. It
can supplement but not replace. Does it prepare one better than having no
process exposure at all? Definitely. Does it prepare a graduating chemist better than most of what is offered as process
chemistry in academia? I think so. I’m trying
hard. As it stands right now, I think kilomentor can replace academic training
in its present form but can’t replace process development experience. Kilomentor
can give new chemical graduates a leg up for an entry level position.
As far as the overall
usefulness of organic synthesis as preparation for other chemical functions, I
have seen throughout my career synthetic organic chemists switching with
outstanding success into other jobs- analytical, regulatory affairs, and in my
own case, solid dosage formulation. I have seen none move the other way. My
conclusion is that thinking about any chemical problem from a base perspective
of structure and reactivity has tremendous advantages. I entered solid dosage
formulation with no academic training; just several weeks working with an old
formulator, two days of those intensive classes, a textbook, and the chemical
literature! A synthetic background is the best preparation for any of these
jobs in my experience.
An Overview and Update of the Kilomentor Blog
kilomentor | 12 April, 2012 13:50
The Kilomentor educational blog is likely to have even more visitors starting next
week. A letter from the author appearing in the letters section of the Journal
of Chemical & Engineering News wil comment on the utility of blogs for
educational purposes. Therefore, a
review of the history, purpose, and perspective of the blog may be appropriate.
The Kilomentor blog was begun in 2006. The goal was to provide training and updating in
the methods for chemical process development, emphasizing scale up of organic
synthesis, particularly for high value pharmaceutical products. The author
recognized that there were textbooks, symposia, and courses for this purpose but
they were expensive and not equally available anywhere in the world. Moreover,
in academia, the treatment of chemical process development was neither
widespread nor generally thorough enough. The Kilomentor blog would be free, have post graduate content, and
be available wherever access to the world wide web was possible.
The particular perspective of this blog was that designing synthetic routes selection has become
easier than in in the past.
Today, ever since electronic sub-structure searching became readily available,
finding promising potential schemes has been simplified. What has not been made
easier is choosing between these hypothetical process routes and finding rugged/fast methods of
isolating process intermediates. Kilomentor has chosen to highlight techniques
that both work on scale and make isolating clean intermediates easier. The articles encourage
development chemists and chemical engineers to choose processes for scaling that comprise
more of such easily purified intermediates. It has suggested that routes containing
more of these rugged intermediates may even be preferable to processes with
fewer apparent steps. Thus routes that can have higher throughput tend to win
out over routes that just have fewer ‘ on paper’ transformations.
The Kilomentor blog has been well received. It has had more than 184,000 visits
since its start and there are now more than 115 separate blog articles. Visits, in 2012, have
reached about 200 per day. A search tool on the site will look for key
words and identify relevant articles.
Thanks for participating; and if you are new, then welcome!
A Large Rigid Acid for Making Crystalline Salts : 4’’-n-pentoxy-[1,1’:4,1’’]-terphenyl-4-carboxylic acid
kilomentor | 31 March, 2012 14:41
I have talked about a number of recyclable reagents, such as lasalocid, in the previous blog. The title compound to-day can be used either as a salt forming reagent or as an appendage or protecting group.
It has a structure containing three phenyl groups strung
together, end to end, through their para
positions with an n-pentyloxy cap at one terminal and a carboxylic acid
function at the other. All three rings are therefore para disubstituted. This
acid, after coupling with the echinocandin B macrocycle, gives a product
with excellent pharmaceutical properties. The substructure compound, as an
amide derivative, is part of the drug anidulafungin. Since this linkage would
be broken during the drug’s metabolism and since the drug has been found to be
safe, this would seem to establish that this acid component is fairly nontoxic
to the human metabolism.
This compound is a high melting solid and would be predicted to give
high melting salts and other covalent derivatives. For example, the esters and
amides are more likely to be solids using this rather than other carboxyl
functionalities. The rigidity of the carbon skeleton might be expected to cause
it to have a significant steric effect even remote to the point of attachment with
another substructure. The expense of this potential protecting group would be
ameliorated because the acid could be easily recovered and recrystallized to a
dependable purity for recycling. Its use might be particularly promising where
the process intermediates are expected to be liquids, oils, or waxes, such as in
Lasalocid and (-)-DAG: Practical Recyclable Chiral Acid Resolving Agents for Making Diastereomeric Salts.
kilomentor | 22 March, 2012 14:46
When performing a chiral resolution at scale it is important whether the resolving agent can be re-isolated,
crystallized to a consistent purity and thus
practically reused. When the resolving
agent is a carboxylic acid, this is simpler when the the carboxylate salt of an
alkali or alkaline earth metal is soluble in water while the free acid precipitates
from water. Two common chiral acids have this characteristic: lasalocid and
Lasalocid sodium is a veterinary pharmaceutical available in large quantities.
It is a chiral carboxylic acid that can be used to form diastereomeric salts
with racemic amines. Based on tested examples it is predicted to work most
dependably for primary amines that have their chiral centre at the alpha or
beta position as well as tertiary amines with a proximate chiral centre with
respect to the nitrogen atom. The ligand is capable of multipoint binding with
the amine as it forms hydrogen bonds to many different oxygens. The ligand
contains many different chiral centres. The molecule is made by fermentation.
The acid is relatively inexpensive. It was covered by US 4,129,580 which
expired in 1998.
acid hydrate also called (-)-DAG
(-)-DAG is also
a water insoluble chiral organic acid that can be used to resolve chiral
It is a
relatively inexpensive compound that is used an intermediate in the synthesis of Vitamin
C. It was first prepared by Reichstein
et al. Helv. 17, 311 (1934). Its use for resolution was taught in the expired
US patent 3,682,925 (1972).
More Discussion of Exhaustive Digestion in Chemical Process Development
kilomentor | 07 March, 2012 11:24
Myslinski has asked me to elaborate on the technique of purification by
‘exhaustive digestion’. This is a phrase that I used in my blog entitled, Crystallization
and Recrystallization Particularly On Scale. In this blog I make the point
that chemists scaling up a laboratory process often fail to do adequate
preliminary purification of a solid and as a consequence get a lower recovery
than is possible if a preliminary treatment is done. I propose ‘exhaustive
digestion’ as one pretreatment before recrystallization. I don’t know whether
there are any widely accepted differences between digestion and trituration but,
when used in my blogs, first, the noun trituration can be taken as applying
equally to oils or solids while digestion will only apply to solids. Second,
trituration can be taken to be done with ambient temperature or cold solvent
while trituration and particularly exhaustive trituration is more often done
with hot liquids, usually rather poor solvents for the desired product.
In the old literature, the progress of purification by digestion on the
laboratory scale is followed by observing the change in boiling point in the digesting
solvent or solvent mixture using a Beckmann thermometer, which is a thermometer
that can measure differences as small as one tenth of one degree. I don’t know
how long it has been since I have seen anyone use a Beckmann thermometer but,
at least in Phys. Chem., it was done quite often in my younger days. The range
of a Beckmann thermometer can be adjusted by removing or adding mercury to the
column of mercury used for measurement by moving mercury back and forth from an
attached mercury reservoir.
I will illustrate exhaustive digestion by quoting from an experiment taken from
Laboratory Technique in Organic Chemistry by Avery Adrian Morton, First Edition ,
McGraw-Hill, 1938, pg. 128-229.
“The apparatus consists of a tube or flask of such size that a large part of
the bulb is covered by the solid material being extracted , a thermometer
graduated in tenths of a degree, a reflux condenser, and a water or oil bath.
Place the sample in the container, insert the thermometer so the bulb is
immersed in or covered by the sample, and add solvent to cover the solid and
the thermometer bulb. The solvent is usually one that will dissolve impurities
without appreciable quantities of the desired material. Petroleum ether or
ligroin is often suitable. A mixture of solvents may be employed without
effecting their utility in this experiment, as long as the composition of the
mixture is not varied . Reflux the solution [I think slurry is intended] using
a water or oil bath as a source of heat. After about 20 minutes the temperature
has reached a constant level. Usually no trouble will be experienced from
superheating as long as solid phase is present, although a little agitation
with the thermometer is sometimes needed. Filter either by pouring into a
Buchner funnel or by using a filter stick. Add more of the same solvent, reflux
once more, and observe the temperature after equilibrium conditions have been
reached. Continue the operation until the temperature of two or more successive
operations are identical. The more soluble impurities have now been removed,
and the solution contains only the pure compound or the compound with [less
soluble type] impurities that have not been entirely removed…. The operation
may be continued, if desired, until all the product has been dissolved. A
change of solvent [may be implemented at some point for further purification]…..
Constancy in the boiling points of successive portions constitutes further
evidence of the purity of purity of the material, whereas a drop in the boiling
point is evidence of the exhaustion of still another component. Usually the
final portions of exhaustive digestion are pure materials.”
In light of the current interest in PAT, I cannot resist including the short paragraph that followed the quote from
this 1938 text.
“Use the same apparatus to follow the progress of a reaction in which a solid
phase is present, e.g., conversion of a triarylcarbinol to the chloromethane
compound while under a mixture of petroleum ether and acetyl chloride.”
By continuously measuring and recording
the changes, in real time, of the boiling point without any sampling this is an
early application of a PAT technology. With the improvements in the size and
sensitivity of temperature measuring devices, this following of small changes
in boiling point under conditions where superheating does not occur perhaps
deserves more frequent consideration.
Exhaustive digestion if used at-scale would require some more rugged method for
measuring small changes in the boiling point of the slurry than A Beckmann
thermometer loaded with mercury. Stirring using the thermometer as described in
the above laboratory procedure of course out of the question. In a small amount
of a thick slurry, the volume could easily fall below the minimum stirrable
volume in the reactor so a plow type stirrer might be needed.
Allophanates- Potentially Useful Reversible Highly Crystalline Solid Derivatives of Alcohols
kilomentor | 28 January, 2012 15:44
In scale-up during chemical process development, Kilomentor has proposed that it is not just minimizing the number of synthetic steps that leads most often to the most cost efficient process, but more often the simplicity and ruggedness of the steps. This is because it is the time and other resources that get invested in the separation and purification functions rather than the reaction per se that contributes most to the overall cost. It is quite possible that preparing a solid derivative and then converting it back to the original functionality may have cost advantages and purity advantages.
Alcohols that are liquids at ambient temperature in particular may be better isolated as a solid derivative rather than as selected fractions from a fractional distillation. Similarly even if an alcohol is a solid, if it is low melting and present in a product mixture, a better yield of pure product may be available by making a derivative and then converting it back to free alcohol. No alcohol derivative introduces more polarity in terms of hydrogen bond donors and acceptors for the same small increase in molecular weight than the allophanate derivative.
The allophanate derivative ( C-O-CO-NH -CO-NH2) is formed by condensation of the alcohol function with two equivalents of isocyanic acid, O=C=NH (or which can also be represented as its tautomer cyanic acid).
The formation of the allophanate can be expected to increase the water solubility relative to the parent alcohol compound and decrease its solubility in organic solvents. Residual cyanuric acid formed during the preparation of allophanates is somewhat soluble in cold water but particularly in hot water. It is insoluble in cold methanol, ether, acetone, benzene and chloroform. Because of its acidity it: Ka1 6.31X 10-8 pKa1 7.20; Ka2 7.94 X 10-12 pKa2 11.14; it is quite soluble in alkaline media.
Except for those arising from lower alcohols, allophanate are well melting, highly crystalline compounds suitable for isolation . The compounds are easily recrystallized and the parent alcohol can be regenerated by warming the allophanate with methanoic alkali.
Depolymerization of cyanuric acid can be done at 360-400 C in a slow stream of carbon dioxide. The gas can be absorbed directly into the neat alcohol or the reagent can be absorbed in an organic solvent such as ether to create a 30-35% by weight solution.
Consider the hypothetical reduction of 4-methyl-3-penten-2-one, (mesityloxide), bp 129 C, by hydrogenation. There are three possible products that are alcohols: 4-methyl-2-pentanol, bp 132 C, (ketone and double bond reduced); 4-methyl-4-penten-2-ol, bp 131.7 C, (only ketone reduced): and 4-methyl-3-penten-2-ol, bp 132 C, (ketone reduced and double bond isomerized). It is likely that reaction conditions can be found that lead to substantially one desired substance but such an enriched mixture still could not be separated based on boiling points. A solution that could be considered is the formation of the allophanate derivatives and recrystallization to achieve the predominant compound pure followed by hydrolysis back to the parent alcohol.
The Prins reaction is expected to produce a 1,3-diol from formaldehyde and an olefin. This is not necessarily a clean reaction. In fact the infrequency of its application suggests that it may lead to multiple products. Formation of the bis-allophanates as a method to obtain a pure crystalline product seems to me to be interesting.
The literature suggests that allophanates are derivatives that can be expected to crystallize from even quite difficult mixtures. For example the method was useful in the isolation and purification of various vitamins from natural sources. Fieser & Fieser in Organic Chemistry, the Third Edition, Reinhold Publishing Company, 1956. “The isolation of two pure factors from wheat-germ oil concentrates in 1936 was simplified by the discovery of crystalline derivatives, allophanates, resulting from esterification of the factors with cyanic acid….. on hydrolysis of the derivatives, the two pure active factors were obtained as highly active pale yellow oils named alpha and beta tocopherol.”
Similarly the derivative was used by Windaus and coworkers in isolating Vitamin D3 from an irradiation mixture. This is reported in Fieser & Fieser’s, Reagents for Organic Chemistry Vol. 1 pg. 171:
“Treatment of the crude, oily mixture with isocyanic acid afforded directly a solid product easily purified by recrystallization from acetone and converted into pure vitamin by hydrolysis.” Vitamin D3 has mp 82-84 C while the allophanate had mp 173-174 C, so one can see the inherent advantage. The co-products of the hydrolysis are conveniently totally water soluble!
The Chemical Manufacturing Process Route Selection: The Inclusion of Phase Switching and its Relationship to Validation
kilomentor | 20 January, 2012 11:31
Neal G. Anderson in his monograph, Practical Process Research & Development, says nothing about validation but he does speak about the need “to freeze the final process early, that is, to identify early the most desirable final chemical transformation to prepare the drug substance.” He credits this concept to P. Shutts. [“Freeze the Commercial Process-Issues and Challenges” a talk contributed at the Third International Conference on Process Development Chemistry, Amelia Island, FA, March 26 1997]. Earlier in his book, Anderson says specifically: “In order to establish routine impurity profiles and levels, ideally the final isolation of drug substance should be optimized, and this process should be used for the preparation of material destined for toxicological studies and later Phase I studies. The types of impurities found in drug substance will be largely determined by the starting materials and reagents used in the final step to prepare the drug substance. Thus the ideal penultimate compound should be identified in investigations, and parallel development work should converge on this penultimate compound.” In this particular passage that the last step Anderson is talking about is not the formation of a pharmaceutical salt. Almost certainly he is talking about a covalent bond forming step that completes the final structure. Also Anderson is speaking about the ideal situation, but what this assumes is not just that the last step, from penultimate compound to actual API, does not change, but also that this final intermediate is so efficiently purified that the steps preceding do not contribute to its impurity profile other than as inconsequential trace substances such as would be below 0.1%. It would only be in such instance that different routes to this penultimate compound would not leave behind any of their own impurities in the drug substance. If this is translated into the vocabulary of process validation, this would declare that there should be no critical steps before the purification of the ultimate intermediate! This is indeed a truly ideal situation and would constitute an impractical goal for real-life penultimate compound purifications to routinely seek.
Even though Anderson does not actually use the word validation in his book, even if we accept that validation is the most significant economic outcome of a developed pharmaceutical process; Anderson does convey the idea that certain characteristics of the final process step that simplify validation, are very, very important.
To achieve this requires a very selective choice of the penultimate compound. It must be easily and efficiently isolated and purified. As Kilomentor has argued this suggests that the step in which it is prepared and worked up involve several phase switches, because it is phase switches that provide purification and itt is compounds containing acid and/or basic functionalities that provide the most common opportunities for simple phase switches.
A Question from the LinkedIn Organic Process Research & Development Group that Fosters Practicing Creativity
kilomentor | 14 January, 2012 15:24
A question was asked at the LinkedIn Organic Process Research & Development Networking Group http://www.linkedin.com/groupItem?view=&srchtype=discussedNews&gid=1902030&item=88459764&type=member&trk=eml-anet_dig-b_pd-ttl-cn&ut=07TQuSi7kXnl41 “How could one remove a small amount of ethoxytrialkylsilane from a solution of ethanol containing some water at a scale of a large commercial waste stream.
I do not know whether a good answer is possible for this question. What interests me is what instruction the question can provide to students of process chemistry about how one searches for possible constructive leads for research.What is the entire real question?
Larry Fertel who posed the problem says, “I've got a stream of ethanol contaminated with <0.09 wt. % of a low boiling, trialkylethoxysilane. Any ideas on how to remove this (adsorbtion, coordination with a flocculent, etc.)? Distillation is not feasible, as the b.p.s are similar. Note that this is on a commercial scale….There actually is 9% or so water in the stream, water content is not an issue….The problem/issue is that the silane has a b.p. within 2 deg C of the azeotrope, sorry for leaving that piece of info out….Let me say that the ethanol stream is many millions of gallons per month, so this is not a research project. The ethanol is a by-product of a commercial process, so replacing it is not an option. Any treatment must work within the economics of reselling the ethanol into the marketplace (i.e., can only cost so many cents/lb.) . One option we are looking at is to convert to ethyl acetate for reselling. We are considering carbon treatment, one can build large columns in parallel that can be used on a continuous basis and get switched around once the carbon gets saturated in the impurity (assuming it works)…. I work for a contract research company, we were approached by a customer with this issue (good for us ($$), bad for them!!) So we need to be the problem solvers. I worry about the kinetics of adding a small amount of a reagent to a small amount of impurity. Considering that it is a bimolecular reaction, wouldn't the rate be slow? Not sure of the spec needed for the final product, at this point we are at the "screening" level.”
The problem is that there is <0.09% of an unacceptable compound contaminating millions of gallons per month of ethanol/water which cannot be removed by distillation because the undesired trialkethoxysilane distils close to 78.2 C, the ethanol/water azeotrope.
The problem is not a solution of ethanol/water. It is the presence of the trialkylethoxysilane at the level of <0.09%. There must be some low level that is acceptable otherwise the problem is to reach an undetectable level at low cost and that is almost certainly impossible. We must assume that the actual level is around 0.09% and it needs to be substantially reduced to let us say 0.01%
The problem is that the process as designed gives this level. It is not that that the level is around 0.09% because of some deviation in the process. There was no time when the level was acceptable. The unacceptable level is always present in the exiting waste stream from the process.
To solve the problem we might argue that we cannot remove the ethanol/water from the stream. That is we might say we cannot act on the major content of the stream because that would be expensive but apparently that is not the case. Larry Fertel says that distillation is rejected because it does not work not because it is too expensive. This opens the possibility that if we could selectively convert the trialkylethoxysilane to something separable from ethanol/water it would constitute a possible solution. Similarly it would seem that somehow changing the distillation temperature of the silane would be a possible solution and it would be particularly advantageous if it were the impurity that vaporized at lower temperature. Adsorption or adsorbtion onto or into another phase would be advantageous and probably most preferred because the predominant ethanol-water would be unchanged.
The hydrolysis and adsorption ideas have been pretty well explored by Stone, Tyrell, and others in the LinkedIn discussion. Kilomentor would like to look at the distillation and reaction possibilities a bit more.
One way to increase the degree of separation between the points of distillation of the trialkylethoxysilane and the ethanol/water is to add something to the mixture which will enhance the rate of vaporization of the more lipophilic material. This is the idea of extractive distillation. At the same time one does not want to leave anything behind in the ethanol/water that you are trying to decontaminate. Suppose we were to add an essentially immiscible perfluorocompound (expensive) or a much more volatile pentane (cheap). Would the vaporizing added compound sweep with it the undesired silane? I am reminded that there is a patent that claims the removal of traces of DMSO by co-vapor entrainment with ??.
It has already been suggested that a hydrophobic styrene-divinylbenzene cross linked resin might work. What about just using beads of solid paraffin? These would be cheap, sturdy and insoluble. They could be created by dropping molten paraffin into the stirred bulf solvent. They would float on the surface of the ethanol/water and could be filtered off and melted to drive off the impurity.
Remember guys, during the idea generation stage of problem solving criticism is supposed to be put on hold!
Now what can we get by playing with the idea of reaction of the impurity? What distinguishes the undesired impurity from the bulk? Molecular weight. The presence of silicon. The presence of a silicon oxygen bond. The presence of carbon-silicon bonds. The presence of silicon in the impurity seems to jump out at me. What is the strongest bond that silicon can form because that is what I want to do if I am going to drive this product into some other substance? The strongest silicon bond is one to fluorine. So perhaps the ion exchange resins that are being contemplated should have fluoride ion loaded on them. The conversion R3SiOEt + F- + H2O going to R3SiF + H-OEt + OH- would have an enthalpy advantage of about 27 kcal/mole. Hydrolysis in contrast has no enthalpy advantage, I think.
Anyway, problem solving is a combination of a good statement of the problem, a non-critical atmosphere for generating ideas, and most important well loaded retrievable associative memory banks. Note that you cannot come up with good ideas based on stuff you can’t recall but need to look up. Looking stuff up can be used to refine ideas but it cannot help to create them. You need to know things well enough that they can be recalled by associative memory when presented with the clues offered by the problem itself.
The Strategy of using Split Runs in Chemical Process Development
kilomentor | 23 December, 2011 15:25
Split run does not appear to be a widely understood term; however, in my personal chemical process development experience it is a common term with a well defined meaning. Split runs are experiments done to test different work-up/isolation and/or purification procedures starting with equal portions from a single large reaction mixture. As a consequence, even if a reaction mixture is both complicated and uncharacterized, a chemist can compare different follow-on procedures for working it up without being concerned about reaction differences that may still exist from batch to batch because the reaction process itself has not yet been finalized and critic parameters completely controlled.
In the general case, the isolated yield fraction, (which when expressed as a percentage is often called simply the percentage yield), is the product of the reaction yield fraction (determined by an assay of the crude reaction mixture before work-up) multiplied by the isolation yield fraction (the fraction representing the effectiveness of the recovery of pure product from the crude reaction mixture). In the case of comparing split runs, the ratio of isolated split run yields is proportional to the ratio of the isolation yields because the pure reaction yield is the same for each, since they come from the same reaction batch. Thus, in comparing the recovered product from split runs, the difference represents only the effectiveness of the isolation protocols….it is independent of and not confounded by the exact details of the reaction conditions. In this way, even at an early stage in developing the particular reaction step, the development chemist can get some idea of the comparative effectiveness of different treatments of the crude reaction mixture.
Of course it is not just the best isolation yield that is of interest and of importance to the process development chemist but also the relative purities of the two products and the amounts of different impurities within each.
The benefit in using split runs results because the optimization of the reaction conditions and the screening of potential isolation methodologies can proceed in parallel. It is not necessary to wait until the optimized processing conditions have been selected to work on the best isolation/purification means. The work can proceed more quickly and an earlier completion date can be set for some project milestones.
This does not mean that reaction optimization and isolation optimization can be completely decoupled. This can be seen to be intuitively obvious if one realizes that in an extreme case, if the assay yield in the reaction becomes 100%, there will be no residual starting material and no by-products formed and so this could simplify the work-up beyond all normal proportion and perhaps allow the most mundane isolation to give a fine outcome.
How thinking about Separating Acetic acid and Acetic Anhydride can sharpen our Chemical Process Development Skills
kilomentor | 16 December, 2011 10:50
Ganesh Wagh in the Chemical Process Development Group on LinkedIn asked the question, “What is the best way to separate a mixture of 60% acetic anhydride and 40% acetic acid to recover the acetic anhydride?” This solicited quite a few responses. The particular forum is now closed and although Kilomentor did contribute one suggestion to the forum I think the topic deserves some further discussion that doesn’t fit well into the question-comments format.
A plurality of people suggested either distillation of distillation under reduced pressure. These solutions may very well be the best if one has the proper fractional distillation equipment and it is not busy performing more valuable separations, but there is the implication in the question itself that, at least in the questioners hands, there was some difficulty with these solutions, since they are the first thing to come to a process chemist’s mind. Indeed Ganesh later noted that he did encounter problems in his hands.
Several people (Antonio Manka, Anton Frenkel, Marta Krawczyk, Pravinchandra Vankawala) suggested fractional distillation, either with some variation of the distillation condition for atmospheric pressure or under reduced pressure. I am not an expert on distillation. That is perhaps why it is not clear to me why distillation at reduced pressure would be expected to simplify separating these two liquids compared to an atmospheric pressure fractionation. Also, whatever the pressure under which the fractionation is done, these methods appear to require the complete removal of the acetic acid before the acetic anhydride distillation and that must leave some lost still bottoms and column residues that must subtract from the recovery. On top of this, I would imagine that fractional distillation must be slow and labour intensive. One suggestion, variation of the packing in the distillation column,which would change the height equivalent of a theoretical plate, is typically not readily accessible in a multipurpose plant particularly where the object is no more than reagent recycling.
Mark Bratt contributed the idea “Any thoughts as to a non-aqueous separation e.g. cyclohexane or methylcyclohexane with methanol or acetonitrile, with an organic base?” Although I don’t endorse trying a non-aqueous, two liquid extraction that uses methanol, which would be both miscible with and reactive towards acetic anhydride (this is almost certainly just a slip – it is true that methanol is very frequently a partner phase in such applications), the idea is suggestive of a type of strategy that is adapted and taken up by others later in the discussions. Perhaps we could simplify this idea. What would happen if we just added enough triethylamine to neutralize the acetic acid? Wouldn’t the triethylammonium acetate form a separate liquid phase aside from the acetic anhydride? Even if it didn’t, wouldn’t distillation of this mixture amount to an reactive distillation in which the acetic acid would be held back by its reversible interaction with the triethylamine allowing the acetic anhydride to distil over cleanly first? Acetic acid and triethylamine are reported to give a higher boiling azeotrope, with composition 69% acetic acid 31% triethylamine, that distils at 163 C. If triethylamine didn’t do the job what about neutralization with tributylamine?
Kees Hoek made a suggestionto add acetyl chloride that would, if it is worked, convert the acetic acid back to acetic anhydride thereby, in principle, multiplying the acetic anhydride recovery. My rough bond energy calculation for this reaction between acetyl chloride and acetic acid gives an enthalpy difference of close to zero so if the kinetics were practical it most likely would produce an equilibration of starting material and products. Driving out the hydrogen chloride would be expected to drive the equilibrium towards acetic anhydride according to LeChatelier. The usefulness might boil down ( pun intended) to how fast one can scrub hydrogen chloride. It might be attractive if one also had in the plant an alkaline waste that needed neutralizing.
Pedro Guivisdalsky, among other ideas, suggested adding toluene to form the binary lower boiling azeotrope with acetic acid that distils at 105.4 C. This is a variant on the extractive distillation idea whereby the acetic acid is made relatively more volatile. Waste toluene could be used and the toluene/acetic acid washed to recover the toluene.
John Knight, Scientific Director at Scientific Update, suggests neutralizing the acetic acid using the completely non-volatile basic ion-exchange resin. As he comments, this risks the exotherm that accompanies every mode of purification that features a neutralization. It would also involve bashing around a lot of insoluble resin that may be ground down and give some tedious still bottoms. Nevertheless, the core idea is that insoluble nonvolatile ionic exchange resins should come to mind when neutralization is considered in separations generally. Also, Dr. Knight again relates back to the idea of using some agent to convert the acetic acid to acetic anhydride. “What about phosphorus pentoxide”, I am musing to myself? The coproduct will be non-volatile and can be rinsed out of the still-pot after careful hydrolysis.
Many contributors to the discussion recommend looking at how acetic anhydride is made commercially as a pointer to an economic method. This is of course excellent advice since the cheapest starting material for acetic anhydride must be acetic acid. The methodology that is used for commercial product may of course be out of the question for recovering usable acetic anhydride in a multipurpose plant where the cost reduction for an accompanying process may be the actual motivation for the question.
Ilya Avrutov seems to have garnered the most endorsements for his suggestion that the acetic acid in the mixture could be made non-volatile by treatment with sodium carbonate, preferably anhydrous sodium carbonate. Others would include a drying agent. All understand the problem that the carbonate leads to water that must be taken into account. Ilya’s concept is another variant of a non-aqueous neutralization like that of Mark Bratt, above; it is simplified by not requiring further organic solvent additions but complicated because the coproduct of his neutralization can react with acetic anhydride to regenerate acetic acid.
This in turned stirs up another pertinent question. Are there cheap agents that can neutralize acetic acid without producing something that would react with acetic anhydride? Two chemicals that come to mind are calcium hydride and calcium carbide. These substances should come to mind whenever one wants to generate calcium salts of carboxylic acids for isolation or derivatization. In this instance the coproduct with calcium hydride would be hydrogen gas which would be innocuous but needs to be treated carefully. Using calcium carbide the byproduct would be acetylene gas that needs to be recognized and handled safely. In each case exactly how these solids are contacted with the acetic acid/acetic anhydride mixture would need to be worked out carefully. I suggest adding either as a slurry in more acetic anhydride.
I think I can still take credit for the most off-the-wall suggestion in the discussion. It was based on a simple question. What is the relation between the water hydrolysis of acetic anhydride and pH? Couldn't one stir the mixture of acetic anhydride and acetic acid with an aqueous salt solution buffered at the most unreactive pH and quickly extract the acetic acid into the aqueous phase? Stirring should be gentle to reduce interfacial contact and the temperature should be kept low to minimize reaction, and of course, separation of the layers should be done promptly. The idea here is that the difficulties that often stop us from reaching a simple solution simply may be mistaken. Here we assume that acetic anhydride will react with water and make a water wash unthinkable. But how well does this reaction actual take place and can it be slowed down enough to perform washing? Remember the Hinsberg reaction of a sulfonyl chloride is performed in contact with an aqueous alkaline solution!
Anyway, there were lots of ideas proffered. Thanks to everyone who contributed. My apologies if I created any distortion in presenting your contribution. The discussion was at http://www.linkedin.com/groupAnswers?viewQuestionAndAnswers=&discussionID=83697798&gid=1902030&commentID=60647184&goback=%2Egmr_1902030%2Eamf_1902030_76918230%2Eamf_1902030_49383814&trk=NUS_DISC_Q-subject#commentID_60647184
If you want to blast me at length please send e-mail to firstname.lastname@example.org. Short blasts can go in the comments here at kilomentor.
An Example of a Possible Kilomentor Type Separation at Scale
kilomentor | 22 November, 2011 10:22
Thanks for the problem. Although you do not provide enough information to exactly identify the compounds that is OK. I understand there may be intellectual property or privacy issues. I think I can make some suggestions that are at least mentally stimulating and which certainly exemplify the kilomentor approach.
Marto has asked for suggestions of how to achieve a particular separation.
He wrote: “I am having a problem in the isolation of mixture containing biphenyl benzylacetate compound along with diarylated benzylacetate copound. I have been carrying out this reaction on 100 g- 200 g scale, the ratio of biphenyl and diaryl compounds is 10:1, isolation by recrystallization was unsuccessful. Both have similar retention factor in TLC. Suggest me any purification method for the isolation of these two compounds.”
First, I would hydrolyze the acetates. When compounds are strongly hydrophobic, separation should most often be tried on the most polar functional groups available.
In the case where there is a difference between the number of substituents ortho to the primary alcohol function, the compounds can probably be separated by the difference in the rates they form the O-sulfate derivatives. Sulfonate the mixture with only enough sulfonating agent ( say chlorosulfonic acid in one of the solvents pyridine or dimethylaniline; see the kilomentor blog on making O-sulfonates). The sulfonated material and the non-sulfonated are easily separated by acid-base extraction and the sulfonation is easily reversedto give two separated alcohols.
In the situations where there is no difference in the number of ortho substituents between the alcohols one must depend upon a difference in inductive and electronic effects and the best chance comes when the primary alcohol mixture is converted into two carboxylic acids. The required hydrolysis and oxidations of the esters can be done together without isolation and the mixture of two carboxylic acids can be obtained cleanly and easily by simple acid-base extraction.
I think there will be a good chance that these acids can be separated by what is called extractive crystallization. (kilomentor has also written a blog about extractive crystallization). Extractive crystallization works by taking advantage of any small difference in pKas of the two acids augmented by any small difference in the solubilities of the two acids in an immiscible organic solvent selected to augment this pKa difference. The goal is to get one compound overwhelmingly as a salt in water and the other compound overwhelmingly as free acid in the organic solvent.
In any case, even if this cannot be made to work, as free acids there are many more salts, both organic and inorganic, that are accessible to find substances that do fractionally crystallize.
Also when one has a mixture of acids one can screen a portfolio of enzymes to find one that esterifies only one of these acids. Thus one uses the most discriminating of reagents, enzymes, catalytically in their favorite reversible reaction, ester formation/ hydrolysis.
Once separated the alcohol-acetates can easily be reformed if that is what is required. The extra steps in both directions are simply probably high yield and the separations are trivial acid/base extractions.
If you are reforming the acetates remember that the ester (non alcohol) can probably be easily separated from any residual alcohol by treatment with lithium bromide or calcium bromide or calcium chloride in hexanes where the alcohol is likely to form an insoluble inorganic complex leaving the ester in hexane solution.
What Might be the Best Solvent for Difficult Cleaning Jobs on the Reactor Walls of a Plant Reactor
kilomentor | 24 October, 2011 07:36
The walls of a large scale reactor can sometimes be difficult to clean. The problem is compounded because they are not easily accessible and cannot be inspected closely. Methods that can be applied in the laboratory for many reasons are off limits. Scrubbing is impractical, dangerous, and potentially damaging to the equipment. What is needed is a powerful but innocuous solvent that can work by vapor condensation not just below the surface of the refluxing liquid cleaner but above the surface and on the reactor walls where the reaction mixture may have splashed, caked and baked.
In the very old literature a common, inexpensive, and innocuous compound was claimed to be the best solvent known and one that would dissolve both organic and inorganic materials; salts as well as uncharged covalent molecules. It was molten boiling acetamide. Acetamide can be synthesized in situ in the reactor by heating ammonium carbonate and acetic acid and distilling out water. This in fact is the first preparation in the First Collective Volume of Organic Synthesis. Acetamide has bp760 222.0 C ; bp100 158 C; bp40 136 C; bp20 120 C; bp10 105 C; or bp5 92 C. As a white solid it has mp 82.3 C. The solubility is 2 grams per ml of water. Acetamide has been advocated as a “green” solvent [http://acs.confex.com/acs/green07/techprogram/S3384.HTM]The ninth edition of the Merck Index describes it as “Solvent; molten acetamide is an excellent solvent for many organic and inorganic compounds. Solubilizer; renders sparingly soluble substances more soluble in water by mere addition or by fusion.” Way back in 1933, Professor O.F. Stafford of the University of Oregon wrote that acetamide dissolved more different chemicals than any other known solvent. [J. Am. Chem. Soc., 1933, 55 (10), pp 3987–3988].
Process chemist sometimes forget that their responsibility is for the minimized overall cost of the process and this is much more than the chemicals only cost. The throughput per unit time is a major factor in the overall cost. That time includes the equipment cleaning time required between batch runs and between the end of one campaign and a new one for another product to be run in the same equipment. It makes little sense to invest extensive research efforts in reducing processing time when the same throughput efficiencies can be more easily achieved by reducing cleaning time between runs.
Many plants use a standard cleaning protocol implemented as an SOP. Only when it fails to remove all the contamination are special cleaning procedures resorted to. In some cases the standard cleaning or rinsing will even exaggerate the problem. For example, in the synthesis of adamantane described in Organic Synthesis Coll. Vol. III pg. 16-19, specific instructions are provided to avoid treating the vessel with water until acetone is used first to completely remove the tar.
The development chemists are the first to get an indication that unusual cleaning difficulties could arise after certain processing. Giving the plant scale-up people a heads-up and some suggestions will improve both teamwork and overall efficiency.