Editing Vanillin (section)

==Production==
{{anchor|Chemistry}}

===Natural production===
Natural vanillin is extracted from the seed pods of ''[[Vanilla planifolia]]'', a [[vine|vining]] [[orchid]] native to Mexico, but now grown in tropical areas around the globe.  [[Madagascar]] is presently the largest producer of natural vanillin.

[[File:Glucovanillin_v2.svg|200px|thumb|right|[[Glucovanillin]], the β-{{sc|D}}-glucoside of vanillin]]
As harvested, the green seed pods contain vanillin in the form of [[glucovanillin]], its β-{{sc|D}}-[[glucoside]]; the green pods do not have the flavor or odor of vanilla.<ref name = walton>[[#walton2003|Walton 2003]].</ref>  Vanillin is released from glucovanillin by the action of the enzyme [[β-glucosidase]] during ripening<ref>{{cite journal | doi = 10.1111/j.1365-2621.1943.tb18011.x | title = ACTION OF a β-GLUCOSIDASE IX THE CURING OF VANILLA | date = 1943 | last1 = Arana | first1 = Francisca E. | journal = Journal of Food Science | volume = 8 | issue = 4 | pages = 343–351 }}</ref><ref>{{cite journal | title = Changes in vanillin and glucovanillin concentrations during the various stages of the process traditionally used for curing Vanilla fragrans beans in Reunion | author = Odoux, Eric | date = 2000 | journal = Fruits | volume = 55 | issue = 2 | pages = 119–125 }}</ref> and during the curing process.<ref>{{cite journal | doi = 10.1021/jf00058a019 | title = Determination of Glucovanillin and Vanillin in Cured Vanilla Pods | date = 1995 | last1 = Voisine | first1 = Richard | last2 = Carmichael | first2 = Lucie | last3 = Chalier | first3 = Pascale | last4 = Cormier | first4 = Francois | last5 = Morin | first5 = Andre | journal = Journal of Agricultural and Food Chemistry | volume = 43 | issue = 10 | pages = 2658–2661 | bibcode = 1995JAFC...43.2658V }}</ref>

After being harvested, their flavor is developed by a months-long curing process, the details of which vary among vanilla-producing regions, but in broad terms it proceeds as follows:

First, the seed pods are [[blanching (cooking)|blanched]] in hot water, to arrest the processes of the living plant tissues.  Then, for 1–2 weeks, the pods are alternately sunned and sweated: during the day they are laid out in the sun, and each night wrapped in cloth and packed in airtight boxes to sweat.  During this process, the pods become dark brown, and [[enzyme]]s in the pod release vanillin as the free molecule.  Finally, the pods are dried and further aged for several months, during which time their flavors further develop.  Several methods have been described for curing vanilla in days rather than months, although they have not been widely developed in the natural vanilla industry,{{efn|[[#dignum2001|Dignum 2001]] reviews several such proposed innovations in vanilla processing, including processes in which the seed pods are chopped, frozen, warmed by a heat source other than the sun, or crushed and treated by various enzymes.  Whether or not these procedures produce a product whose taste is comparable to traditionally prepared natural vanilla, many of them are incompatible with the customs of the natural vanilla market, in which the vanilla beans are sold whole, and graded by, among other factors, their length.}} with its focus on producing a premium product by established methods, rather than on innovations that might alter the product's flavor profile.

====Biosynthesis====

[[File:Vanillin Biosynthesis.gif|thumb|Some of the proposed routes of vanillin biosynthesis]]

Although the exact route of vanillin biosynthesis in ''V.&nbsp;planifolia'' is currently unknown, several pathways are proposed for its biosynthesis. Vanillin biosynthesis is generally agreed to be part of the [[phenylpropanoid pathway]] starting with {{sc|L}}-phenylalanine,<ref name="digital.library.unt.edu">{{cite journal | last1 = Dixon | first1 = R.&nbsp;A. | title = Vanillin Biosynthesis – Not as simple as it seems? | url = https://digital.library.unt.edu/ark:/67531/metadc279692/m2/1/high_res_d/Dixon%20vanillin%202011.pdf | journal = Handbook of Vanilla Science and Technology | year = 2014  | page = 292 }}</ref> which is deaminated by [[phenylalanine ammonia lyase]] (PAL) to form t-[[cinnamic acid]]. The [[Arene substitution pattern|''para'']] position of the ring is then [[hydroxylate]]d by the [[cytochrome P450]] enzyme cinnamate 4-hydroxylase (C4H/P450) to create ''p''-[[coumaric acid]].<ref name="nature.com">{{cite journal | last1 = Gallage | first1 = N.&nbsp;J. | last2 = Hansen | first2 = E.&nbsp;H. | last3 = Kannangara | first3 = R. | last4 = Olsen | first4 = E.&nbsp;C. | last5 = Motawia | first5 = M.&nbsp;S. | last6 = Jørgensen | first6 = K. | last7 = Holme | first7 = I. | last8 = Hebelstrup | first8 = K. | last9 = Grisoni | first9 = M. | last10 = Møller | first10 = L.&nbsp;B. | title = Vanillin formation from ferulic acid in ''Vanilla planifolia'' is catalysed by a single enzyme | journal = Nature Communications | year = 2014 | volume = 5 | page = 4037 | doi = 10.1038/ncomms5037 | pmid=24941968 | pmc=4083428| bibcode = 2014NatCo...5.4037G }}</ref> Then, in the proposed ferulate pathway, 4-hydroxycinnamoyl-CoA ligase (4CL) attaches ''p''-coumaric acid to [[coenzyme A]] (CoA) to create ''p''-coumaroyl CoA. [[Hydroxycinnamoyl transferase]] (HCT) then converts ''p''-coumaroyl CoA to 4-coumaroyl [[shikimate]]/[[quinate]]. This subsequently undergoes oxidation by the P450 enzyme coumaroyl ester 3’-hydroxylase (C3’H/P450) to give caffeoyl shikimate/quinate. HCT then exchanges the shikimate/quinate for CoA to create caffeoyl CoA, and 4CL removes CoA to afford caffeic acid. Caffeic acid then undergoes [[methylation]] by caffeic acid O-[[methyltransferase]] (COMT) to give ferulic acid. Finally, vanillin synthase hydratase/lyase (vp/VAN) catalyzes hydration of the double bond in ferulic acid followed by a retro-aldol elimination to afford vanillin.<ref name="nature.com"/> Vanillin can also be produced from vanilla glycoside with the additional final step of deglycosylation.<ref name = walton/> In the past ''p''-hydroxybenzaldehyde was speculated to be a precursor for vanillin biosynthesis. However, a 2014 study using [[radiolabelled]] [[precursor (chemistry)|precursor]] indicated that ''p''-hydroxybenzaldehyde is not used to synthesise vanillin or vanillin glucoside in the vanilla orchids.<ref name="nature.com" />

===Chemical synthesis===
The demand for vanilla flavoring has long exceeded the supply of vanilla beans. {{As of|2001}}, the annual demand for vanillin was 12,000 tons, but only 1,800 tons of natural vanillin were produced.<ref>[[#dignum2001|Dignum 2001]].</ref> The remainder was produced by [[chemical synthesis]]. Vanillin was first synthesized from eugenol (found in oil of clove) in 1874–75, less than 20 years after it was first identified and isolated. Vanillin was commercially produced from eugenol until the 1920s.<ref>[[#hocking1997|Hocking 1997]].  This chemical process can be conveniently carried out on the laboratory scale using the procedure described by [[#lampman1977|Lampman 1977]].</ref> Later it was synthesized from lignin-containing "brown liquor", a byproduct of the [[sulfite process]] for making [[wood pulp]].<ref name="#hocking1997|Hocking 1997"/> Counterintuitively, though it uses waste materials, the lignin process is no longer popular because of environmental concerns, and today most vanillin is produced from [[guaiacol]].<ref name="#hocking1997|Hocking 1997"/> Several routes exist for synthesizing vanillin from guaiacol.<ref>[[#vanness1983|Van Ness 1983]].</ref>

At present, the most significant of these is the two-step process practiced by [[Rhodia (company)|Rhodia]] since the 1970s, in which guaiacol ('''1''') reacts with [[glyoxylic acid]] by [[electrophilic aromatic substitution]].<ref>{{cite journal |author1=Fatiadi, Alexander |author2=Schaffer, Robert |name-list-style=amp |title=An Improved Procedure for Synthesis of {{sc|DL}}-4-Hydroxy-3-methoxymandelic Acid ({{sc|DL}}-"Vanillyl"-mandelic Acid, VMA) |journal=Journal of Research of the National Bureau of Standards Section A |year=1974 |volume=78A |issue=3 |pages=411–412 |doi=10.6028/jres.078A.024|pmid=32189791 |pmc=6742820 |doi-access=free }}</ref> The resulting [[vanillylmandelic acid]] ('''2''') is then converted by 4-Hydroxy-3-methoxyphenylglyoxylic acid ('''3''') to vanillin ('''4''') by oxidative decarboxylation.<ref name="#esposito1997|Esposito 1997"/>
: [[File:Synthesis vanillin 4.svg|600px]]

Although guaiacol can be obtained by pyrolysis of wood, the type intended for vanillin production is mainly produced by petrochemistry.<ref name=Bomg/><ref name="#hocking1997|Hocking 1997"/>

====Wood-based vanillin====
15% of the world's production of vanillin is produced from [[lignosulfonates]], a byproduct from the manufacture of [[cellulose]] via the [[sulfite process]].<ref name="#hocking1997|Hocking 1997"/><ref name="Fache et al 2015"/> The sole remaining producer of wood-based vanillin is the company [[Borregaard]] located in [[Sarpsborg]], [[Norway]].<ref name=Bomg/> For this kind of use, softwood is preferred because there are more guaiacyl units convertible to vanillin.<ref name="#hocking1997|Hocking 1997"/>

Early production of wood-based vanillin would involve four plants: a sulfite pulp mill, a fermentation plant, a vanillin plant, and a Kraft (sulfate) pulp mill. The sulfite mill provides the brown liquor to the fermentation plant, which makes use of the residual sugar. The spend liquor is sent to the vanillin plant, which uses alkaline oxidation with air at 160–170 °C and 10–12 atm pressure, toluene extraction, and back-extraction with NaOH to obtain a crude sodium vanillate. Addition of [[sulfurous acid]] affords easy separation of the soluble sulfide addition compound of vanillin from insoluble impurities such as [[acetovanillone]]. The vanillin is extracted, and the remaining liquor is sent to the Kraft mill for burning to recover energy and sodium sulfide, both important for a Kraft mill.<ref name="#hocking1997|Hocking 1997"/> This process went out of favor in North America due to the large amounts of caustic liquids that needs to be disposed by the mill at the end: 160&nbsp;kg for every 1&nbsp;kg of vanillin produced. The recovery of sodium sulfide also became less and less profitable as the sodium-to-sulfur ratio became more and more unbalanced.<ref name="#hocking1997|Hocking 1997"/>

Borregaard is able to keep operating because it runs its own pulp mill. They have improved a process from Monsanto by using ultrafiltration<ref name="#hocking1997|Hocking 1997"/> to concentrate the incoming [[lignosulfonates]], which reduces the amount of NaOH used and waste produced. The basic chemistry is unchanged: alkaline oxidation using a metal catalyst such a copper salt.<ref>{{cite web |last1=Evju |first1=Hans |title=Process for preparation of 3-methoxy-4-hydroxybenzaldehyde |url=https://patents.google.com/patent/US4151207A |date=24 April 1979}}</ref><ref>[[#bjorsvik1999|Bjørsvik and Minisci 1999]]</ref> According to ''Scientific American'', vanillin produced this way contains armoatic impurities that add strength and creaminess to its flavor.<ref name=Bomg/> This is probably due to [[acetovanillone]] being present.{{efn|name=lig-vanillone}}

===Fermentation===
The company Evolva has developed a genetically modified yeast which can produce vanillin. Because the microbe is a [[processing aid]], the resulting vanillin would not fall under U.S. GMO labeling requirements, and because the production is nonpetrochemical, food using the ingredient can claim to contain "no artificial ingredients".<ref name=Bomg>{{Cite web|last=Bomgardner|first=Melody M.|date=2016-09-14|title=The Problem with Vanilla|url=https://www.scientificamerican.com/article/the-problem-with-vanilla/|access-date=2020-10-19|website=Scientific American|language=en}}</ref> The biosynthetic process starts with glucose, or any sugar that can be converted into erythrose 4-phosphate (which leads to 3-dehydroshikimic acid).<ref>{{cite web |last1=Hansen |first1=Joergen |last2=Hansen |first2=Esben Halkjaer |last3=SOMPALLI |first3=Honey Polur |last4=Sheridan |first4=Joseph M. |last5=Heal |first5=Jonathan R. |last6=Hamilton |first6=William D. O. |title=WO2013022881A1 Compositions and methods for the biosynthesis of vanillin or vanillin beta-d-glucoside |url=https://patents.google.com/patent/WO2013022881A1/en |language=en |date=14 February 2013}}</ref> The end product is 98% pure and is also considered natural in the EU.<ref name=Agostini>{{cite web |last1=Agostini |first1=Francesca Degli |title=Sustainable natural vanillin production through yeast fermentation - Europabio |url=https://www.europabio.org/sustainable-natural-vanillin-production-through-yeast-fermentation/ |date=3 October 2024}}</ref>

Using [[ferulic acid]] (a chemical found in rice) as an input and a specific non GMO species of ''[[Amycolatopsis]]'' bacteria, vanillin can be produced. Many other bacteria, either GMO or non-GMO, can be used for the same purpose. However, because vanillin inhibits the growth of free-floating bacteria, yields have been low. This can be overcome through the formation of [[biofilms]], which has been done with the non-GMO ''[[Bacillus subtilis|B. subtilis]]'' strain CCTCC M2011162.<ref>{{cite journal |last1=Yan |first1=Lei |last2=Chen |first2=Peng |last3=Zhang |first3=Shuang |last4=Li |first4=Suyue |last5=Yan |first5=Xiaojuan |last6=Wang |first6=Ningbo |last7=Liang |first7=Ning |last8=Li |first8=Hongyu |title=Biotransformation of ferulic acid to vanillin in the packed bed-stirred fermentors |journal=Scientific Reports |date=6 October 2016 |volume=6 |issue=1 |page=34644 |doi=10.1038/srep34644|pmid=27708366 |pmc=5052561 |bibcode=2016NatSR...634644Y }}</ref> However, using ferulic acid as the starting material does not qualify for "natural ingredient" in the EU.<ref name=Agostini/>

Biotransformation of eugenol (from cloves) into vanillin by non-GMO microorganisms has also been reported.<ref>{{cite journal |last1=Singh |first1=Archana |last2=Mukhopadhyay |first2=Kunal |last3=Ghosh Sachan |first3=Shashwati |title=Biotransformation of eugenol to vanillin by a novel strain Bacillus safensis SMS1003 |journal=Biocatalysis and Biotransformation |date=4 July 2019 |volume=37 |issue=4 |pages=291–303 |doi=10.1080/10242422.2018.1544245}}</ref> The same has been reported for guaiacol and guaicyl lignin (from conifers).<ref>{{cite journal |last1=Xu |first1=Lingxia |last2=Liaqat |first2=Fakhra |last3=Sun |first3=Jianzhong |last4=Khazi |first4=Mahammed Ilyas |last5=Xie |first5=Rongrong |last6=Zhu |first6=Daochen |title=Advances in the vanillin synthesis and biotransformation: A review |journal=Renewable and Sustainable Energy Reviews |date=January 2024 |volume=189 |pages=113905 |doi=10.1016/j.rser.2023.113905|bibcode=2024RSERv.18913905X }}</ref><ref>{{cite journal |last1=Zuo |first1=Kangjia |last2=Li |first2=Huanan |last3=Chen |first3=Jianhui |last4=Ran |first4=Qiuping |last5=Huang |first5=Mengtian |last6=Cui |first6=Xinxin |last7=He |first7=Lili |last8=Liu |first8=Jiashu |last9=Jiang |first9=Zhengbing |title=Effective Biotransformation of Variety of Guaiacyl Lignin Monomers Into Vanillin by Bacillus pumilus |journal=Frontiers in Microbiology |date=11 May 2022 |volume=13 |doi=10.3389/fmicb.2022.901690|doi-access=free |pmid=35633711 |pmc=9130762 }}</ref> These starting materials do not qualify for "natural ingredient" in the EU.<ref name=Agostini/>