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`The Oxidation of Nicotine.
`
`I.
`
`Kinetics of the Liquid Phase Reaction
`Near Room Temperature
`
`Robert H. Lionel!
`
`Department of Chemistry. University at Vermont
`Burlington. Vermont. U.S.A.
`
`
`
`"[613286L0§
`
`(35°C. and 75% relative humidity)
`although ozddase is ‘datroyed. The
`heat treated tobacco absorbed Oz in-
`tensely initially but decreased with
`time whereas non-heat
`treated'to—
`bacco ruches a maximum 02 uptake
`rate after some time and then de—
`creases. Recently Wada(3) has oxi-
`dized nicotine at 30°C. with pure O2
`and found about 20% of-the nicotine
`oxidized in 4 weeks. He found am-
`monia, methyl amine, nicotinic acid,
`myosmine, cotinine, nicotyrine and
`unidentified products.
`
`Experimental
`Nicotine was vacuum distilled in
`3 Todd Column packed with glass
`helixes and the fraction at 97-98“
`(10 mm.) taken (reflux ratio 10:1).
`Frahly distilled, water—clear nicotine
`turns yellow in a few hours when
`exposed to air, but when sealed off
`in a glass ampule under vacuum it
`remains colorless for months. Pre-~
`liminary work was done—using an
`AMINCO Warburg apparatus at 20—
`40“ and with various shaking rates.
`It was found that with N, in the
`Warburg there was not volume
`change, with 0, there was a rapid
`uptake of gas which was roughly 5
`times the uptake rate using air: rates
`increased with shaking speeds and
`increased with temperature. Nicotine
`acidified with H01, H280.
`and
`HOAc did not have any 02 uptake
`activity indicating that the free base
`is involved in the oxidation. To ob-
`tain kinetic data an apparatus was
`constructed which consisted of a flat
`bottom flask of about 50 ml. volume
`fitted with a ground glass stopper
`for filling and a side arm which was
`connected to a 100 ml. gas burette
`
`Noven Ex. 1022
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`Page 1 of 3
`
`! agricultural
`sent level of
`\enL
`
`ecl
`
`reficients of
`Friction Be.
`- and Various
`Unpublished
`Agricultural
`-tate College,
`
`'I coeflicients
`l materials.
`ing 35 (6):
`
`-). Coeflicient
`Ir corn and
`ublished re
`—52Rl, Inter.
`-mpany, Chi.
`
`S. C. W. Sys-
`Bright Laf
`Yorth Caro-
`
`Neight and
`can Science
`
`
`
`
`
`
`
`
`introduction
`Although the biochemical degrada-
`tion and oxidation of nicotine has
`been extensively investigated (1), the
`kinetics of the reaction of oxygen
`with nicotine has not previously been
`
`reports ‘ that
`reported. 0babko(2)
`heating tobacco containing 25-28%
`H20 (in an inert N, atmosphere) to
`70.75°C. does not deprive the tobacco
`of its ability to bind 02 from air
`under fermentation conditions
`
`
`
`30
`
`20
`
`IO
`
`
`
`ML02once
`
`(Tobacco Science 89)
`
`TIME sec . IO'2
`Figaro I.
`
`IOO
`
`ISO
`
`
`
`Noven Ex. 1022
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`Page 1 of 3
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`

`

`
`
`
`
` an;AM‘
`-2:him
`o'er
`5.s
`-r
`'1‘.
`
`containing sat. NaCl-Hfi and a 3-
`way stop-cock for flushing the apps-
`ratus with gas. Oxygen gas at 1 at-
`mosphere pressure was used in all
`experiments. The reaction flask was
`mounted on a shaker which had
`variable amplimde and speed and was
`immersed in a water bath regulated
`to i 01°C. Experiments were per-
`formed at various shaking ampli-
`tudes and speed with nicotine sam—
`ples of 1.00-5.00 ml. A sample size
`of 2.00 ml. and shaking at 440 cyclm/
`minute with an amplitude of 7—8 mm.
`(measured at the sample) yielded
`rates of oxygen uptake that were
`independent of shaking for the tem—
`perature range of 15—40° used in this
`study. Nicotine stored in contact with
`air'decrensed in rate of oxygen up—
`take compared to firmth vacuum
`distilled nicotine Vacuum storage
`prevents aging but air contact dur-
`ing sample removal gradually aged
`the stored nicotine: this problem was
`solved by using Mathaon Prepuri—
`lied N2 (8 ppm 0,) to flush out the
`nicotine storage flask after each sam-
`ple removal and in this way identical
`mtas could be obtained from the
`same nicotine over a period of one
`week.
`Oxygen was bubbled into nicotine
`held at temperatures from 50-70°C.
`The nicotine rapidly turned brown
`and an infrared band at 5.9 microns,
`absent in pure nicotine, increased in
`intensity as
`the experiment
`con—
`tinued. The infrared band suggests
`cotinine is being formed(4). Oxida—
`tion for several days and then using
`Frankenburg‘s(4) technique for iso-
`lation of cotinine indimted complex
`reaction products. Addition of a free
`radical
`initiator, a,
`a'—
`'-isobu-
`tyronitrile (AIBN), in a similar ex—
`periment,
`resulted in more rapid
`darkening of the nicotine and simi-
`lar. if not identical, reaction prod-
`ucts. Addition of an antioxidant,
`4-methyl—2, 6—di-tert-butylphenol
`(BET), greatly decreased rate of
`coloration of nicotine on oxidation
`and inactivated nicotine to 0, for
`several hours.
`Several experiments were done on
`a burley tobacco containing about
`5% nicotine. Samples of the tobacco
`were heated in an atmosphere con—
`taining N2, air or 02 and the nico-
`tine was determined by the method
`of Cundifl(5). No change in nicotine
`content was found with N, at tan-
`peratures from 85-125°C. With 0,,
`the nicotine was reduced to one—half
`its original value in 300 hours at
`85‘, 24 hours at 110' and 3 hours at
`125°. The oxidized tobacco had a
`pleasant aromatic odor.
`
`f.
`'~.~‘.
`,w
`335:?
`‘.’3
` a“
`httl|egacy.library.ucsf.edultid/nnal4d00/pdf
`
`
`Table 1. linear Rates of Oxygen Absorbed by Liquid Nicotine
`
`ML0.(5r?)
`
`induction
`x lO‘
`period
`sec—ll x ml.
`Temp. °C. nicotine“1 sea): In“
`15
`1.08
`5.60
`
`20
`
`25
`
`1.35
`
`1.61
`
`2.46
`
`1.60
`
`i
`
`2
`
`‘
`
`I
`
`Remarks
`Nicotine stored
`24 hrs. under N2.
`Nicotine stored
`10 days under N2.
`Nicotine «fresh
`
`from vacuum dist.
`
`Run
`IX—ZO
`
`XIV-24
`
`V-18
`
`VII-19
`
`XIII-23
`
`XIV-25
`
`XVI—25
`
`XVII-26
`XII—22
`
`VIII-20
`
`X—21
`
`XI—21
`
`III—16
`
`111—16
`
`I'V-16
`
`No storage.
`Nicotine stored
`24 hrs. under N2.
`Nicotine stored
`10 days under N,.
`Nicotine stored
`14 days under N...
`Nicotine stored
`14 days under N2.
`Added 50 mg. 3111'
`Nicotine stored
`4 days under N2.
`Nicotine stored
`24 hrs. under N}.
`Nicotine stored -
`3 days under N1.—
`Nicotine stored -
`4 days under N2.
`Nicotine yellow
`from storage" ime
`month in poor
`vacuum.
`Reused nicotine
`from 11—16.
`immediately.
`Reused nicotine
`from III-16 24
`hours later.
`
`
`25
`
`25
`
`25
`
`25
`
`25
`30
`
`35
`
`35
`
`40
`
`25
`
`25
`
`25
`
`1.51
`
`1.59
`
`1.47
`
`1.54
`
`none
`1.73
`
`2.09
`
`2.10
`
`2.34
`
`1.21
`
`0.80
`
`0.18
`
`2.12
`
`2.22
`
`1.98
`
`1.70
`
`29
`1.58
`
`0.86
`
`0.88
`
`0.64
`
`2.10
`
`——
`
`—
`
`Results and Discussion
`In Figure 1 typical plots of vol-
`ume 0, used vs. time are shown.
`Table 1 tabulates all the results
`where rates are those from the linear
`part of the reaction. An Arrhenius
`plot of the maximum rata of table
`1 yields an activation energy of 6.81
`kcal (least squares). This is a low
`activation energy compared to the
`uncatalyzed oxidation of hydrocar-
`bons such as tetralin where the over—
`all activation energy is 14.9 i 0.6
`km]. from 65-130'C.(6) If the‘linear
`rates are extrapolated back to zero
`oxiy'gm uptake, and this time (allied
`the induction period tind. a plot of
`log (ll‘ind) vs 111‘ (assuming first
`order kinetics) would yield an activa—
`tion energy for the removal of in-
`hibitor (initiation). A value of 14.5
`keel. is found. The induction period
`could be due to formation and de-
`composition of a nicotine hydro-
`peron‘de but it is premature to as-
`
`ITobacco Science .90)
`
`sociate an activation energy with this
`proposed mechanism. A small appar—
`ent evolufion of gas in the initial
`periods of lower temperature rate
`studies is probably due to decreased
`gas solubility in the nicotine at lower
`temperatures:
`In the experiments reported here,
`2.00 ml. nicotine. at room temp, was
`used. corresponding to 1.88 x 104
`moles. For the consumption of one
`mole 0: per mole nicotine we would
`therefore require 421 ml. 02 at STP.
`Most experiments were run to a total
`0: consumption of 20-30 ml. (STP)
`and inhibition (deviation from linear
`rate) was noted at about 15 ml.
`(STP) 02 consumption. The low 0,]
`nicotine mole ratio at which inhibi—
`tion is found (about 0.03) together
`with inhibition by BHT and cam-lynx.
`by AIBN provides strong evidence
`of the free-radical nature of the en'—
`dation. This indicates nicotine oxi-
`dation follows the general mechanism
`
`
`
`ZATZRGIBG
`
`Noven Ex. 1022
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`

`

`of olefin oxidationfl) and a nicotine
`hydroperoxide may be expected as
`an intermediate. The proposed hydro-
`peroxide could be unstable at room
`umperature, yielding- H20,
`(which
`could form nicotine N-oxide). cati-
`nine, 320. and radicals which could
`mntinue the chain reaction and form
`complex polymers. Further work is
`underway to isolate the proposed
`hydroperoxide and provide more de-
`hils on the mechanism of the re-
`action.
`
`Aclmowlodgmenl'
`The author thanks the Tobacco
`Industry Research Committee which
`provided financial assistance making
`this work possible.
`
`Liferahlre Cii'ed
`
`1. McKennis, H. In, Tumbull, L. B..
`Wingfield, H. N. Jr, and Dewey,
`L. J., J. Am. Chem. Soc. 80, 1634
`(1958); ibid 6597.
`2. Obabko, V. A_, Vscsoyuz. Nanak.-
`Issledovaiel. Inst. Tabach. i Mak—
`harach. Prom. No. 142,
`105—72
`
`(1940); ibid. pp. 176-207. Chem.
`Abs. 36, 1439.
`3. Wade, E., Khaki, T. and Saito.
`K, Arch. Biochem. and Biophys.
`79, 124 (1959).
`4. Frankenburg, W. G. and A. A.
`Vaibekunas, J. Am. Chem. Soc. 79,
`149 (1957).
`5. Cunditf, R. H. and Markunas, P.
`C., Anal. Chem. 27, 1950 (1955).
`6. George, P.. Ridefl. E. K. and
`Robertson. A., Proc. Roy. Soc. 185,
`288 (1946).
`7. Bateman, L., Quart. Rev. 8, 147
`(1954).
`
`
`
`-' with this
`all tappar-
`he initial
`ture rate
`decreased
`2 at lower
`
`'ted here,
`211113., was
`38 x 10"
`n of one
`are would
`at STP.
`to a. total
`i. (STP)
`rm linear
`15 ml.
`low 0.]
`I inhibi-
`together
`:afalysis
`Evidence
`the oxi-
`Ene oxi.
`:hanism
`
`http:/lleg
`
`y.librel'y.ucsf.edulfid[nna14d00lpdf
`
`(Tobacco Science 551)
`
`€61286LOS
`
`l
`
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