Spec ochimica Ac a Pa A: Molecula and Biomolecula Spec oscopy 290 (2023) 122303
A ailable online 31 Decembe 2022
1386-1425/© 2023 The Au ho s. Published by Else ie B.V. This is an open access a icle unde he CC BY-NC-ND license (h p://c ea i ecommons.o g/licenses/by-
nc-nd/4.0/).
A jou ney ac oss dopamine Me abolism: A o a ional s udy o DOPAC
Miguel Sanz-No o
a
, Lucie Kolesniko ´
a
b
, A an Insaus i
c
, Jos´
e L. Alonso
a
, Ike Le´
on
a
, Elena
R. Alonso
a
,
*
a
G upo de Espec ocopía Molecula (GEM), Edi icio Qui ima, Labo a o ios de Espec oscopía y Bioespec oscopía, Unidad Asociada CSIC, Pa que Cien í ico UVa,
Uni e sidad de Valladolid, 47011, Valladolid, Spain
b
Depa men o Analy ical Chemis y, Uni e si y o Chemis y and Technology, Technick´
a 5, 166 28 P ague 6, Czech Republic
c
Depa amen o de Química Física, Facul ad de Ciencia y Tecnología, Uni e sidad del País Vasco (UPV/ EHU), 48940 Leioa, Spain
HIGHLIGHTS GRAPHICAL ABSTRACT
•DOPAC (3-dihyd oxyphenylace ic acid)
is a ele an sca old in dopamine
me abolism.
•DOPAC was ans e ed in he gas phase
and p obed by high- esolu ion o a-
ional spec oscopy.
•Union o lase abla ion and o a ional
spec oscopy p o ides accu a e s uc-
u al da a.
•Th ee dis inc s uc u es o DOPAC ha e
been iden i ied by o a ional
spec oscopy.
ARTICLE INFO
Keywo ds:
Neu o ansmi e s
Con o ma ional pano ama
High- esolu ion spec oscopy
Ro a ional spec oscopy
ABSTRACT
DOPAC, a ele an sca old in dopamine me abolism, was p obed in he gas phase and in e oga ed by high-
esolu ion o a ional spec oscopy. He ein, h ee dis inc con o me s we e isola ed in a supe sonic je and
iden i ied o he i s ime h ough an examina ion o he end o he o a ional cons an s and he dipole
momen selec ion ules. Addi ionally, we examined he plausible elaxa ion pa hways o he low-ene gy con-
o me s o DOPAC, which helped us o claim he indi ec de ec ion o wo addi ional con o me s, p o iding
conclusi e expe imen al e idence o he lexible na u e o his biomolecule. The cu en in es iga ion sheds some
ligh on he di e ences be ween je -cooled o a ional expe imen s and ma ix-isola ion in a ed spec oscopy.
1. In oduc ion
Neu o ansmi e s a e molecules in ol ed in he chemical signaling
p ocesses in he b ain. G-p o ein–coupled ecep o s (GPCRs) in e ac
wi h he neu o ansmi e s and ansmi he in o ma ion h ough
chemical in e ac ions ha commonly occu h ough he “lock-and-key”
in e ac ion model. [1] In he accep o - ecep o in e ac ions o
neu o ansmi e s wi h GPCRs, i is assumed ha he molecules p esen
speci ic complemen a y geome ic shapes.
GPCRs ecep o s a e a la ge g oup o molecules in ol ed in di e se
biological unc ions, and hey a e one o he majo esea ch ocuses in
pha macology. [2–3] Conc e ely, one o he mos s udied GPCRs is he
dopamine ecep o , whose p ima y endogenous ligand is he dopamine
(DA) molecule. Dopamine is e y common in he e eb a e ne ous
* Co esponding au ho .
E-mail add ess: [email p o ec ed] (E.R. Alonso).
Con en s lis s a ailable a ScienceDi ec
Spec ochimica Ac a Pa A:
Molecula and Biomolecula Spec oscopy
jou nal homepage: www.jou nals.else ie .com/spec ochimica-ac a-pa -a-
molecula -and-biomolecula -spec oscopy
h ps://doi.o g/10.1016/j.saa.2022.122303
Recei ed 5 No embe 2022; Recei ed in e ised o m 27 Decembe 2022; Accep ed 28 Decembe 2022
Spec ochimica Ac a Pa A: Molecula and Biomolecula Spec oscopy 290 (2023) 122303
2
sys em. This ho mone and i s ecep o s a ec mo emen , emo ions, and
he ewa d sys em in he b ain. The e o e, i is no su p ise ha dopamine
and i s ecep o s a e in ol ed in many diseases, such as Pa kinson’s and
schizoph enia. [4–5] This has led o dopamine and i s me aboli es being
a new a ge o s udying and de ec ing di e en diseases. Nowadays,
he e is g owing a en ion on s udying DA me abolic pa hways (see
Fig. 1) [6–7] o unde s and dopamine- ela ed biological unc ions.
P ecise knowledge o he p ope ies o DA p ecu so s and me aboli es
becomes essen ial o disco e ing and de eloping new d ugs agains
se e al degene a i e diseases.
In dopamine me abolism, some s udies in mice and a s using b ain
mic odialysis samples combined wi h liquid ch oma og aphy ha e
demons a ed ha he p ima y me aboli es o dopamine a e 3-dihyd ox-
yphenylace ic acid (DOPAC) and homo anillic acid (HVA) (see Fig. 1).
[8] HVA does no p esen e y high biological ac i i y and may be
conside ed he las link o dopamine me abolism. Howe e , he DOPAC
molecule, one o he mos abundan in e media e in dopamine ca abo-
lism, is in ol ed in di e se biochemical eac ions. Fo example, in he
signaling p ocesses, some s udies showed ha DOPAC ac s as a dopa-
mine agonis in GPCRs and could ha e an an agonis ic e ec on GABA
ρ
1
ecep o . [9–10] I was also con i med ha he DOPAC molecule in-
e ac s wi h
α
-synuclein p o ein, he cen al molecule in ol ed in syn-
ucleinopa hy diseases. [11–13] Fu he mo e, ela ed o he DA
me abolism, he pa ien wi h Pa kinson’s and schizoph enia p esen s
al e a ions in he DOPAC molecule’s bio-p oduc ion h ough DA’s
me abolism. [14–15] Fo his eason, DOPAC is an excellen bioma ke
o iden i y he loss o dopamine gic neu ons. [16–17] All his indica es
ha he DOPAC molecule is no a negligible p oduc in dopamine
me abolism and is also in ol ed in se e al biological p ocesses and
diso de s.
Dopamine has been p ima ily s udied as he p incipal neu o ans-
mi e molecule, ba e and in he enzyme-ligand complexes. [18–20]
Besides, ad anced echniques using lase apo iza ion me hods ha e
shown i s in insic s uc u e in isola ion condi ions, p esen ing a e y
ich con o ma ional landscape wi h se en de ec ed con o me s. [21]
Mo eo e , ou g oup has made g ea e o s o epo s uc u al da a on
he ela ed L- y osine [22] and L-DOPA [23], and we plan o ex end hese
in es iga ions o ul ima ely co e he comple e me abolic ou e o
dopamine (see Fig. 1). In his con ex , li le is known abou he con o -
ma ional p ope ies o he DOPAC molecule. E en hough DOPAC is
expec ed o p esen simila con o ma ional lexibili y as dopamine,
expe imen ally, only one con o me was de ec ed in bo h a gon and
xenon c yogenic ma ices. [24] In-silico, he au ho s p edic ed he pos-
sibili y o de ec ing ou con o me s, so hey a ibu ed he non-
obse a ion o mo e con o me s o he molecula in e con e sion o
he mos s able one. The c ys al s uc u e did no b ing much in o ma-
ion abou i s con o ma ional landscape because hey o med hyd ogen-
bonded dime s in he c ys al la ice, cha ac e is ic o he ca boxylic
acids. [25].
The in insic con o ma ional p e e ences o DOPAC and he possible
in amolecula in e ac ions esponsible o hei s abiliza ions become
undamen al o unde s anding he biological media’s dopamine
me abolomics. In his con ex , we p esen a comp ehensi e s udy o
DOPAC using he combina ion o lase - apo iza ion, je -cooled o a-
ional spec oscopy, and high-le el densi y unc ional heo y (DFT)
calcula ions.
Fig. 1. Schema ic classical biosyn hesis and deg ada ion pa hways o dopamine. [26] We highligh in solid black squa es molecules ha ha e al eady been s udied in
ou labo a o y, while in g ey squa es, we ema k molecules cu en ly being in es iga ed, including DOPAC.
M. Sanz-No o e al.
Spec ochimica Ac a Pa A: Molecula and Biomolecula Spec oscopy 290 (2023) 122303
3
2. Me hodology
2.1. Theo e ical calcula ions
The unequi ocal con o ma ional assignmen o he obse ed o a-
me ic species is gene ally achie ed by di ec ly ma ching he expe i-
men ally de e mined o a ional cons an s wi h hose p edic ed
heo e ically o a se o candida e s uc u es. Al hough se e al se s o
heo e ical calcula ions a e al eady a ailable in he li e a u e [24] we
pe o med ou heo e ical compu a ions o es ima e o he pa ame e s
ele an o ou expe imen s, such as he elec ic dipole momen com-
ponen s, besides o a ional cons an s. Keeping in mind he i e possible
o a ions a ound single bonds shown in Fig. 2, a sys ema ic con o ma-
ional sea ch was conduc ed. In a i s s ep, we employed as molecula
mechanics me hods (MMFFs) wi h he “La ge scales Low Mode” and
Mon e Ca lo-based sea ch algo i hms in Mac oModel (www.
sch odinge .com) o loca e all possible minima ene gy s uc u es on
he po en ial ene gy hype su ace. Subsequen ly, each molecula
s uc u e was u he geome ically op imized a he B3LYP-GD3BJ/
aug-cc-pVTZ le el o he heo y (Gaussian 09 package [27]) wi h
G imme D3 dispe sion and Becke-Jonson damping aken in o consid-
e a ion, which is po ayed as he bes pe o ming le el o heo y o
dopamine- ela ed molecules (see Appendix A, Theo e ical benchma k
o addi ional de ails). Finally, equency calcula ions we e pe o med
o iden i y he s a iona y poin s as ue minima (all hei equencies a e
eal) and calcula e he Gibbs F ee Ene gies. The s uc u es o he i e
lowes -ene gy con o me s a e shown in Fig. 2, and hei spec oscopic
p ope ies a e collec ed in Table 1. We ollow he no a ion used in a
p e ious s udy o he s uc u ally simila neu o ansmi e dopamine o
label hese con o me s. [28] A combina ion o h ee symbols is
employed: a capi al le e G o A speci ies he gauche ( olded) o an i
(ex ended) a angemen o he −CH
2
COOH chain, he Roman nume als
indica es he ene gy o de (ΔE) o he di e en con o ma ions ela ed
o he posi ion o he cis −COOH g oup wi h espec o he di ec ion o
ca echol hyd oxyl (–OH) g oups, and subsc ip s a and b deno e he
di e se o ien a ions o he ca echol −OH g oups.
2.2. Expe imen al me hods
A comme cial sample o DOPAC (m.p. >127–130 ◦C) was inely
powde ed and blended wi h a comme cial binde . Then he mix u e was
p essed using a hyd aulic p ess o o m app oxima ely 2 cm long
cylind ical ods. Small amoun s o an adequa e binde (me hylcellulose,
5 %) a e necessa y o gi e enough consis ency o he solid od. A e -
wa d, we employed a Nd:YAG picosecond lase (355 nm, 10 mJ pe
pulse) o apo ize he sample. We employ a mo o con olle which al-
lows a DC mo o (O iel Mo o Mike 18074) o o a e and ansla e he
od up and down along he injec ion sys em o achie e homogeneous
exploi a ion o he od. Also, he epe i ion a e o he expe imen is se
o 2 Hz o p ese e good acuum condi ions in he acuum chambe and
educe sample consump ion. A e wa ds, he apo ized p oduc s we e
supe sonically expanded in o he acuum chambe o he spec ome e
using he pulsed low o a ligh ine ca ie gas (Ne a a backing p essu e
o 10 ba ), and hen p obed by mic owa e spec oscopy. In ou lase
abla ion b oadband chi ped pulse Fou ie ans o m mic owa e spec-
ome e (LA-CP-FTMW), [29] a 24 Gsamples/s a bi a y wa e o m
gene a o (Tek onix AWG7122B) c ea es a as chi p mic owa e pulse,
which is subsequen ly ampli ied using a 300 W a eling wa e ube
ampli ie (IFI, GT186-300) o mac oscopically pola ize he molecules a
equencies om 6.0 o 12.0 GHz. Up o 80.000 indi idual F ee Induc-
ion Decays ( ou FIDs on each al e cycle) we e a e aged in he ime
domain and Fou ie - ans o med o ob ain he equency domain spec-
um. We used a Kaise –Bessel window o imp o e he baseline esolu-
ion. The unce ain y o he line measu emen s was es ima ed o be
be e han 20 kHz. Also, since he sample injec ion is pe pendicula o
he mic owa e ield, he ansi ime o he pola ized molecula je is
ela i ely sho and we usually achie e line wid hs ( ull-wid h-hal -
maximum, FWHM) o app oxima ely 100 kHz.
3. Resul s and discussion
The b oadband CP-FTMW spec um o lase -abla ed DOPAC be ween
6 and 12 GHz is shown in Fig. 3. A ew lines co esponding o pho o-
agmen a ion p oduc s such as inylace ylene (CH
2
CHC
2
H) o inyl-
diace ylene (CH
2
CHC
4
H) we e quickly spo ed and elimina ed. The
emaining, ela i ely ich spec um was hen ca e ully o e iewed.
Acco ding o he alues o he o a ional cons an s in Table 1, all con-
o me s will show he cha ac e is ic pa e n o a nea -p ola e asym-
me ic op. Among he domina ing ea u es o he spec um, a
cha ac e is ic a- ype R-b anch p og ession assigned o K
a
=1 ← 1
ansi ion exempli ied in Fig. 3, could be easily ecognized and was
asc ibed o he i s o ame ic species labeled as o ame I. In he cou se
o he analysis, weake lines ollowing p ac ically iden ical spec al
pa e ns we e disco e ed on he igh side o a- ype R-b anch ansi ions
Fig. 2. The i e lowes -ene gy con o me s o DOPAC we e calcula ed a he B3LYP-GD3BJ/aug-cc-pVTZ le el.
M. Sanz-No o e al.
Spec ochimica Ac a Pa A: Molecula and Biomolecula Spec oscopy 290 (2023) 122303
4
o o ame I. These sa elli e lines we e a ibu ed o a- ype R-b anch
ansi ions o o ame II (see Fig. 3). The possibili y ha hese sa elli es
should be assigned o he ib a ionally exci ed s a e o o ame I could
be uled ou by he na u e o he expe imen ; his would be signi ican ly
depopula ed a he low empe a u e achie ed in he supe sonic expan-
sion. An i e a i e p ocedu e o i ings and p edic ions made i possible
o iden i y b- ype R-b anch and Q-b anch ansi ions o bo h o ame s.
Howe e , ansi ions obeying c- ype selec ion ules we e obse ed only
o o ame II. Once he o a ional lines o o ame s I and II we e
emo ed om he b oadband spec um, new se s o b- ype and c- ype
ansi ions a ibu able o o ame III we e ound (see K
a
=2 ← 1 p o-
g ession in Fig. 3). Finally, a- ype ansi ions we e p edic ed bu no
obse ed o o ame III. All o a ional ansi ions collec ed o hese
h ee o ame s we e submi ed o he analysis employing Wa son’s
asymme ic op semi igid- o o Hamil onian in A- educ ion and I
- ep-
esen a ion. Ob ained spec oscopic cons an s a e collec ed in he i s
h ee columns o Table 1. The measu ed expe imen al equencies a e
ga he ed in Tables B1 – B3 o Appendix B.
Ro a ional cons an s c i ically depend on he mass dis ibu ion o
each o ame , and hei alues a e usually conclusi e in he con o ma-
ional assignmen s. Howe e , an inspec ion o Table 1 shows ha he
obse ed o ame s ha e simila alues o he o a ional cons an s as
co esponding o con o me s wi h simila mass dis ibu ions. These
alues a e only consis en wi h hose p edic ed o he gauche con-
o me s GIa, GIb, GIIa and GIIb, bu he di e ences a e no su icien ly
signi ican o disc imina e be ween hem unambiguously. In cases like
his, one needs o call o ano he disc imina ing ool. We can p o i om
he ac ha he alues o he dipole momen componen s d i e he
o a ional selec ion ules. Hence, he obse a ion o nonobse a ion o a
gi en ype o o a ional ansi ion can be, in a o able cases, aken as a
de ini i e ool in he con o ma ional assignmen s. Ro ame III is he only
species o which no a- ype ansi ions we e obse ed. Acco ding o
Table 1, his obse a ion is only consis en wi h he p edic ed nea ly
ze o alue o
μ
a
dipole momen componen o con o me GIIa o which
o ame III should be assigned.
Ou o he wo emaining o ame s (I and II), h ee candida es s ill
Table 1
Expe imen al and p edic ed spec oscopic pa ame e s o he DOPAC con o me s.
Expe imen al Theo y
a
Pa ame e
b
Ro ame I Ro ame II Ro ame III GIa(II) GIb(I) GIIa(III) GIIb AIb
A /MHz 1998.53474 (67)
c
1932.52255 (43) 1979.58731 (59) 1940.9 1998.3 1977.7 1946.5 2236.6
B /MHz 521.91410 (19) 533.58405 (16) 525.42333 (30) 533.5 522.8 526.6 530.6 480.6
C /MHz 453.32765 (17) 455.20909 (19) 452.99409 (32) 455.4 453.7 454.0 456.9 438.2
Δ
J
/kHz 0.06255 (80) 0.04321 (86) 0.0397 (17) … … … … …
Δ
JK
/kHz 0.8478 (43) 0.3578 (47) 0.619 (12) … … … … …
Δ
K
/kHz 1.130 (67) … … … … … … …
δ
J
/kHz 0.01269 (34) … … … … … … …
δ
K
/kHz −0.586 (48) … … … … … … …
|µ
a
|/D Obse ed
d
Obse ed No obse ed 0.7 1.9 0.2 1.8 0.5
|µ
b
|/D Obse ed Obse ed Obse ed 0.4 0.9 3.2 3.6 2.1
|µ
c
|/D No obse ed Obse ed Obse ed 0.3 0.1 1.1 0.3 0.9
N 75 79 48 … … … … …
σ
ms
/kHz 3.5 6.4 8.6 … … … … …
ΔE / cm
−1
… … … 0 106 128 147 156
ΔG / cm
−1
… … … 108 100 168 166 0
a
Calcula ed a he heo y’s B3LYP-GD3BJ/aug-cc-pVTZ le el wi h G imme dispe sion.
b
A, B, and C a e he o a ional cons an s; Δ
J
, Δ
JK
, Δ
K
, δ
J
, and δ
K
a e he
qua ic cen i ugal dis o ion cons an s (A- educ ion); |
μ
a
|, |
μ
b
| and |
μ
c
| a e he absolu e alues o he elec ic dipole momen componen s along he ine ial axis a, b, c;
N ep esen s he numbe o dis inc equency lines in he i ;
σ
ms
is he oo mean squa e de ia ion o he i ; ΔE and ΔG ep esen he ela i e ene gy (including ZPE
co ec ion) and Gibbs ee ene gy (T =298 K), espec i ely, o he global minimum.
c
The numbe s in pa en heses a e 1
σ
unce ain ies in he las decimal digi uni s.
d
Expe imen al obse a ion o a gi en ype o ansi ion.
Fig. 3. B oadband o a ional spec um o 3,4-dihyd oxyphenylace ic acid measu ed by LA-CP-FTMW spec oscopy. Co esponding symbols highligh he mos
p ominen lines o each o he de ec ed o ame s. A p og ession o K
a
=1 ← 1 a- ype R-b anch ansi ions is illus a ed o o ame s I and II, while K
a
=2 ← 1b- ype
R-b anch p og ession is shown o o ame III. A decomposi ion line belonging o inylace ylene is also indica ed.
M. Sanz-No o e al.
Spec ochimica Ac a Pa A: Molecula and Biomolecula Spec oscopy 290 (2023) 122303
5
exi : GIa, GIb, and GIIb. No c- ype ansi ions we e ound in he eco ds
o o ame I, while he opposi e is ue o o ame II, which di ec ly
poin s o o ame I as con o me GIb. This assignmen can be u he
con i med by e alua ing di e ences in he alues o he o a ional
cons an s o GIb con o me using hose o he al eady assigned GIIa as
e e ence. These wo con o me s di e only in he opposi e o ien a ion
o he ca echol hyd oxyl g oups (see Fig. 4). This sub le s uc u al
change induces a small bu speci ic change in he alues o o a ional
cons an s. Hence, in going om GIb o GIIa con o me , he p edic ed
changes in o a ional cons an s a e ΔA = − 20.6 MHz, ΔB =3.7 MHz,
and ΔC =-0.3 MHz, and hese nicely ma ch wi h he obse ed changes
o ΔA = − 18.95 MHz, ΔB =3.51 MHz, and ΔC = − 0.33 MHz passing
om o ame I o o ame III, hus con i ming he iden i ica ion o
o ame I o GIb and o ame III o GIIa (see Fig. 4).
Ro ame II, which exhibi s all dipole momen selec ion ules, can be
asc ibed o ei he GIa o GIIb con o me s. The ques ion o which one
should be assigned canno be answe ed di ec ly. Once again, we in oked
he end o a ia ion in o a ional cons an s in Fig. 4. Con o me s GIIa
and GIa di e in he o ien a ion o he ca boxylic g oup, p oducing
dis inc i e shi s in o a ional cons an s ha can suppo he assignmen .
As shown in Fig. 4, he p edic ed changes in o a ional cons an s in going
om GIIa and GIa a e in good ag eemen wi h hose expe imen ally
de i ed in going om o ame III o o ame II, leading o he conclusion
ha o ame II is GIa. Mo eo e , in his case, he p edic ed |
μ
b
| and |
μ
c
|
dipole momen componen s o con o me s GIa (|
μ
b
|
GIa
=0.4 D and
|
μ
c
|
GIa
=0.3 D) and GIIb (|
μ
b
|
GIIb
=3.6 D and |
μ
c
|
GIIb
=0.3 D) a e
di e en . This ac is expec ed o gene a e a no iceable di e ence in
ela i e in ensi ies when compa ing he co esponding b- and c- ype
ansi ions o con o me s GIa and GIIb, espec i ely. Consequen ly, since
we obse e b- and c- ype lines o compa able in ensi y o o ame II i is
easonable o asc ibe i o con o me GIa, u he co obo a ing he
assignmen .
Finally, we pu sued he spec um analysis o sea ch o he o he low-
ene gy con o me s. A p io i, we could expec ha bo h GIIb and AIb
con o me s should be indeed p esen in he supe sonic expansion since:
a) The mo e ex ended con o me , AIb, is p edic ed o be he mos
abundan species a 298 K (a ending o ΔG), b) The abundance o GIIb
should be simila o ha o GIa, and also i s la ge dipole momen com-
ponen s should ease he con o ma ional sea ch. No e ha ela i e pop-
ula ions a he hypo he ical equilib ium o he lowes -ene gy con o me s
o DOPAC (a 298 K) ha e been calcula ed om he Gibbs ene gies a he
B3LYP/aug-cc-pVTZ le el, gi en in Table 1, o be GIa ≈19 %, GIb ≈20
%, GIIa ≈14 %, GIIb ≈14 % and AIb ≈32 %. Howe e , no spec al
signa u es a ibu able o he GIIb and AIb con o me s we e ound,
equi ing a deepe analysis. This beha io could be asc ibed o plausible
con o ma ional in e con e sion p ocesses, also known as con o ma-
ional cooling. I is well es ablished ha o je -cooled o a ional ex-
pe imen s, con o ma ional elaxa ion can occu h ough collisions wi h
he ca ie gas i he in e con e sion ba ie s a e low enough (abou ~
400 cm
−1
and lowe ), [29–32] as obse ed, o ins ance, o se e al
amino acids and dipep ides. [33–35] Thus, we ha e examined he co -
esponding in e con e sion ba ie s o es whe he he same p ocess is
occu ing. As shown in Fig. 5, calcula ed in e con e sion ba ie s be-
ween con o me AIb and con o me GIb, and be ween con o me GIIb
and AIb a e 22 and 190 cm
−1
, espec i ely, which helps us a ionalize
he absence o con o me s GIIb and AIb in he supe sonic expansion. A
his poin , we pe o med ela i e in ensi y measu emen s on di e en b-
ype ansi ions o con o me s GIa, GIb, and GIIa, since he in ensi ies
a e p opo ional o he squa e o he co esponding dipole momen
componen s. We obse ed a clea p edominance o con o me GIb
(abou ~ 55 %), which is in g ea disc epancy wi h he p edic ed
abundances based on ΔG. This poin s o a popula ion ans e om
ei he he AIb o GIIb con o me s in o con o me GIb, which appea s
en iched in he spec um, and help us o indi ec ly co obo a e he ex-
is ence o bo h AIb and GIIb con o me s in he je .
Addi ionally, he elaxa ion p ocess can be con i med by di ec
Fig. 4. Expe imen al and calcula ed changes in o a ional cons an s induced by he di e en o ien a ions o he ca echol hyd oxyl and cis ca boxylic g oups.
M. Sanz-No o e al.
Spec ochimica Ac a Pa A: Molecula and Biomolecula Spec oscopy 290 (2023) 122303
6
compa ison wi h he ela ed L-DOPA. [23] In con as o DOPAC, he
in oduc ion o a pola side chain (NH
2
g oup) in L-DOPA hampe s he
in e con e sion due o he o ma ion o s abilizing OH––N in a-
molecula hyd ogen bonds, which hinde s he o a ion o he
H
–
C
–
C
–
O dihed al angle. Hence, all he low-ene gy con o me s a e
de ec ed in he supe sonic expansion o L-DOPA, while only h ee
con o me s, which exhibi a cis con igu a ion o he ca boxylic g oup,
a e conclusi ely cha ac e ized o DOPAC. Ne e heless, cu en esul s
p o ide compelling e idence o he whole con o ma ional landscape o
DOPAC and comple e he p e ious s udy by ma ix-isola ion in a ed
spec oscopy, [24] whe e only he mos s able GIb con o me su i ed
upon ma ix deposi ion and was subsequen ly obse ed. Mo eo e , ou
esul s poin ou he no o ious di e ences be ween he cha ac e is ic
cooling o he supe sonic je , a ibu ed o collisions wi h he ca ie gas,
and he con o ma ional cooling co esponding o ma ix isola ion
echniques, [34,35] which exhibi a adically di e en na u e. Fo he
la e , con a y o je -cooled o a ional expe imen s, ba ie heigh s o
abou 400 cm
−1
a e no high enough o p eclude con o ma ional in e -
con e sion p ocesses. [23] The e o e, his con o ma ional elaxa ion
p ocess should be mo e accused du ing he deposi ion p ocess in he
ma ix as he highe -in-ene gy con o me s appea depopula ed in he
ma ix a e deposi ion. P esen esul s can be used as ele an bench-
ma k in o ma ion when compa ing bo h spec oscopic en i onmen s.
A compa ison be ween he con o ma ional landscapes o DOPAC, L-
DOPA, and dopamine b ings o ligh ele an insigh s. We obse e ha
dopamine- ela ed species exhibi a ela i ely wide con o ma ional
space. The b oade pano ama is ob ained o dopamine, whe e eigh een
con o me s a e p edic ed below 600 cm
−1,
and se en a e obse ed
expe imen ally, all o hem exhibi ing NH—
π
s abilizing in e ac ions. A
sligh ly na owe landscape o L-DOPA - se en con o me s p edic ed
below 600 cm
−1
and ou con o me s de ec ed (plus wo en a i e ob-
se a ions) -, DOPAC, wi h only i e con o me s p edic ed below 200
cm
−1
and h ee obse ed in he expe imen , and y osine, he la e
exhibi ing eigh p edic ed con o me s in he same ene ge ic window and
only wo a e obse ed expe imen ally. This leads o he ollowing end
o he numbe o obse ed con o me s (n): dopamine (7) >L-DOPA (6)
>DOPAC (3) > y osine (2).
As can be seen, y osine p esen s a a he poo con o ma ional space,
which is mos likely due o se e e pho o agmen a ion issues ound o
a oma ic amino acids ha usually make challenging he obse a ion o
hei comple e con o ma ional pano ama. Ne e heless, he o he
dopamine- ela ed species exhibi g ea lexibili y. Al hough, as
explained below, in he case o DOPAC he lack o an –NH
2
g oup allows
se e al con o ma ional in e con e sion p ocesses o occu , which will
be di ec ly ela ed o he obse a ion o a somewha na owe con o -
ma ional pano ama..
4. Conclusions
In he p esen wo k, we epo a s a e-o - he-a spec oscopic s udy
o DOPAC, a ele an dopamine me aboli e, o shed ligh on i s in insic
con o ma ional p ope ies. Th ee dis inc s uc u es ha e been un-
equi ocally iden i ied in he je , showing a cis con igu a ion o he
ca boxylic g oup in all cases. We ha e inally unleashed he en i e
con o ma ional space o DOPAC; wo o he obse ed con o me s, which
emained elusi e un il now, ha e been i e u ably de ec ed o he i s
ime. This in es iga ion p o ides ele an insigh s in o he h ee-
dimensional s uc u e o he isola ed molecule, p obing once mo e he
capabili y o high-quali y o a ional da a o un eil he shape o ele an
o ganics and biomolecules. Fu u e s udies will be ocused on he
in es iga ion unde he isola ion condi ions g an ed by he supe sonic
je o ye unexplo ed dopamine me aboli es (i.e., ad enaline and
no ad enaline), looking o wa d o ackling he whole me abolic ou e
o dopamine.
CRediT au ho ship con ibu ion s a emen
Miguel Sanz-No o: W i ing – o iginal d a , Concep ualiza ion,
Fo mal analysis. Lucie Kolesniko ´
a: W i ing – e iew & edi ing,
Concep ualiza ion. A an Insaus i: Fo mal analysis, W i ing – e iew &
edi ing. Jos´
e L. Alonso: W i ing – e iew & edi ing, Concep ualiza ion,
Resou ces, Funding acquisi ion. Ike Le´
on: Fo mal analysis, W i ing –
e iew & edi ing. Elena R. Alonso: Supe ision, Concep ualiza ion,
Me hodology, In es iga ion, Fo mal analysis, W i ing – o iginal d a .
Decla a ion o Compe ing In e es
The au ho s decla e ha hey ha e no known compe ing inancial
in e es s o pe sonal ela ionships ha could ha e appea ed o in luence
he wo k epo ed in his pape .
Da a a ailabili y
No da a was used o he esea ch desc ibed in he a icle.
Acknowledgmen s
The au ho s hank he inancial unding om Minis e io de Ciencia e
Inno aci´
on (CTQ2016-76393-P and PID2019- 111396 GB-I00), Jun a de
Cas illa y Le´
on (VA077U16 and VA244P20), and Eu opean Resea ch
Council unde he Eu opean Union’s Se en h F amewo k P og amme
(FP/2007- 2013)/ERC-2013-SyG, G an Ag eemen n. 610256
NANOCOSMOS.
Appendix A. Supplemen a y ma e ial
Supplemen a y da a o his a icle can be ound online a h ps://doi.
o g/10.1016/j.saa.2022.122303.
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