C8953 NMR strukturní analýza seminář Identification of an unknown compound Jan Novotný, Ondřej Jurček 17 6003@is.muni.cz, ondrej.jurcek@ceitec.muni.cz April 18, 2018 □ if? Task 0: Classification of an unknown substance Assign the general name to displayed substances: CARBOHYDRATE PEPTIDE STEROID TERPENE ALKALOID COUMARINE □ g - g ► -El = ^00,0 Äri' 26, 2(f 7 02VC^ Task 0: Classification of an unknown substance A J AAftA A _aJa----_ J_I_J_ll juüilii 1 JAJ\_ I li ill i k_jlJ._,_JI_*L-j i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—r Ondřej Jurček J—i—I—I—I—I—I—I—I—I—|- m>rV 26, 2S17 •03%®' Task 0: Classification of an unknown substance Terpen e Carbohydrate Streroid Peptide Alkaloid JLii 1 ah A a MA Coumarine JÜU_LjU_L ~i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—r~ Ondrej Jurček ~I—i—i—i—i—i—i—i—i—i—i i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—i—r- -i—i—i—i—i—i—i—i—i— -1—i—i—i—i—i—i—r~ 3 2 1 ppm 3|= >OQ.O S1 m>rV 26, 2M7 O^V^ Task 1: Ci0Hi3NO 5[ppm] Multiplicity Integral 1.05 triplet 3 1.75 singlet 3 3.70 quartet 2 7-7.60 complex multiplet 5 Ondřej Jurček ftr\\ 26, 2(f 7 O*^^ Task 1: Ci0Hi3NO 5[ppm] Multiplicity Integral / \ ^ 1.05 a triplet 3 f / \-1\| C 1.75 c singlet 3 \v_/J \_ 3.70 b quartet 2 e d // 7-7.60 d-f complex multiplet 5 O Ondřej Jurček □ g - = _š = -O c\o ^>ri' 26, 2(f 7 O*^^ Task 2: CnHi5N02 5[ppm] Multiplicity -/(Hz) Integral 1.30 triplet 7 3 3.00 singlet - 6 4.25 quartet 7 2 6.65 dublet 8 2 7.80 dublet 8 2 ^>ri' 26, 2(f 7 O^C^ Task 2: CnHi5N02 IM \ / \\ / \=/\ U. ^ 5[ppm] Multiplicity -/(Hz) Integral \^ n-a O- 1.30 b triplet 7 3 e IM 3.00 e singlet - 6 4.25 a quartet 7 2 6.65 d dublet 8 2 7.80 c dublet 8 2 Ondřej Jurček ^>ri' 26, 2(f 7 O^C^ Task 3: Ci2H17NO - ^/COSY *H NMR (300 MHz, CDCI3) Ol CO H CD 1" Ifl Ol Cl 10 r| •3" CN O r- M tH CO U> CN O Cl Cl M N Cl (S H H Ol Ol in on in «o ri' 26, 2(f 7 o&^VC^ Task 3: Ci2H17NO - ^/COSY COSY (300 MHz, CDCI3) Jiá. Ondřej Jurček F2 (ppm) 1- 4 5 8 i i i i i i i i—i i i in a o » S1 I ► < 1 ► -11 = -O <\ O ^>ri' 26, 2(f 7 0«V(^ Task 3: d2Hi7NO - ^-"C/HSQC.HMBC HSQC (300 MHz, CDCI3) F2 (ppm) 1- I 1 l I I l I I (III I I I I I I I I I I I I I I I 1 [Ml I I I I I I I I lili I I I I i I I I lili lili 1111 140 120 100 80 60 Fl (ppm) 40 20 Ondřej Jurček ^>ri' 26, 2(f 7 OTty®' Task 3: Ci2Hi7NO - ^-"C/HSQC.HMBC HMBC (300 MHz, CDCI3) wméym^m^tm**mH WH—I F2 (ppm) 1- 2 3- 4 5 I I I I I I I I l I I I I I I I I I I I I I I I 1 Q o - i i i i i i ' l l f i I i I I l I I I I I 1 I I I I I I I I 1 1 1 I I I ! I I I I I I l l I l 1 I l l ' I I I I I I I I I 200 180 160 140 120 100 80 60 40 20 0 Ondřej Jurček ^>ri' 26, 2(f 7 OTty®' XH NMR (300 MHz, CDCI3) COSY(300 MHz, CDCI3) J lis .....i.........i.........i.........i.........i.........i.........i.........i.........i......... 7.7 7.5 7.3 7.1 ppm 2.6 2.4 2.2 2.0 1.8 1.6 1.4 1.2 ppm 0.92 1.08 1.84 1.00 2.08 2.05 2.91 2.94 2.02 F2 (ppm) 1- i i i i—i i i i—i i i i—i i i i JL_Lj_AJt_ f 1 ' ' 1 f 9876543210 Fl (ppm) Ondřej Jurček m □ g - = 3 = >OQ,0 ^>ri' 26, 2(f 7 O^O*' General comments • inspect molecular formula CmHh0oNnXx: Degree of unsaturation m + 1 — 0.5(h + x — n) • identify signals of CH3 and exchangeable protons in ID 1H sectrum • arbitrary numbering (e.g., from lower to higher value of chemical shift) of resolved resonances in all spectra • identification of the individual spin systems using DQF-COSY • resolve geminal protons using HSQC • connect molecular fragments/isolated spins using HMBC, NOESY o specify the stereochemistry (relative configuration) by means of J— and NOE interaction • in ID spectrum bottom blue numbers are integrals, labels in violet frames contains the arbitrary label (A-N), multiplet specification (use with caution, automatically determined), and position of a signal in ppm 9 UnHa-UnHb in 2D reffers to correlation of protons a and b of unknown compound Un < n ► < g ► < II ► < ^1 ► 1: ^Q^O ■3 QicV(> ID XH of Ci0H20O 1H ID ).8M DMSO 25CTXI 4.0 Jan Novotný Tr r 3.5 3.0 2.5 fl (ppm) —1— 2.0 —1— 1.5 —1— 1.0 —r 0.5 1H-13C HSQC and NOESY 20 i 30-\ & 40H O 50H 60-^ 70 UnC1-Ha UnC2-Hd > UnC3-H^^ UnC4 _ UnC5-Hk—-- j UnC6-Hh. UnC4-Hf UnC7-Hj [ 30 \ i UnC7-Hb UnC8-Hk UnC8-Hc \ \ / UnC9-Hg UnC10-Hm \ 20 40 50 [ 60 70 E Ä 2 3^ 4^ 4 2 C02-1H (ppm) 6 _i_ uhHa-Hn j_u _1_ UnHd-Hk UnHd UnHk-Hr \ UnHe-Hm Un„s_Hm UnHa-H, J UnHb-Hj^ UnHg-Hn UnHg UnHh-Hm V UnHI-H UnHk-Hm X j " 1 If unMD-j \/ UnHd-HL 1 / I/(/UrÚ / / UnHf-Hj yy |nUf*S / UnHg-Hľ V UnHf-Hi. UnHf-Hm y , . .1 UnHm-Hn « a e I I 4 C02- 1H (ppm) -r- 2 h3 Task 1: J-connectivity of C10H20O E o. 2 Q. 1111 1111 1111 1 1 1 1 1 . 1 1 1 1 UnHc-Hm UnHg-Hm UnHk-Hmi UnHb-Hj UnHb-Hh UnHJ-Hj -cl— IMHb-Hf UnHf-Hq/l ^* UnHe-říh' UnHf-Hi \ UnHf-Hb w 3H UnHm-Hn Jan Novotný □ S Task 1: J-connectivity of C10H20O • methyls la,2d connected to CH 51 O remaining crosspeak of CH 51 to CH 9g • methyls la,2d diastereotopic =>- chiral carbon 9 e a 2 4 ■ UnHg-Hm —r~ 4 UnHb-Hh i UnHb-Hj r UfHb-Hf 3 2 G)2-1H (ppm) r r Jan Novotný 4/5 Task 1: ./-connectivity of CioH2oO Task 1: J-connectivity of C10H20O • CH 9g has crosspeaks with deshielded 10m =4> OH group (n) 9 CH 9g has two crosspeaks with diastereotopic protons 4if H fH H f i e a 2 4 ■ tJnHg-Hm" nnTTm—TTn UnHb-Hh i UnHb-Hj r UfHb-Hf UnHil-Hj 3 2 G)2-1H (ppm) r f * * Jan Novotný 4/5 Task 1: J-connectivity of C10H20O E o. 2 Q. 3H 1111 UnHc-Hm UnHg-Hm' UnHk-Hmi UnHm-Hn Jan Novotný UnHdl-Hj \ UnHf-Hb i 1 -po^O Task 1: J-connectivity of C10H20O • CH lOm connected with CH2 8ck 5 4 3 2 1 5 4 3 2 1 G)2-1H (ppm) Jan Novotný 4/5 Task 1: -/-connectivity of C10H20O Jan Novotný Task 1: J-connectivity of CioH2oO CH2 8ck weakly coupled with CH2 7bj =>- closing ring protons b and k coupled to CH 6h which is connected to methyl 3e other expected crosspeaks in DQ-COSY crowded/overlapped, found topology confirmed in HMBC (3/4Jhc) e S 2 4 ■ UnHc- UnHg-Hm* UnHJt-Hm" UnfliD-Hn —i— 4 DnHŠ-Hj 3 2 G)2-1H (ppm) r f * * Jan Novotný 4/5 Task 1: J-connectivity of CiqH2oO 30- 35- E Cl O. O co 40- 45- 50- Jan Novotný j_i_ j_i_ j_i_ HSQC HMBC „ IT.UnC6-He UnC6-Hki----^' x - 30 UnC7-He^ UnC6-Hf UnC7-Hk - 35 UnC7-Hj UnC7-Hb UnC7-Hi - 40 UnC8-He UnC8-Hm UnC8-Hn UnC8-Hk„ „n TTl\ UnC8-Hb UnC8-Hh Hn \ xUnC8-Hh \ \ \\ - 45 UnC9-Hn UnC9-Hm UnC9;H1 UnC9-Hd °n""f \ UnC9-Hi \ Un""Ha UtiGQ-HgS1 - 50 = 1= c\(y Task 1: J-connectivity of CioH2oO • CH2 8ck weakly coupled with CH2 7bj =>- closing ring • protons b and k coupled to CH 6h which is connected to methyl 3e 9 other expected crosspeaks in DQ-COSY crowded/overlapped, found topology confirmed in HMBC (3/4Jhc) MENTHOL Jan Novotný 30 35 e q. a. O 40 45 50 4 _i_ HSQC HMBC UnC6-Hh UnC6-He UnC6-Hi UnC6-Hk UnC7-Hk' UnC7- UnC6-Hb 30 UnC7-He^ UnC6-Hf 35 UnC7-Hb UnC7-Hi 40 UnC8-Hm UnC8-Hn UnC8-He UnC8-Hk„ „„ \ UnC8-Hb kUnC8-Hh -Hn \ ---- \UnC8-Hh \ "7 \ \ \W ITnPQ-Hl UnC8-Hc UnC9-Hn UnC9-Hm unLy.H1 UnC9-Hd 45 UnC9-Hg UnC9-Ha 50 4 2 (02 - 1H (ppm) 4/5 Task 1: Stereochemistry of menthol C10H20O lo2: homonuclear/heteronuclear couplings • large couplings preserved in ID slices of HSQC: axial H - 2 visible interactions (geminal and vicinal) x equatorial H - only geminal o ID TOCSY: selective decoupling =^ simplification of complex multiplets • DQF-COSY: analysis of phase sensitive spectrum o lo3: NOE contacts (axial strong) Hy vH, HB 3JX.A = small 3JX.B = large C 3JHC= 6-8 Hz r 2Jur = 5-7 Hz HB 3JX.A = small 3JX.B = small C 3JHC = 1-3 HZ HB 2J = 0-2 Hz Jan Novotný 5/5 Task 1: Stereochemistry of menthol C10H20O lo2: homonuclear/heteronuclear couplings • large couplings preserved in ID slices of HSQC: axial H - 2 visible interactions (geminal and vicinal) x equatorial H - only geminal o ID TOCSY: selective decoupling =^ simplification of complex multiplets • DQF-COSY: analysis of phase sensitive spectrum o lo3: NOE contacts (axial strong) Jan Novotný 5/5