12.158 Lecture 7 - MIT OpenCourseWare12.158 Lecture 7 • Sterols part 2 – Sterol biosynthesis...

12.158 Lecture 7

• Sterols part 2 – Sterol biosynthesis review and revision – Steroids as age and environment

indicators – Enigmatic steroids 2- and 3-alkyl and

carboxysteroids

1

Sterols, Eukaryotes and O2squaleneepoxidesqualeneepoxide 29292929squalenesqualene1.14.99.7 + O21.14.99.7 + O2 28282828

OO

2222222221212121 2626262624242424

25252525

LANOSTEROL BRANCHLANOSTEROL BRANCH 5.4.99.7 5.4.99.85.4.99.7 5.4.99.8 CYCLOARTENOL BRANCH 1212

CYCLOARTENOL BRANCH1212

18 20 2318 201818 2320 2320 23

17171717 2727272711111111 1313131319191919 D 16DDD 161616CCCC1111 9999141414142222

8 15888 151515PROTOSTEROLSPROTOSTEROLS 10101010BBBBAAAA

77773 5333 555H OH O HOHO

lanosterollanosterol cycloartenol 4 64 6cycloartenol 4 64 6

2.1.1.41 1.14.13.70+ 3 O22.1.1.41 1.14.13.70+ 3 O2 smt1smt1

24-methylenecycloartenol24-methylenecycloartenol

smo1 + 3 O2 , 1.1.1.170, 1.1.1.270smo1 + 3 O2 , 1.1.1.170, 1.1.1.270eburicoleburicol4,4-dimethyl-5?-cholesta-8,14,24-trien-3?-ol4,4-dimethyl-5?-cholesta-8,14,24-trien-3?-ol H OH OHOHO HOHO

1.14.13.70+ 3 O21.14.13.70+ 3 O2 1.3.1.701.3.1.70 cycloeucalenolcycloeucalenol

4,4-dimethyl-5?-ergosta-8,14,24(28)-trien-3?-ol4,4-dimethyl-5?-ergosta-8,14,24(28)-trien-3?-ol 5.5.1.95.5.1.94,4-dimethyl-5?-cholesta-8,24-dien-3?-ol4,4-dimethyl-5?-cholesta-8,24-dien-3?-ol

HOHO HOHO HOHO

1.14.13.72, 1.1.1.170, 1.1.1.270 +3 O21.14.13.72, 1.1.1.170, 1.1.1.270 +3 O2 obtusifoliolobtusifoliol1.3.1.701.3.1.70

1.14.13.70+ 3 O21.14.13.70+ 3 O24,4-4,4 dimethylfecosterol-dimethylfecosterol 4?-methyl zymosterol4?-methyl zymosterolH OH O H OH O

HOHO

1.14.13.72, 1.1.1.170, 1.1.1.270 +3 O21.14.13.72, 1.1.1.170, 1.1.1.270 +3 O2 1.14.13.72, 1.1.1..170, 1.1.1.270 +3 O21.14.13.72, 1.1.1..170, 1.1.1.270 +3 O2 4?-methyl-5?-ergosta-8,14,24(28)-trien-3?-ol4?-methyl-5?-ergosta-8,14,24(28)-trien-3?-ol

4?-methylfecosterol4?-methylfecosterol 1.3.1.701.3.1.70zymosterolzymosterol

H OH OHOHO H OH O

1.14.13.72, 1.1.1..170, 1.1.1.270 +3 O21.14.13.72, 1.1.1..170, 1.1.1.270 +3 O2 5.3.3.55.3.3.5 4?-methylfecosterol4?-methylfecosterol

5.3.3.55.3.3.5fecosterolfecosterol 5?-cholesta-7,24-dien-3?-ol5?-cholesta-7,24-dien-3?-ol

HOHO H OH OHOHO

erg2erg2 1.3.1.721.3.1.72 24-24 methylenelophenol-methylenelophenol

episterolepisterol smt2smt2HOHO lathosterollathosterol HOHO H OH O

erg3+ Oerg3+ 2O2 1.3.3.2 + O21.3.3.2 + O2 24-ethylenelophenol24-ethylenelophenol

5,7,24(28)-5,7,24(28)ergostatrienol-ergostatrienol smo2 + 3 O2 , 1.1.1.170, 1.1.1.270smo2 + 3 O2 , 1.1.1.170, 1.1.1.270 H OH O 7-dehydro-cholesterol7-dehydro-cholesterol HOHO

HOHO

1.3.1.211.3.1.21erg5 + Oerg5 + 2O2 24-ethylenelathosterol24-ethylenelathosterol

5,7,22,24(28)-5,7,22,24(28)ergostatetraenol-ergostatetraenol 1.3.3.2 + O1.3.3.2 + 2O2HOHO cholesterolcholesterol HOHO

HOHO

1.3.1.711.3.1.71 24-methylene 5-dehydroepisterol24-methylene 5-dehydroepisterol

1.3.1.211.3.1.21ergosterolergosterolHOHO

HOHO

isofucosterolisofucosterol

dwf1dwf1H OH O

sitosterolsitosterol12 O2 11 O2 11 O2 H OH O

Burial & diagenesis ergostane cholestane sitosterane

2

The effect of oxygen on biochemical networks and the evolution of complex life.

Jason Raymond and Daniel Segre' Science 311, 1764-1767 (2006)

Cholesterol

This image has been removed due to copyright restrictions.

squalene hopene,tetrahymanol

3

http://prelude.bu.edu/O2/networks.html

4



Oxidosqualene Cyclase


5

Symbiogenesis:the phylogenetic

tapestry ofeukaryotes

http://www.life.umd.edu/ This image has been removed due labs/delwiche/ to copyright restrictions.

6

http://www.life.umd.edu/labs/delwiche/

http://www.life.umd.edu/labs/delwiche/

Eukaryote Diversity & Chloroplast Endosymbiosis

Anaerobes stem of tree?

Algae Forams

Radiolaria


©Jacob Waldbauer, 2007 Keeling et al. 2005

7

Phytosterols Whereas vertebrates and fungi synthesize sterols from epoxysqualene through the lanosterol, plants cyclize epoxysqualene to cycloartenol as the initial sterol. Q. Presumably lanosterol biosynthesis predates cycloartenol biosynthesis? What might have driven the lanosterol-cycloartenol bifurcation?

HO cycloartenol

HO β-sitosterol 24-ethyl-5-en-3β-ol HO

stigmasterol 24-ethyl-5,22-dien-3β-ol

HO ergosterol 24-methyl-5,7,22-trien-3β-ol

8

Phytosterols C30 sterols are generally minor components of organisms and immature sediments. However, when they occur, they have distinctive structures that are easily recognised in the ancient record.

red algal spongeHO sterol HO sterol

HOHO dinosterol 4a-methyl-24-ethyl

cholestan-3β-ol

9

Oxidosqualene Cyclase Alignment


10

Microorganism Microalgae Bacillariophyceae Bangiophyceae Chlorophyceae Chrysophyceae Cryptophyceae Dinophyceae Euglenophyceae Eustigmatophyceae Haptophyceae Pelagophyceae Prasinophyceae Raphidophyceae Rhodophyceae Xanthophyceae Cyanobacteria: Methylotrophic bacteria Other bacteria Yeasts and fungi Thraustochytrids

Major or common sterols

C28D5,22, C28D5,24(28), C27D5, C29D5, C27D5,22 C27D5, C27D5,22, C28D7,22 C28D5, C28D5,7,22, C28D7,22 C29D5,22, C29D5 C29D5,22, C29D5, C28D5,22 C28D5,22 4Me-D0, dinosterol, C27D5, C28D5,24(28) C28D5,7,22, C29D5, C28D7, C29D5,7, C28D7,22 C27D5 (marine) or C29D5 (freshwater) C28D5,22, C27D5, C29D5,22, C29D5 C30 D5,24(28), C29D5,22, C29D5, C28D5,24(28) C28D5, C28D5,24(28), C28D5 C29D5, C28D5,24(28) C27D5, C27D5,22 C29D5, C27D5 C27D5, C29D5, C27D0, C29D0 (evidence equivocal) 4Me-D8 C27D5 C28D5,7,22, C28D7, C28D7,24(28) C27D5, C29D5,22, C28D5,22, C29D5,7,22

The nomenclature is CxDy where x is the total number of carbon atoms and y indicates the positions of the double bonds. In general, C28 sterols have a methyl group at C-24, and C29 sterols have a 24-ethyl substituent. Table adapted from data in Volkman (1986); Jones et al. (1994) and Volkman et al.

11

Uncommon Marine Sterols

HO HO 24-methyl-27-nor sterols and stanol 24-nor sterols known in sponges: C27 compounds a range of algae & invertbrates: C26

H H19-nor sterols A-nor sterolsR=H, CH3, C2H5, methylene spongesΔ22 trans unsaturationprobably formed by de-alkylationof algal sterols

HO

H

H

R

HOH2C

12

Uncommon Marine Sterols (2)

HO HO

Aplysterol Gorgosterol

13

How to identify sterols by GC-MS of TMS and acetate derivatives

• Relative Retention Times of Nematode Sterols

• Mass Spectra of Nematode Sterols

• Mass Spectral Data for Nematode Sterols, Analyzed as Steryl AcetateDerivativesa Steryl acetate________Mass spectrum (m/z, relative intensity tobase peak)__Cholesta-5,7,9(11)-trienol 424 (5), 364 (100), 349 (33),251 (31), 209 (64), 197 (43), 195 (52)Cholest-8(14)-enol 428 (100),413 (18), 368 (6), 353 (13), 315 (16), 288 (7), 273 (6), 255 (21), 229(42), 213 (43), 81 (80), 55 (83)Cholesterol 368 (100), 353 (14), 260(15), 255 (13), 247 (18), 213 (14), 147 (48), 145 (37), 81 (74), 55(71) Cholestanol430 (12), 415 (2), 370 (33), 355 (16), 316 (3), 276(28), 275 (18), 257 (4), 230 (17), 215 (100), 201 (18), 147 (32), 81(53)

14

Sterol Reading #1

Text has been removed due to copyright restrictions.

Please see http://www.springerlink.com/content/q05172k241v60328

15

http://www.springerlink.com/content/q05172k241v60328

Reconstruction of past biota

16

Biomarkers and Fossils what can they can say:

Who are the major groups of marine primary producers today?

Which groups dominated at different periods in the geologic past?

How is plankton growth recorded in rocks – and oil?


What factors influence the completeness of the fossil record of marine plankton?

How has long-term ecological succession of marine plankton affected the evolution of other organisms and biogeochemical cycles?

17

Marine Primary Producers

Today, 2 major groups:

Acquired photosynthesis by secondary endosymbiosis

Were preceded by red/green algae & prokaryotic phototrophs

Left a rich body & molecular fossil recordRose to ecological prominence relatively

recently

Picocyanobacteria Prochlorococcus/

Marine Synechococcus

These images have been removed due to copyright restrictions.

Chl a+c Phytoplankton Diatoms

Dinoflagellates Coccolithophorids

18

Eukaryote Diversity & Chloroplast Endosymbiosis

Anaerobes stem of tree?

Algae Forams

Radiolaria


©Jacob Waldbauer, 2007 Keeling et al. 2005

19

The Fossil Record of Phytoplankton - one form of bias: where sediment is deposited

20

Another form of Bias:Age of the Ocean Floor

200 0 NOAA NGDC

21

White Cliffs of Dover

Rocks Made of Plankton Cretaceous coccoliths

Diatomaceous Earth Mine, Wallace Co., Kansas Grace Muilenburg, Kansas Geol. Surv.

K~T Boundary at Stevns Klint Denmark

Courtesy of Organic Geochemistry Group, Universitat Bremen. Used with permission.22

Succession in the Photosynthetic Plankton of the Ocean: A Perspective Drawn from

Chemical Fossils

Roger E. Summons & Jacob R. Waldbauer (MIT)Andrew H. Knoll (Harvard University)

John E. Zumberge (GeoMark Research)

Successions in Biological Primary Productivity in the Oceans. In Falkwoski P. and Knoll A.H. (eds) The Evolution of Photosynthetic Organisms in the Oceans, 2006.

23

Evolutionary Trends from Rocks & Oils: Present-day sample localities


24

Evolutionary Trends from Rocks & Oils• Sedimentary organic matter is the direct geologic legacy of

primary production • Massive accumulations of organic matter in petroleum systems

worldwide record ocean biogeochemistry • Oils are widely available, accessible, abundant & carry the same

kind of evolutionary information that is buried in sediments • Oils reflect the natural ‘average’ in the variation in source rock

organofacies • GeoMark Database: Biomarker parameters from over 1800

microbial-sourced oils (no terrigenous input) have been averaged to obtain 133 petroleum systems from the Neoproterozoic to Miocene

• The source rock type and age for many of the oils in GeoMark’s database are known based on extensive integration of geological and geochemical frameworks

25

0.6

PaleoLatitude vs Carbonate-Sourced OilsC

29/C

30H

1.4

1.2

1.0

0.8

carbonate marl shale

0.4

0.2 -80 -60 -40 -20 0

Paleo Latitude 20 40 60 80

26

Pentacyclic Terpane Ratios C

31R

/H

0.60

0.55

0.50

0.45

0.40

0.35

0.30

0.15

0.20

0.25 carbonate marl shale lacustrine

0.10 0.2 0.4 0.6 0.8

C29/H 1.0 1.2 1.4

27

• Clearly, these groups rise to paleontological prominence in Mesozoic and Cenozoic oceans, but…

• Is there an earlier, “cryptic” evolutionary history? – Unmineralized or poorly mineralized stem diatoms or

coccolithophorids? – Non-diagnostic fossils of dinos (sans archeopyle) among

older acritarchs?? – Help from molecular clocks?? – Sedimentary record:Biomarker molecules are most

informative

28

C28/C29 Sterane Ratios: The Rise of Modern Plankton

29

Dinoflagellate Biomarkers-Petroleum Record

C25

C16

C15

C17 C12

C21 C22

C2

C1

C4

C3

C5

C8

C9

C6

C7

C18

C19 C20

C10

C14

M8

M1

M10

M6

M4

M9

M3 M2

M7

M5

DS67

DS12 DS38

DS3

DS26DS8

DS11

DS52

DS49

DS27

DS10

DS16

DS57

DS69 DS62

DS24

DS60 DS55

DS14

DS68

DS50

DS13

DS61

DS1

DS20

DS19

DS65 DS17

DS46

DS42

DS2

DS6

DS43

DS71

DS41

DS59

DS56

DS34

DS70

DS73DS64 DS63

DS9

DS4

DS66

DS47

DS53

DS37

DS39

DS15 DS40

DS44

DS48

DS72

DS51

DS45

DS74

DS21

DS5

DS54

L23

L12

L17

L11

L7

L5

L21

L1

L20

L18

L6

L16

L8

L15

L4

L3

L13

L14

L19

L2

L9

L24

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

0 50 100 150 200 250 300 350 400 450 500 550 600

Source Rock Age (mybp)

Aro

mat

ic D

inos

tera

nes

(3/3

+6)

carbonate marl distal shale lacustrine End Cretaceous End Permian End Proterozoic

30

Aro

mat

ic d

inos

tero

ids/

2+3-

met

hyls

Distribution of dinosteroids in Phanerozoic

sediments This image has been removed due to copyright restrictions. Moldowan and Talyzina

Science 281,168-1170, 1998

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C26 steranes

21-nor Unknown source



32

Secular Change in C26 Sterane AbundanceApplication of 24-norcholestanes for constraining source age of petroleum

A. G. HOLBA et al. Org. Geochem. 29,1269 -1283, 1998 Rampen et al., AGU 2004:��..Another specific biomarker is 24-norsterol. Its value as an age diagnostic biomarker was already reported (3), but the source of this sterol was still unknown although a diatomaceous source was assumed. We have now found this sterol in the diatom species Thalassiosira aff. Antarctica. In combination with the knowledge that the 24-norsterol production increased substantially during the Cretaceous this may provide a tool to predict the mutation rate of the Thalassiosirales. Our data show that molecular paleontology can assist in obtaining more reliable estimates of the molecular clock rate and thus be an important tool

217

Diatom sterol Diatom sterane

in reconstructing the evolution of diatoms.

HO 24-nor

33

C26 steranes elution pattern


Moldowan et al GCA 55, 1065, 1991

34

Secular Change in C26 Sterane Abundance Application of 24-norcholestanes for constraining source age of petroleum

A. G. HOLBA et al. Org. Geochem. Vol. 29, pp. 1269 -1283, 1998

Courtesy Elsevier, Inc., http://www.sciencedirect.com. Used with permission. 35

http://www.sciencedirect.com

http://www.sciencedirect.com

Diatom-specific HBI & their Geologic Occurrence Highly branched acyclic isoprenoid alkenes and alkanes

The Rise of the Rhizosolenid Diatoms Jaap S. Sinninghe Damsté, Gerard Muyzer, Ben Abbas, Sebastiaan W. Rampen, Guillaume Massé, W. Guy Allard, Simon T. Belt, Jean-Michel Robert, Steven J. Rowland, J. Michael Moldowan, Silvana M. Barbanti, Frederick J. Fago, Peter Denisevich, Jeremy Dahl, Luiz A. F. Trindade and Stefan Schouten

23 APRIL 2004 VOL 304 SCIENCE

C20 HBI C25 HBI

•Identified in the diatom genera


Rhizosolenia, Haslea, Navicula, and Pleurosigma only •Biological products have 1-6 unsaturations; fossils fully saturated

36

C30 Desmethylsteranes

37

C30 Desmethyl Steranes Oil from Southern Oman

OM20 24-i-propylcholestanes 414 217

NZ Kora 24-n-propylcholestanes 414 217

59:00 1:00:00 1:01:00 1:02:00 1:03:00 1:04:00 Time

ααα 20R

ααα 20S

αββ 20R+S

38

C30 Desmethyl Steranes Eastern Siberia Oils

59:00 1:00:00 1:01:00 1:02:00 1:03:00 1:04:00 Time

ES36 414 217

ES89 414 217

i / n > 1

i / n < 1

39

Stratigraphic column of Huqf Supergroup withrepresentativ e lithology, biostratigraphy and geochronologica This image has been removed

l constraints. due to copyright restrictions.

GD Love et al. Nature 457, 718-721 (2009) doi:10.1038/nature07673

40

Catalytic hydropyrolysis (HyPy) biomarker geochemistry applications

� Pyrolysis assisted by high H2 pressure (15 MPa) and a molybdenum catalyst (active phase is MoS2)

� A powerful tool for releasing bound biomarkers

� high yields of biomarker hydrocarbons

� less structural/stereochemical alteration

� Love et al. (1995) Org. Geochem. 23, 981

� info on bonding (D2)

41

Temperature-programmed, open-system pyrolysis (fast residence time of volatile

Hydropyrolysis apparatus

High pressurehydrogen

Thermocouple

Reactor tube

Sample bed with dispersed Mo catalystElectrical

connectors

Gas collection And measurement

Mass-flow controller

Pressure transducer

Cold trap

Temp prog. amb. – 500oC @ 8oC/min H 2 pressure15 MPa sweep gas flow10 dm3/min

42

Comparison of free and kerogen-bound hydrocarbons TICs of saturates for a Minassa-1(A1C) sample

05 May 17 10 17 19 std.100 * Free saturated hydrocarbons 15 Ph 21 23 * = std.

% 25 hopane/sterane regioni18Pr 27

x x 29 31xx x x x x

2 05 May 31 08

17 19 HyPy of Kerogen

100

15 21 23

% Ph 25 hopane/sterane region

i18 27Pr x x x x x 29 31 x x x x 2 Time

15.00 20.00 25.00 30.00 35.00 40.00 45.00 50.00 55.00 60.00 65.00 70.00 75.00 80.00 85.00 90.00

Numbers refer to carbon chain lengths of n-alkanes X= series of mid-chain methylalkanes (unknown origin)

43

MRMGC-MS ion chromatograms of C26–C30 desmethylsteranes

released from catalytic hydropyrolysis of a Masirah Bay Formation (JF-1) and a Ghadir

Manquil Formation (GM-1) kerogen.


GD Love et al. Nature 457, 718-721 (2009) doi:10.1038/nature07673

44

21418 b/c +std 50ng/1000ul 2- & 3-methylsteranes 03100216 100 6

60.82 2.11 414.423 > 231.212

62.82 4-methylsteranes

% 58.6459.04 63.06 58.15

65.12 65.63

0 03100216 100 62.80 414.423 > 217.19661.32

57.57 58.62%

59.81 63.00s

52.24 53.10 55.95 56.86 64.23

o0

03100216 100 61.20 400.407 > 217.196

59.91 56.11

% 57.12 58.76

55.39

0 03100216 100 61.14 404.432 > 221.221

%

51.57 55.02 58.23 64.050 Time

MRM –GCMS howing C30 steranes f a marine sediment

52.50 55.00 57.50 60.00 62.50 65.00 67.50 70.00

46


2- & 3alkylsteranes

Note the exact co-elution of the

four synthetic isomers with the

equivalent peaks of the

Phosphoria oil

47

2- & 3alkylsteranes

Note the different isomer

This image has been removed due to copyright restrictions. preference for

marine vs lacustrine sediments

48

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12.158 Molecular Biogeochemistry Fall 2011

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12.158 Lecture 7 - MIT OpenCourseWare12.158 Lecture 7 • Sterols part 2 – Sterol biosynthesis...

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