Data Inventory Sheet

...
1
Commodity D~

-~ 2. ..AutD..9 r Wt..Kc\:' L.i~- ~'"

3. Title (or description)
\" :....! ,... T,' _,
J ~.
:-\, ..... -" .

ROBINSON, H. E. I solation pr0c edu re s for ke r ogens and associate d soluble

CJrga n ic ma terlals .• Y,tOrganic CE'och em istry: Heth ods a nd Res ults. Sprin ge r -
Verl ag , New York, Chapter 6 , 1969, pp. 181- 195.
4. Date Iq I., q
5. Reference CJ...-. . ~
b~_~c....~ ," ~:'0C.I·" ('I·:: ' .~l\l . \ ( ~~u.. ,,-Q:,'.t. )nl. .t.<Af f-..,- ()
s,~~ \)0...
I
·".cr
6. Source
L"J l~~ ~ l- I\. W..l c:A.. "C (.J

7. Location of Data
~~~~~' . Lcl:'~ I:_ ~" ~ ~\:.~
8. Form of Oat a
9. Type of Work

10. Description uf Work

11. Types of Data

12. Qua "tity of LJata

13. Quality of Data
14. Pri ority
 Reprint from
Organic Geocbemistry (Eglinton/Murpby)
Springer-Verlag Berlin' Heidelberg· New York 1969
Printed ir. Germany. Not for sale

W. E. Robinson

Isolation Procedures for

Kerogens and Associated Soluble Organic Materials
 HAPTER 6

Isolation Procedures for Kerogens

and Associated Soluble Organic Materials
W. E. R OB! 0

Laramie Pel roleum Reseurcl! C ellter,

U.S . Bureau of Milles .
Laramie, Wyomillg

Cont en t ~
181
I. Introduction. . . .
18?
II. I olation of Kerogen
I 6
Ill. IsolatIOn of Bitumen
188
IV. Isolation of Acids
V. Isolation of Porphyrins
190
191
VI. I olation of Amino Acid
191
VII. Isolation of Carbohydrates
192
VIII. o ncluding Remarks
193
References . . . . . . . . . .

I. Introduction
Kerogens are of geocnemical intere t because they represent one of the mos'
abundant forms of carbonaceou3 material s. The term •. kerogen ", or" oil-former ",
was originally given to the organic matter in oil haies. torbanite, kukersitt', and
others. Recently, b~oader usage has been made of the term in describing the
insoluble organic material pre ent in nonreservoir sedimentary rock s and other
rocks. The soluble organic materials associated with oil-shale kerogens are of
particular intere t becau e they are usually im':igenous to the formation and have
had little opportul1lty for migration becau e of the low poro itl' and low perme-
ability of the formations.
DOWN and H!M US ~l] published a cia ificativn of hales and coal ba ed upon
a number of properties including the yield of oil on distillation. By thi classifica-
tion, the term " kerogen rocks" riescribe sedimentary de po its containing insol-
uble organic kerogen which on dis, :1!'ltion yields an oil equivalent to more tha 1

L
1 2 W. E. ROBINSON :

50 pert;e nt of the organic content of the rock . im ilarly, the term .• kerogen coals ,-
de cribe edimentary materials containing organic matter which upon distillation
yie ds an oil equivalent to more than 10 percellt a nd less than 50 perc;:nt of the
organic content.
In the present discussion, the term" ke"ogen -, will be used to describe the
insoluble organic material pre ent in kerogen rock . The term "bitumen ,- will l)e
used to de cri!Je the soluble organic material present in the kerogen rock be sed
upon olubility in a hydrocarbcn solvent. Other nomenclature will t -. elf-
explanatory. or example, the material olubilized by methanol will be referred
to as " methanol- oluble material ".
The di cussion cover the i olation dkerogen, bitumen , fatty acids, porphyrin s,
ammo acids, and carbohydrate from kerogell rock~ . The procedures as described
are intended to be informative without any implication of recommended usage.
Other methods are applicable and in ome cin;umstances ma y be preferred . Like-
\ lise, the articles reviewed wI: re chosen for the subject matter discus~ed and no
attempt has been made to completely review the ubject.

II. Isolation of Kerogen

One important problem in kerogen onstitutional &t udies is the isolation of
unaltered kerogen from it as ociated mineral. The problem of the i olation of
kercgen is complicated by extreme d;rferences in mineral content and mineral
composition of the kerogen roc! . Some ke,'oge n rock s contain a little as 5 percent
mineral while other may contain as much as 95 percent mineral. ome ediments
are highly argillaceou while other highl y calcareous. Some kerogen rock
contain large amounts of pyrite but others contain little pyrite. Pyrite is extremely
difficult to separate from the organic material becau se o f an apparent attraction
for ,he kerogen . Thus, !t is understand:Jble that the best avai lable separation
technique do not find universal application.
Separation techniques sho uld ideally remo ve a ll mineral matter from th
organic kerogen without fractionating the kerogen into dissi milar component
or altering the kerogen chemically. t the present time thi has not been acco m-
plished . The methoos to be described con i t of the modified Qua s _ sink-n at,
acid dige tion, and pyrite r\:moval. In studies of oi l shale at the Bureau of Mine
laboratory, techniques other than tho e to be described such as electro t::ttic.
magnetic, ultrasonic, and notation eparations have been tried with only limited
succe .
Hand-picked samples of oil shale from the Bureau of Mine demon tration
mine near Rine, Colorado, are crushed to 1- to 2-in('h pieces by using a la rge-
size jaw crusher. The I - to 2-inch piece. of sa mple are crushed to mesh or smaller
in a small- izejaw crusher. The -mesh oil- hale sa mple is then cru heel to 100 m 's h
or smaller in a hammer mill or more recently in a disc mill, consisting o f crushing
di cs con tamed in a barrel th a t vibrate in an eccentric manner. Tnis method
quic~ : ly cru he the oil hale to very small particle sizes. The possibility exist that
minor degradati n or oxidation of the o rganic material in the kerogen rock may
occur during the crushing procedure.
 Isolation Procedures ror Kerogens and Associated Soluble Organic Materials 1113

A. Modified Quass Method

The method a used by Q UAS [2] on outh African kerogen rock is an adapta-
tion of the amalgamation pr ocess used for coal. Th e method efficientiy reduced
the ash c~' ntent of the outh African kerogen rock from 40 percent to about
~-
5 percent. H.\1 uS and BAsAK [3] u ed the method on kerogen rocks and obtained
significant mine. ,,; IClnoval from only one of four different samples. The modified
Quass method as 'Ised successfully 011 Green Ri ve r Formation kerogen rock is
described by MITH and HIGBY [4].
This method is based upon the principles of differential wetting of the organic
kerogen and the inorganic mineral by two immi sc ibl e liquids such as oil and
wa.<!r. The organic kerogen is wet by the organic liquid pha se and the mineral i'
wet by the water pha se re ulting in the kerogen being retained in the oil phase
and the mineral beir.g relea sed into the water ph os:::. Peri odic changing of the
water phase results in mineral reduction . ew surfaces on the oil-kerogen phase
are created b y so me form of kneading. mixing, or grinding ac ti o n .
Pri or to thi s concentration procedure. the ~ erogcn ro~ k IS crushed to pass a IOO-mesh creen,
then ex trGcted ",ith benzene to remove most or th~ soluble organic material in th e kerogen rock .
Arter air drying. the extracled and crushed rock is leached with dilute acid to remove mineral ca rbonates.
The next tep in the procedure is the preparation or a pastelike ball or the kerogen rock and an
oily phase. Co nsiderable practice IS required in preparing a paste ball that \\ ill retain its adhesive
nature in the ",ater phase. This is done by mixIng tht' extracted and acid-leached kerogc:1 rock with
an oIly phase in the container or an att . ition g'inder. Sufficient oily phase is stirred with the crushed
, r
rock to rorm a paste with the consistency or a light putty. Mineral oil or n-hexadecane have pruven
satisractory ror th e oily phase; however. num erous othcr organic materials may be use:ul. Thc n-
hexadecane has the aJvantage over mineral oil or co ntributing only one impurity orknown composition
in case the oily phase ca nn ot be 'o mpletely remov~d from tile kerogen .
Water is added to the container along \\ ith ceramic balls and a grineing and mixing aClion is
sta rted . The mineral is preferenlially wet by the water and tends to accumula te in the water phase.
The organic kerogen is prererentially wet by the oily phase and tends to be retained in the oily phase.
Periodic removal or al'd replenishing the water IJhase reduces the minera! content or the kerogen
rock. The mineral "rallout" is quite rapid during the .:arl) part or the grinding; ow.equently. the
water phase should be changed rrequently during the first rew hours or grinding. This process is con-
tinued until a minimum a h va lu e is obtained lor the sample. With some kerogen rock s. the disappear-
ance or the X-ray difTraction peak ror soda re ldspar can be rollowed to show the reduction in mineral
Lontent.
At the end or the grinding, the water phase is removed b) suction and sufficient ac~tone is added
to destroy the adhesive nature or the oily phase and the kerogen . The kerogen concen,rate, oily phase.
and the ::cetone arc removed rrom the grinding balls by suction arter \\ hich the acetone and some
or the oil is removed rrom thr keroren concentra te by filtration . The sample is air dried and e trarled
with benzene to remove the remaiOl,.g oily phase rrom the kerogen concentrate. The sample is then
dried at 60° C under reduced pressure. Arter dryinb, the sa mple IS washed w;;h hot wate.· first by
decanting and finally bye. traclion in a soxhlet extractor ror 24 hours. Th~ sample is then dried at
60° under reduced pressure. nushing occasionally wi th nitrogen.
A comparison of th e decrease 111 mineral con tp;ll a nd othe r dat a ob tain ed for
II kerogen rock s is shown in Table I. Red~! ::tion in a h CCl~tent amounted to
95 percent or more for two of the k e r oge n rocks. In only o n ,' kerogen rock was
the reduction in ash content less than 50 perce nt. There was cor siderable variation
in the amount of the tot a l organic n~? terial ro::presentcd in the (oncentrate. ranging
from 24 to 99 percent for the Bra z il ia n and outh Afric:1I1 ke :ogen rock, respec-
tively. The assay oil / carbon ratios va ried from 0.43 h) 0.05 for the II kerogens
which rahged in age from 12 to 300 million yea r s. The oil carbon ratio were not
I 4 " . E. ROB I so .. :

Table I. Approx imar e age. assa) oil ro orgwlic c llrh ? 1I rari()~, mId rhe decrease ill ash co lll elll by a
cOIICl'lIIrariulI procedllr~ jor 11 uii shales

arne and location Approximate Wt percent assay Ash before Decrease in ash
age oil to "t percenl concentration content by a
(million years) organic carbon (percent) concentration
ratio 0 procedure
(percent)OO
---- ------
Alas a (Howard Pass) 120--215 0.95 34.1 96
Argentina (San Juan) 19 215 0.61 82.6 5
Brazil ( ao Paulo) 12 1\52 75.0 83
Canada (New Glasgow) 300 0.44 84.0 60
Colorado (Piceance k) 60 . 5 05.7 ~O
Fran~e (SI. Hilaire) 215 0.43 66.3 73
ew Zealand (Orepuki) 60 0.54 12.7 70
Oregon (Shale City) 40 0.70 48.3 25
Scotland (Dunnet) 250 0.67 77.8 76
South Africa (Ermelo) 215 300 0.72 33.6 96
Spain (Puerto llan o) 215- 300 0.68 62.8 59

o Reported by ROBINSON and DINNEEN [5].

00
once lllrated by the Modified Quass Method [4].

related to :lge but undoubtedly 'ue related to the molecular tructure of the
kerogen~ . The kerogens with low oil carbon ratios have high conten ts of condensed
aromatic ~tructure that produce low yields of oi l and high yield of carbon
residue upon pyrolysis. On the other hand, kerogens with high oil/carbon ratics
have higt. contents of oil-producing materials such as aliphatic and alicyclic
structure.
The main advantage of this concentration procedure is the absence of drastic
chemical treatment, producing essentially,] chem ica lly unaltered kerogen concen-
trate. However, the possibility of fractionating the kerogen of some kerogen rock s
does exi t. A disadvantage of the concentration procedure i the time required to
prepare the concentrate, which is usually severa l days. Also, the mineral reduction
for some kerogen rocks is very low and with mo t kerogen rocks the reduction in
mineral content i not complete. However, the method is ideal for some kerogen
rocks, for example Alaskan and outh African kerogen rocks (Table I).

B. Sink-F loat Method

The ink-float method of concentrating kerogen takes advantage of difTerences
in the pecific gravity o f the mineral components and the organic kerogen. For
example, Green River Formation kerogen has a specific gravity of 1.07 [6] , and the
mineral components have a range of specific gravity from about 2 to 5. Separation
is accomplished hy centrifugation in a dense liquid medium where the lightweight
kerogen tends to float and the mineral tend to ink.
The sink-noat technique was used by lUTS [7] in lowering the ash content of an "oil shale "
kerogen rock sample to about 5 perceni. In this procedure. finely powdered rock is suspended in a
olution of CaCl, having a specifi,; gravity of 1.06 to 1.\5. The mixture is centrifuged at 3.000 rpm
until 010 t of the organic material rises to the surface of the liquid and the min<!ral sinks to the b{Jllom
of the contai ner. The organic material is filtered and washed free of CaCI 2 ·
 Isolation Procedures for Kerogens and Associated oluble Organic Materials 185

H HHARD el al. r] centrifuged kerogen rock In mixture " of carbon tetrachloride and benzene
having specific gravllies f~om 1.15 to 140. In thi. method t!"le kerogen rock. ground to pass a 100-
mesh screen, is extracted with benzene to remo~e benzLne-so luble material and leached with dilute
acid to remove mineral carbonates. The kerogen rock, extra ted. leached. and dried, is suspended
by stirri ng in a 1.40-density carbon tetrachloride-benzene medium. then cen trifuged . Thc noating
material is removed from the cen trifuge tube by aid of suction, then filtered , washed with acetone.
and dried. The material that sinks is treated similarly. Portions of the .. noat" sample a nd tl.e "siny "
sample arc retained for analyses. The remainder of the "noat " sample from the first ce ntrifuga tion
is uspended in a 1.20-density carbon tetrachloride-benzene medium and centrifu ged in a simi lar
manner. Likewise, a pordon of the material tha t noated on the 1.20-density medium is suspended
in a l.I 5-density ,arbon tetrachloride-benzene medium and cen trifuged. By this procedure, a total
of three "noat " sam ples a nd three "sink" sa mples of different kerogen concentration are obtaIned .
The first flOat concent,':!!e, representing 39 per-::ent of the kerogen, contained
27 percent ash. The econd float concentrate, representing 6 percent of the kerogen,
contained 14 percent a h. The final float concentrate with 9 percent a h repre ented
only 1 percent of the total kerogen . A total of 12 percent of the kerogen was ob-
tained in concen trate having 15 per",ent or less a h.
The H/C ratio and the assay oil/organic carbon ratios of the ink and float
concentrates suggest that little or no fractionation of the kerogen occurred. For
examr le, the H/C ratios of the concentrates had maximum devia tion of only
1.2 percent and the assay oil/C ra tios of the concentrate had maximum deviation
of only 3.4 percent. If the kerogen had been r.o mposed of materials of widely
different compo itions and the e materi als had been selectively fractionated by
the concentration procedure, greater diffe rences would ha ve been obtained for the
two properties.
The advantages of this r.o ncentration me thod are (1) the kerogen is not altered
chemically, and (2) a concentrate of less than 10 percent mineral ca n be obtained.
Elemental analy es can be determined easily at vario u stage of concentration;
therefore, it is easy to extrapolate to zero ash co ntent a nd obtain elemental
compositions of the mineral-free k~rogen. The di sadvantages of the concentration
method are (I) low yield s of good c:mcentrate are obtained, and (2) some kerogen
may be fractionated.

C. Acid Treatment
One commonly u ed method of preparing kerogen concentrates is acid digestion using hydro-
chloric acid, hydronuoric acid o r combinations of these acids. The acid digestion method dS described
by SMITH [6) consists of treating the finely ground and benlene-extracted rock In a polyethylene or
other resistan t container wi lh sufficient methyl alcohol to form a thick pa~te . The purpose o f the
a lcohollrea tment is to wet the sample so that good contact bet"een acid and rock is achieved. Mineral
carbona tes are removed by treatment of the m0 :st rock with a dilute solution of hydrochloric acid
( I part concentrated HCI to 9 parts water) at room temperature ulllil evolution f CO, ceases. The
~uspension is filtered and the residue is washed with water. The residual rock is treated then with I : I
concentrated HC I (37 perce.,t) and water at room temperatl're for I hour followed by filtering and
washing with water to remove mineral carbonates. The washed residue is treated with a I : I mixture
o f concen trated HCI (37 percent) and II F (48 percent) and e\aporated 10 dampness on low heat of
abou t 100° C to volatilize the reaction products of the si licates. With mo t rocks it is necessary to
repeat the acid trea tment and evapora tion . The damp residue is suspended in sa tu rated boric acid
soluti on to prevent the fo rmati o n o f insoluble nuorides and then filtered . T hi s prucedure is repeated
a nd the resiclue is washed finally with water. The resIdue is ex 'racted then "ith I : I HCI a nd H 2 0,
j
filtered , and washed with water. The residue is suspended in hot water and" ashed succ ;;sively by
decanting and filteri ng until the filtrate is free of chloride ions. The kerogen concen trate dried at

L
 1 6 \) . E. R OB I'" ON :

O' C under red uced pressure. \ ilh Green River oil shale. Ihe co ncen lra le fro m Ihis procedure co nlains
3 10 5 percent min eral o f wh ich abo ul 80 percenl o f Ihe mi nera l is pyri le.

Thf! advantage of this meth od are (I) mo t of the minerals (exce pl pyrite) a re
rem oved from the k e rog~ n rock. (2) there i little opportunit y for frat:ti ona ti on
of the orga nic ma terial. and (3) the meth od i ra pid. Th e main di sadvantage of the
methvd is the possible altera ti on of the orga nic kerogen by the a ti on of strong
mineral acids.
D. Pyrite Removal
Pyrite i one con tituelll of the rock mineral th at is di ffi cult to remove from
the kerogen by phys ical meth oci 3 beca use of a n a ppa rent a tt racti on of the pyrite
for the kerogen . Pyrite ca n be removed by nitric aci d di gest:on; howeve r. the
nitric acid ox idize a nd nitrates the kerogen. Reducti on with zin c and hydro-
chloric acid remove the pyri te l,lwl y but the ex tent of reduction of fun cti onal
groups pre en t in the kerogen is not ea ily pred icted. LAW LOR el 01. [9] removed
pyrite qu a nt ita tively by redu ction usi ng li thium aluminium hydride, re ulting in
specific alterati on of kerogel. fun cti onal groups.
T he la ller melhod consisl s of Irculing finel y cru shed . bcn zen e-cxlracled . and carbo nale-free
kerogen r'lck wilh a le lra hydro furan solull on o f LiAI H, . Fi l e pa rts of sa mple 10 o ne pa ri o f LiAI H,
is placed in a nask equipped \I ilh a renu x conden ser. A l o lume o f IClrahydro fur a n cqui,alcnI 10
7.5 ml per gram o f sample is slo l\ Iy inlrod uced. The reaclio n mix lUre is renu \ ed r,) r 30 minul es. coo led
10 room lemperalUre. and I acuum flllercd. slo pping Ihe flll ra lio n while Ihe resi due is still mo isl
The mo isl residue is Iransferrcd as quickl) as poss ible in small po rti o ns 10 a beaker ,'o nla ining " a lcr.
Th(' Ilaler reaCIS I\lIh and de ~ iro)' s Ihe exces; LiAIH, . Th e rcsulling mixlUre is aCldiflc,1 wi lh I HC\.
healcd 10 boili ng. and flllc lcd . s a re~ ull o f Ihe acid Irea lmCn! Ihe pyrili c sulfur is e' I,ea as II .
and Ihe pyrilic iro n remai ns in so luti on as ferro us io ns. The lilhium and aluminium o f Ihe co mple \
arc di sso lvcd by Ir~aling the re s id~ c sufficicllll) lI ilh acid So lUlio n 10 assure complelc I·: mo val o f
alumi nium io ns. as dCle rmined by leslin g Ih e wa shings \\lIh a mm o nium h)droxidc. Th e produci
is washcd free o f chlo ride io ns by dcc~ '1lin g and filler \lashing \\ ilh hOi \l a lcr a nd is fin ally dried
under vac uum al 60' C.
Lithium aluminium hydride treatment succe fu ll y ~e m ove d py rite from a
ca rbona te- free keroge n rock and a keroge n concentrate con tainin g origi nally
3. 1 and 5.3 percent pyrite. res pective ly. Quantita ti ve remova l ,,[ pyrite fro m the
sample was hO\v n by chemica l a naly i for pyrite by a modifica tio n of the Mott
meth od [1 0]. The di a ppea ra nce of th e major X-ray diffraction pea k for pyri te in
each of the sa mples ubstantia ted the co ncl usion tha t the remova l of PY rite was
complete.

III. Isolation of Bitumen

An important a nd int res ting ph ase of a stud y of a ke rogen roc k is the in-
vestiga ti on of the compo ilion of the so luble ma terial associa ted with th e kerogen.
urrentl y, th e geochemica l as pec ts of thi type stu dy are of con iderable interest
beca u e of the emphasis placed on findin g new petroleum ou rt:e beds by the
aid of geochemica l info rm a tio n and techn iq ues. Mo t ,- oii shale'- keroge n rock
a re uniqu e a nd a re of part icul ar interest beca use the orga nic ma teria l is ind igenou s
to the forma ti on with litt le opportun ity to migra te to or from other loca tions.
Many dim' rent technique for the ex tracti on of so luble orga nic materia l from
kerogen rocks have been devised a nd are being used [ 11- 14} Fo r exa mple. ome

L
 I olation Procedure for Kerogens and A sociated oluble Organic Materials 18i

inve tigators u e a range of solvent and extract the rock ucces ively with olvel1ls
of increasing polarity. Thi tends to eparate the sc..luble material into fractions
of different degrees of polarity. Other in estigators u e mixed solvents, usually
a hydrocarbon olvent plus a polar solvent. These extractions provde quantities
of polar materials not usually removed with hydrocarbon olvents.
FERGuso, [15J tudied variables in the extraction of bitumen from sediment .
Yield of bitumen roughly doubled when particle ize of the ample was reduced
from 30 to 10 microns by hammer-mill cru hing. n extraction time of 24 hours
was con idered sufficient to remove more than 99 percent of the bitumen. Benzene
was found to be more ati factory for removing hydroca rbons than mi ed olvents.
A pecially designed extrl\ction apparatus where Ihe ample was continuou Iy
tirred in boili'lg ~olvent wa more efficient that a sox hlet extractor. BAKER [16J
used the techr.ique of FERGU ON in tudies of extra-:ts from sediments of the
Cherokee Group. CUMMINS and ROI3JNSON [17J extractri bitumen from Mahogat~ .Y
zone kerogen rock of the Green River Formation. ROBJNSO I!I al. [18J studied
the bitumen obtained from sections of a 900-foot core of the Green Ri ver For-
mation. B RLINGAME el al. [19J isolated steranes a nd triterpanes fwm .reen
River Formation kerogen rock and a Precambrian shale . To RTELOT and FROST
[20J studied the extractable organic material in nonmarine and marine shale
of Cretaceous a2e. HILLS el al. [21J identified gammacerane. a pentacycl ic tri-
terpane, in bitumen of the Green River Formation kerogen rock . EGLINTON el al.
[22J extracted hydrocarbons of biological origin from <: I-billion-year-old edi-
ment. ROlliNSON and DIN EEN [5J e tracted bitumen from I ~ different kerogen
rocks of different age. different envirollment and of different orga nic content.
OR6 el al. [23J isolated and identified n-alkanes ranging from CI~ to C n COl11-
pounds in fungal spores. Odd-carbon-numbered ompounds predominated in
the C 2S to C 29 range. ALVIN and Me ARTilY [24J isolated the C l 7 isoprenoId
hydrocarbon (2.6,10-trimethyltetradecane) from an a ncient sedimen t.

A. Extraction Technique
The method to be described i a simple benLene ex tracti on of the kerl)gen
rock . Variations of the extraction techniq ue. u ed by other investigators, are
satisfactory and for certain applications may be more deSirable. everal investi-
gators hzve made u e of ultrasonic vibr:J tion in the extraction process.
The kerogen rock. crushed to pass a IOO-mesh screen. is placed in an extracto r of appropriate
stze. For samples up to 500 grams. a soxhlet e.~tractor of si7c suitab le for the sample is used . Consider-
able channeling OCCI' rs in the large extraction thimbles "nd it IS deSIrable to periodically stop the
extraction and stir the sample. Because of this channelin~ , it is best to use ;mall sample,. Equipment
simi lar to that described by FERcusoN [15] "ould be desirable. Filter aids such as sand or other
sui table materials can be u~ed where anal}sis of the residual kerogen rock is not to be determined.
For samples of several kilograms. it has been found comcnicnt to extract in 20 liter carbo}s at room
temperature with con'~alll stirring. Three or four charges of fres:, sohent remo\es most of the soluble
material.
After placing the sample in a suitable extractor frc e.• purified benzene IS added to the extractor.
The extraction proce is started and continued unti l \cry lillie colored matcrial is remo\ed in thc
extract. Thi. u ~ ually is ncarl} complete in hours. but. in most cases It is cOll\enientto let the cxtraction
continue for 24 hours. The sohent is distilled from the cxtract at reduced pressures.
 W. E. ROBINSO :

B. Fractionation Technique
A ,"edification of the fraclionation technique used by CUMMI and ROBI SON
[17] follows. The crude extract is dissoived in a 40 to I volume ratio ofn-pentane
to ample and a ll owed to stand overnight at 0 0 C and then filtered . The pentane-
insoluble material remo ved by filtration is washed with a small quantity of cold
pentane (0 0 C) and is then dried.
One part of pentane-soluble material is placed on a prewetted column of 25 parts of alumina
(Alcoa, F 20, 0-200 mesh, extracted" ith pentane, activaied 400· C for 2 hours) and eluted with
purified solvents. The column IS exhau tively eluted first with pentane. followed by benzene, then a
mix1llre of 10 percent methanol 1:nd 90 percent benzene. Each of the eluted fractions is stripped free
of s01vents and dried . The pentane-eluted material is referred to as hydrocarbon conce ntrate. the
benzene-elu!ed materials as re.ir.~ I and the benzene-methanol-eluted material as resins II.
One part of hydrocarbun concentrate IS placed on a J: ' ewetted column of 2 parts of silica gel
(Davidson, 200 mesh, extracted with pentane, dried at 100· C for 2 nours) and eluted with purified
olvents. The column is exhaustively eluted first with isooctane, followed by benzene ar.d finally
2-propanol. The e fractions contain mainly alkane, aromatic, and polar compounds.
The aromatic oils were further fractionated on a column of alumina eluti~g successively with
isooctel1c, benzene. and isopropanol. The alkanes are fractioned into normal alkanes and isoalkanes
plus cydoalkanes on molecular sieves (Linde, 5 A. extracted with pentane, activaten at 240. C under
vacuum overnight [25]. The method consists of placing the alku'les on the sieves with hot isooctane
and sub~equcntly removing the trapped n-alkanes with n-pentane at room temperature.
When large quantities of branched-cyclic alkanes are prepared. the material is fractionated into
10· C cut in a molecular still at 10 J Torr pres ure starting at room temperature. The first few cuts.
being the lowest molecular weight material, are redistilled in a pinning-band distillation column at
I Torr pressure. Fractions from the me-Iecular distillation, having boiling points too high for the
spinning-band distillation, are placed on a column of alumina (200 parts alumina to I part 3mrle)
and eluted with n-pentane, collecting 25 ml fractions o f the eluting material. The solvent is removed
from each fraction by evaporation on a hot-water bath . using dry nitrogen to speed evaporation .
The resulting fractions from the vacuum distillation and the fractions from the a:"mi~a co lr :mns ap!
suitable for tra p ping from GLC columns. The trapped fractions. from repeated trappings, a'e identified
or classified by utilizing mass. infrared. and NM R spectral analysis.

Thermal iITusion [17] has been used in lieu of molecular di tillation for
fractionation of the isoalkane and cycloalkane . Although thermal t:;ITusion
oITers some ad van tage in type eparation. it i much more time-con uming than
molecular disti lation .

IV. Isolation of Acids

The presence of fatty acids in recent a nd ancient sediments is of geochemical
interest, contributing eVIdence for the organic origi n of petroleum a nd other
fossil fuels. It is of special intere I to tud y the relationship of fatty acid and the
corresponding n-alkanes in the sediment. The presence of other a lkane acids,
' uch a the iso- and anteiso-acid , i oprenoid acids, a:1d resin acids, are a lso of
interest from geochemical considerations.
Fatty acids up to 3.\ were identifIed by MEl SCHEI and KE ! EY [13] in an
aerobic soil sample. COOPER [26] identified fatty acids ranging from C to C
t4 30
frol11 recent and ancient 'ediment . A mechanism explaining the conver ion of
even acids to odd acids and odd alkanes, was proposed by OOPER and BRAY
[27]. ASLESON el al. [28] i.'xtracted fatty acid fro:n several source including

L
 Isolation Procedures for Kerogens and Associated oluble Organic Materials 189

the reen River Formation where fa tty acid up to t were identified. LAWLOR
and RObiN () [29J idl!ntified C tO to C 3S fatt y acids from Mahogany zo ne oil
shale of the Green River Formation. The di ·tribution of Fatty acids in ome
Lower retaceous sections of the Powder River Basin was studied by K VEN-
VOLDEN [30]. CA ~ON a nd GRAHAM [31J isolated C 14 , CIS , C 19 , and 20 i oprenoid
acids from a petroleum crude. EGU TON e l al. [32J i olated C t9 and C 2 0 chain
i oprenoid acid from Green River Formation kerogen rock . Branched- hain
fatty acids were isolated from marine ediment and from Green River Fo rmaticn
oil shale by LEO and PARKER [33].

A. Ex traction Techniques
In a tecnnique described by LAWLOR and ROBI NSON [29] a sa mple of ((JO-mesh kerogen rock
is treated with dilute acid to remove mineral carbonates and to convert salts of acids to free acids
The resi dual rock is treated then with 8 percent mcthanolic KOH at renux temperature for 18 hours.
The aqueous solution " f the potas ium salts of th e free acids, acids fo rmerly presen t as sa lt s, and ac id s
~ nerly pre ent as esters is sepa rated f~o m the insolubl : residu2 by filtration . Water and ca rbon
tetrachloride are added to the filtrat e and the mi xture is shaker. vigorously. By this treatment, the
potassium alts of the acids arc concentra ted in the wa ter p~d se a nd the nonaci dic material is con-
centrated in the carbon tetrachloride pha e. After se paration of t~ e water phase from the ca rbo n
tetrachlo ride ph ase, the water phase is acidified with H I. and the free acids are lemOI ed by extraction
with carbon tetrachloride. The solvent is removed leavi ng th e acid produ ct as a residue. The c rude
acids were cO'lverted to methyl esters using methanol and boron trinuori Ie.
In anot her technique, PARKER [34] extracted th ~ rock sam ple with methanol while the sa mple
was stirred and s u !>j~ctcd to ultrasonic vibrations fo r 15 minutes. The methano l extract is recovered
by filtration . The resicue is stirred with chloroform a nd subjecled to ultraso nic vibra tion ,. The extracts
rem oved by methanol and by ch lo roform are combined and sa ponified . The recovered acids arc
converted to methyl esters using m ~! hanol a nd boron trinu oride. This meth od docs not sa ponify
esters associated with any in solublt· kerogen that may be present.

B. Fractionation Techniques
The methyl esters of fa tty acids m'ly be concentrated by adduction with urea.
The traight-chain etas, the branched-chain esters or the total methyl e ter
fractiv.l may be analyzed by gas-liquid chromatography (U,-C). Comparison of
retention times of unknown con tituent with known sl llldarJs are utilized for
component identification. Trapped peaks from the GLC are ana lyzed by ma ,
infrared, and M R spectra to provide posi tive identificat ion of the constituents.
Me ARTHY and D UTHI E [35] descri bed a meth od for the separation of free fatt y acids (F FA )
from .; ther lipids using a column of silicic acid. Coar e particles of silicic acid a re selected by su pending
100 g o f the silicic acid in 400 ml of methanol and decanting an-i discarding the silick acid no t settling
in 5 minutes. This is repeated o nce with methanol a nd onc(: with 400 ml of aceton~ . fhe si licic acid
is rinsed with diethyl et her and perm itted to air dry.
Five grams of prepared silicic acid are mixed witlJ 10 ml of isopro panol-KOH (50 mg KOH per ml)
and 30 ml of dieth yl ether. After standing 5 minutes, the silicic acid is slurried into a glass column and
washed with tOO ml of diethyl ether.
The sample, dis olved in a small quantity of diethyl ether, IS placed o n the ilicic acid and thor.
oughly washed into the packing by s~ve ral small portion of diethyl ether. Sterols, ster')l esters. and
glycerides are eluted as o ne fract ion with 100 to 150 ml o f dielhyl ether a t a now rate of 5 ml per minute
The FFA are removed from the column with 50ml of 2 percent formic acid in dietlt yl ether (1: I;
followed by 75 to 100 ml of diethyl ether. If phospholipid arc present. they are removed fro m the
column by methanol.

L
 \ . E. ROB I SON :
190

Attempt to i olate alcohols in a h)drolysate of kerogen by utiiizing thin-layer chrorolutography

with silica gel were made by LAWLOR and ROBINSO [36). They also used thin-layer chromatography
to purify methyl esters of kerogen acids prior to mass spectral analyses.

V. Isolation of Porphyrins
The tudy of fo s il pigments a well as the study of other biological fo sils
provine information on the modilica tion of organic debri under geological con-
ditions. TREIBS [37] undertook a sy tematic earch for well-deli ned organic
pig nents in rock and found that extract of" oil hale" kerogen rocks, as old
a Tria ic, exhibited spectral characteri tic similar to those of complex metal
salt of porphyrins. This wa a particularly important discovery becau se por-
phyrins are regarded as remnants of biological precursors, uch as chlorophyll
and hemin .
More recently porphyrins have been tudied in various other sediments and
in petroleums. GROE NI GS [38] developed a technique for the quantitative
determination of porphyrin aggregate in petroleum . The extraction reagent was
a saturated solution of hydrogen bromide in glacia: acetic acid. The ~orphyrin
content of extracts from Green Ri ve r •. oil shale" kerogen rock was determined
by MOORE and DUN ' I G [39]. GIH RA and MCGEE [40] isolated a crystalline
porphyrin from gil onite of the Green Ri ve r Formation. BL MER and OME [41]
isolated porphyrins and chlorins from a Triassic" oil shale " kerogen rock of
Serpiano. Switzerland. A compari on of porphyrins in "oil shale " kerogen rock,
in shale oil, and in petroleum was made by MORA NDI and JE 'S N [42]. BAKER
[43J extracted porphyrins from gilsonite and petroleum a phaitene~ using
methanesulfonic acid . HODGSO el at. [44] reviewed the role of porphy rins in
petroleum , oil sa nds, oil hales, coal. and sedimentary rocks.
The method of extracting porphyrins from kerogen ro.;ks consist of either
(I) extracting the soluble material from the rock prior to treatment of the extract
with a porphyrin extraction reagent, or (2) extraction of the rock directly with
the porphyrin extraction reag'!nt.
MOORE a nd DUNNING [39] used the first technique. The kerogen rock is crushed and is ex t racted
successively wi th diffelent boiling solvents. The firs t solvent is benzene followed by ben7ene-methanol
azeot rope (60 percent benzene 40 percent methanol). benzene-ci.!oroform azeotrope ( ., percent ch lo-
roform 13 percent methanol), methanol a nd finally pyridine. The extraction is continued until the
ex tract is colo rless a nd sh ows no fluorescence under ultraviolet light. Each extraction requires n to
96 h o urs. The extracts are isolated by distilling off most of the solvent a t atm ospheric pressure followed
by heati ng at 50" C under reduced pressure. The porphyrin aggregate from each ex tract is isola ted by
method of UROENNI GS [38).
MORANDI and JENSEN [42] utilized the second technique and extracted the porphyrins directly
from the kerogen rock with the Groennings extraction reagent. Before the procedure could be used. it
wa, necessary to tr~at the rock wi th 10 percent H ~ Ito remove mineral carbonat "so
BLUMER and ( 'MENN [ 41] extracted thc kerogen rock with I : I benLene-methanol mixture and
iso ated the porphyrins and chlorins by solvent partitioning using different concentratio ns of HCl. By
this method. about 1.6 kg of the kerogen rock is exhaustively ex tracted with a benzene-mcthanol
mixture followed by evaporati on of the solvent. The dried extract is redi solved in a minimum amount
of chlo roform a nd the asphaltenes are precipitated by adding ho t isoo tane. The precipitate is filtered
and washed with a mixture of chloroforon and isooctane . The filtrate IS e" a po rated to dryness and
red issolved in warm diethyl ether. The ether solut io n is ex tracted suc('essi';~\Y with 2.4 H I, 4.9 N

,
L
 Isolation Procedures for Kerogens and ssocl3ted oluble Organic Materials 191

HC\. and finally with 6.1 H \. The 2.4 . HCI extract is neutraliLeJ wtlh sodium acetate and the
porphyrin pigment is removed by ether extra:tion. The 4.9 and 6.1 H I extracts are diluted to 1.0
acidity and the chlorins are recovered by ether ex traction.

VI. Isolation of Amino Acids

Proteins and nuc\eoproteins are universal components of all living organi ms.
The i oldtion and identificat ion of proteinaceous materials in sedimentary rock
is therefore of geochemica! interest.
The amino acid content of soils has been studied by various investigator .
STE ':E SON [45] examined soil> and obtained quantitative analy es of hydrolysate
from the ample . ERDMA el a/. [46] made a comparative study of amino acid
present in a recent marine deposit and those present in an Oligocene marine mud .
VALLENTYNE [47] reviewed the occurrence of amino acids, purines, and pyrt-
midine in lakes, ocean, ewage, and terrestrial oils. WAIN [48] analyzed Paleo-
zoic rocks for amino acids.

A. Extraction Technique
STEVENSON [45] described a method for extracting amino acid from soil samples. A 50 g sample
of finely ground, air-dry soil. from which the undecomposed plant resi dues had previously been removed
by flo tation o n water, is reOuxed with 150 ml of 6 HCI for a period of 24 hours. The mixture is filtered
thro ugh a Buchner funnel with suction and the residue wash~d thoro ughly with distilled water. The
filtrate and washings, containing the amino acid , a,e combined and the excess HCI is removed by
repeated evaporation at 50' C under reduced presS ure . ny humin present is se parated by cen tr ifuga-
tion .
8. Fractionation Technique
An aliquot of amino acid solution containing from ::0 to 25 mg of <x-amino acid nitrogen is passed
through a column of a cation exchange resin (Amberlite I R- 120) in the h)drogen form . The material
retained on :he column is \\ashed with di stilled water \'ntilthe eluate is neutral to litmu s paper then
eluted with 2 liters of 5 percent aqueous ammonia foll o\\ed by a washing of 500 mi of di tilled water.
The eluate from the second water washing is discarded until a po itive ninh)drin test is obtained th(n
the remaining eluate is r- tained . The I!luate from the ammonia and second water washings are combined
and the ammonia is removed by gent le evaporation. The solutio n i made slightly alkaline with Ba(OH h
and evaporated to a paste to remove all the combined amm o nia. as indicated by a negative Nessie 's
test. The residue is taken up in dis.illed water and the bari'IOl i rem oved as the sulfate. The final solution
is combined with the original water eluate.
STEtN and tOORE [49] used a column of Dowex-50 resin 10 separate amino acid s. An aliquot of
amino acids soluti on, containing approximately 6 mg of >!-amino acid nitrogen. is addt J to the top of
the column and is washed into the resin with thrle 2 ml portions of distilled water. The column is
eluted with 1.5 N H I until gluta,nic acid appeared in the eluate, with 2 HCI until valine appears.
a1ld finall y with <I N H C\,

VII. Isolation of Carbohydrates

Because of th high content of carbohydrates in plants relati ve to lipid and
proteins, the study of carbohydrates in ediments is particularly important to
geochemical considerati'Jlls. The geochemistry of carbohydrates ha been studied
exten ively ; however, the exact role played by carbohydrates in the formatil)n
of fo si l fuels remains to be learned .

L
192 W. E. ROBINSON :

Free sugars ranging from 10 to 3,000 mg/kg of edimentary organic material

were found in fresh water sediment by VALLE TY:-IE and BID\. ELL [50J and
WHITTAKER and VALLE TYN [51]. D FF [52J obtained wa ter-soluble paly-
accharides from peat and soils. Trace amounts (Ies than I mg/kg) of glucose
were found in h)drolysate of Green River kerogen rock by PALACAS [53].
HABAROVA [54J studied the ca bohydrates in ediments of marine and inland
eas. Sugar released by acid hydroly is of marine sediment from the anta
Barbara Ba in were reported by PR HNOWSKY el al. [55]. P LA AS el 01. [56J
reported the pre ence of carbohydrates in ancient sedimentary rock. WAI. and
ROGERS [57J studied the distribution of carbohydrate residues in Middle Devonian
Onvndaga beds of Pennsylvania and we tern ew York .

A. Extraction Techniques
WAIN and ROGERS [57] described a method of ext ractive degradation of carbohyd rates in sedi-
ment s. leaned and crushed samples are treated with 50 percent sulfuric acid to degrade carbohydra te
materials to furfurals . The resulting solution i tested in triplicate for total ca rbo hydrates by adding a
phenol solution [58). The density of the colored solutions is measured in a colorimeter a nd th p :-,tal
ca rboh ydrates, expressed as glucose equivalent. are calculated by comparison with known samplr. of
D-glucose. Mont)saccharides are extracted and sep' rated irom the rock by acid hydrol:'~:>, The
quantity of ulfuric acid n.~essary to neutraliLe the alkaline constituen ts of the rock is deter,nined by
preliminary treatment with a known amount of acid "110 subsequent back-titration with aOH in the
presence of phenolphthalein. An amount of 0.5 H2 0 4 acid is added to neutw!ize the alkaline
constituents plus an amount reqUired to hydrolyze the rock sample. The sample-acid mixture is boiled
under renux for 8- 10 hours. Then the mixture is centrifuged and neutralized with BaCO , . Desalting
is done first by ethanolic precipitatio"l then by passage through ion-exchange resins. The desalted
hydrolysates are concentrated in a na h evaporator to standard volume and examined by paper
chroma tography.
WHIn'AKER and VALLENTYNE [51] desc ribe a semiquan titati ve method for the determination of
free sugars in lake sediments. The sample, suspended in 70 percent eth anol, is boiled for 5 minutes, then
filtered while the ex tract is still hot. After the filtrate cools it is filtered again to remove any material
precipitated on cooling. The ex tracts are deionized on ion-exchange resins to remove inorganic mate-
rials. The dei o nized extract is concentra ted by evaporation to dryne s under reduced pressure at 45° C.

B. Fractionation Technique
WHIn'AKER and VALU TYNE [51] used unidimensional paper c .....omatography to separate sugars.
A wad of folded filter paper is stapled to the bottom of a sheet of What man o. I filter paper in order
to Illcrease the now and mallltain an even now during the development of the chromatograms. The
equilibrated mixture of butanol, ~thanol, and water (45: 5: 50) is used as the solvent. Ten spots (5 to
50 microliters) of star.dard sugar solution with three spots of unknowns interposed between the
standard spots are placed on each of duplicate chromatograms C hromatograms are developed for
72 hours. Prol'J nged development o f 7 days improves separation of nonpentose sugars; however, the
long developmen t removes pentoses. After -Jevelopment, one chromatogram is prayed with? benzidine
spray which reveals the position of all the subars in the standard mixture and their counterparts in the
unknown. Other chromatograms are spra yed "ith indicators for specific sugars.

VIII. Concluding Remarks

Much progress has been made in isolating kerogen from their associated
mineral ; however, it is apparent that additional research in this area is needed .
This would properly include fundamental re earch of the prop _'rties of the mineral
 IsolatIon Proced ures for Kerogens a nd Associated oluble O rga nic Material 193

and it relation hip to the o rga nic kerogen. It would be of interest t ascerta i.l
the extent and na ture of an y a ttracti ve fo rces between minera l a nd kerogens.
Thi would require study of individua l f0 .Tul1iio ns ince it i unlikely tha t the
properties of each kerogen rock a r~ the same.
Much development work ha been ex pended in modifymg a nd improving
technique for i olating indi vidual compo und a nd pigment s associated with the
insoluble ke rogen in kerogep rocks. Thi rese'lrch has re ulted in the identifica ti o n
of a significant number of compo unds which provide info rma ti o n concern ing
the o rigin a nd tran forma,ion o f ca rbo nClceou materia l . Much mo re data need
to be co llected :.l nrt organi z:!d before logi<;a l cor.c1u ions can be drawn . In relating
a given organic deposit with ano ther, orne impro vement in o rga ni zatio n of
data would re ult if isola tion procedures were sta nd a rdi zed .

References
I. DOWN, A. L., a nd G . W. H IM US: Classilica ti'ln of oil shales a nd cannel coa ls. J. Inst. Petrol. 26,
329-34R (1940).
2. Q A<;S. F. W .: T he a na lysis of the ke rogen of oi l sha les. J. lns t. Petrol. 25, 8 13-8 19 (1939).
3. HI MUS, G. W., a nd G. . B SA K: A na lysis of coal a nc ca r bo nacecu materia ls cont a ining h igh
percentages of inheren t m inera l mailer. Fue l 28,57 - 64 ( 1949).
4. S~IIT lj . J. W., and L. W. H IGBV : Preparation of organic concent ate from Green River oil shale.
A nal. Ch~m . 32, 17-1 (1960).
5. ROBINSON. W. E.. and G . U. 0 1 NEEN : ConHitutional aspects of oil-shale ke rogen. Proc. 7th World
Petroleum ongress, ~exico City, Mexico 0.3,669-680 (April 1967).
6. SM ITH. J. W .: Iti mate co mposition of organic matenal in Green River oil shako .S. Bur. Mines
Rept. Invest. 5725 (1961).
7. L' TS. K.: T i;·~ usc of the Ooat a nd sinl- method for isola tion of organic matter in oi l shales. Brenn-
stoff- hem. 9, 2 17-2 18 ( 1928).
8. H UBBARD, A. B., H. N. MITH, H. H . HEA,' , and W . . ROll iNSON : Method of concentrating kerogen
in ' o lo ra do o il sha le by trea tmen t with acetic acid and gravity sepa ration. U .S. Bur. Mi nes
Rept. In vest. 4872 ( 1952).
9. LAWLOR , D. L.. J. I. FESTER, a nd W. E. ROB INSOr- : Pyrite remo\ al from oil-shale concentrates using
lith ium a luminium hydride. Fuel 42. 239 - 244 (1963).
10. H ESS, W . VAN, a nd E. EARLV : A modilicatio n of MOII's method for the determination of pyri tic
su lfu r in coa l. Fuel 38. 425 - 42 ( 1959).
II. S~'ITII , P. V.. tudies on origin of petroleum : Occurrence of hydrocarbons in recent sedimen t .
Bull. Am . Assoc. Pet rol. Geologists 38. 377 - 404 (1954).
12. Hu T, J . M ., F . STEWART, and P. A. DICKEV: Origin of hydrocarbons of inta Basin, tah . Bull.
Am . Assoc. Pet rol. G eologists 38, 1671 - 169 ( I , : ~ )
13. M EINSCl IEI . W . G ., a ncl G. S. KE NEV : Analysis o f a chromatographIc fraction of organic extracts
of oils. An a l. C hern . 29, 11 53- 11 61 (1957).
14. BR AV. E. E., a nd E. D. EVANS: D ist ribu tion of n-paraffins as a clue to recog nition of source beds.
G cochim. osmochim . ACla 22, 2- 15 (196 1).
15. FERG SO , W . S.: Determ ining hydrocarbo ns in sediments. Bull. Am . Assoc. Petrol. Gcologi ts 46,
1613 - 1620 (1962).
16. BAKER, D. R .: O rganic geochemistry of Cherokee Group in c.~theas t ern Ka n as and orth-
eastern Okl a ho ma. Bull. A m. As OC. Petrol. Geologist s 46, 162 1- 1642 (1962).
17. CUMMI NS, J. J ., a nd W. E. ROBI NSON : ormal 'tnd isoprenoid hyd rocarbons isola ted from oil-shale
bitumen. J. C hern. Eng. Da ta 9, 304 - 307 (1964).
18. ROBINSO , W . E., J. J. CUMMI S, a nd G . . DINr-;EEN: Changes in Green R iver oi l-sha le paraffins
"it h de pth. G eochirn . Cosmochim. Acta 29, 249 - 258 (19651
19. B RLINGAME, A. L.. P. H AUG, T. BELSKV, a nd M. AL\IN : O ccurrence o f biogenrtic sterane and
penta -cyclic triterpa nes in a n Eoce ne sha le a nd in an early P reca mb ri a n s ha le. P roc. at. Acad.
Sci. U.S. 54, 5 (1965).
1:\ Org.lnl eochemlstry
 194
W. E. ROBINSO, :

20. To RTELOT, H. A., and I. C. FROST: Extractable organic material in non -marine and marine s hales
of Cretaceous age. U.S. Geol. urI'. Profess. Papers 525-0, 73 _ 8 I (1965).
21. H IL , I. R., E. V. WHITEHEAD, DE . -.ND£RS, J. j . C MMINS, :lOd W. E. ROBINSON : '\n optica lly
active Irilerpane, gammace,ane, in Green River olorado oil-shale bilumen. hem . ommun.
20, 752 - 754 (1966).
22. EGU TON, G ., P. M . ScoTT, T. BELSKY. A. L. B RUNGAME, and M . CI\LVIN: H ydrocarbo ns ofbiolog-
ica l origin from a one-billion-year-old edimenl. Science 145, 263 - 264 (1964).
23. OR6, J ., J . L. LASETER, and L\ WEBER: Alkanes in fungal spores. cience 154, 399 _ 400 (I 966).
24 CALVIN, M ., and E. D. M CCARTHY : The isolalion and idenlificalion oflhe 17 Isoprenoid saturaled
hydrocarbon. 2. 6, 10-lrimelhyltelradecane : squalene as a possible pr~cu r~or. Div. o f Pel. Chern.,
Am. C hern. Soc., ew York, N. Y oo 87 - 91 ( epl. I I - 16, 1966).
25. O' 0 OR. J . G ., F. H . BUROW, and M . . ORRIS : Delerminalion of normal paraffins in C,o 10
paraffin waxes by molecular sieve: ad orpt ion . Anal. Chem . 34, 2 _ 5 ( 1962).
32

26. COOPER. J E.: Fally acids in recem and a ncienl sedimenls and pelroleum reservoir walers. alure
193,744- 746 (1962).
27. - , and E. E. BRAY : • postulaled lOle offal ty acios In pc" ': ""m formal ion. Geocilllll. Cosmochim .
ACla 27, I I 13 - I 127 (1963,.
28. AUEt:'<lN, P. H ., T . C. HOE RING, and P. L. PARKER : Fally acids in sedimenla ry rocks. Inrern. Ser.
Monographs Earlh Sci. 15. 169 - 174 (1<;64).
29. LAWLOR, D. L., and W . E. ROBINSON: Fally acids in Green River formation oil shale. Div. of Pel.
Chern., A,n. Chern. Soc., Delroit, ~ l ich oo General Papers No. 1, 5 - 9 (April 1965).
30. K V5NVOLDEN, K . A .: Molecular di tribution of normal fall) acids and paraffins in omc Lower
Cretaceous sedimenls. Nalure 209. 573 - 577 (I 96/i).
3 I. ASON, J ., and D. W. GRAHAM : Iso lalion of isoprenoid acids from a California pelroleum . Telra-
;tedron 21. 471 - 48 3 (1965).
32. EGUNTON, G., A. G . Do GLAS, J. R. MAXWELL. J. . RAMSAY, a nd S. STALL8ERG-STENHAGE : Oc-
currence of isoprenoid fally acids in Ihe Green River shale. Scie'lce 153, 1133 - I 135 (1966).
3•. Lro, R . F., and P. L. PARK [R: Branched-ch ain fally acids in sediments. cience 152, 649 _ 650(1966).
34. PARKER . P. L., and R . F. LEO : Fally acids in blue-green algal mal communilies. Science 148,
373 - 374 (1965).
35. M e ARTHY, R . Doo and A. H. DUTH IE: A rapid (,uantilali ve method for Ihe sepa ralion of free fally
acids from olher !JI,;ds. J . Lipid Res. 3, 117 - I 19 (1962).
36. LAWLOR. D. L., and W. E. ROBINSON : Alkanes and fally acids in Green River fo rmal ion oil sha le.
ymposium " In tera ,o' :plinary AspeclS of Lipid Research ". Am . Oil hemislS' I}ciely, Los
A ngele ,Ca lif. (April 1966).
37. TREIBS, A.: hlorophyJl and hemin dcri\alives in organic mineral subslances. Angew. Chern. 49,
b 2 (1936).
38. v~OENN I GS, S.: Quantilat/Ve delerminalion of Ih po rphyrin aggregate in pelroleum. Anal.
C hern . 25, 938 -94 1 ( 1953).
39. MOORE. J. W., and H . . DUNNI 'G: Inlerfacial aClivilies and porphyrin conlenlS of oil-shale
eX lraCIS. Ind. Eng. Chern. 47, 1440 - 1444 (1955).
40. UG IHARA, J . M oo and L. R. M CGEE: Porphyrins in gilsonile. J . Org. Chem. 22, 795 - 79 (1957).
41. BL MER. M ., and G . S. OME N: Fossil porphyrins : ncomplLxed chlorins in a Triassic edimenl.
Geochi m . Cosmochim . Acta 25, 81 - 90 (1961).
42. MORAND I, J. R., and H. B. JENSE : omparison of porphyrins from shale oil, oJ! shale, and petro-
leum by absorplion and mass spectroscopy. Chern . Eng. Dala 11 , 8 1- 88 (1966).
43. BAKER, E. W. : l.1ass spectrometric characterization of pctroporphyrins. J. Am. Chern. Soc. 88,
2311 - 23 15 (1966).
44. HODGSON, G. W., B. L. BAKER, and E. PE~KE : The role of porphyrins in the geochemistry of petro-
leum . 7th World Petrolcum ongr. Mexico City, Mexico (April 1967).
45. STEVENSO , F. J .: Ion exchange chromatography of a mino acids in oil hydro lysa les. Soil ci. Soc.
Am . P roc. 18,373-377 (1954)
46. ERDMAN, J. G ., E. I. MARLETT, and WE. HANSON : Survival of amino acids in marine sediments.
cience 124, 1026 (1956).
47. VALLENTYNE, J . R .: T he molecul a r nature of organic mailer in lakes and oceans with lesser reference
to ewage and terrestrial soils. J. Fisheries R e . Board Can . 14, 33 - 82 (J 957).
 I olallon Procedures for Kcrogens and A ocia lcd oluble Organic Malerial 195

4 ' ,\ ' ' ' . '' 1.. [)"lnbUlion ofsorne organic ;ub;lance In i ' ICOIOI C rocks OfCcOI ral Pennsylvania.
in : Coal clcncc. Wa hlnglon. D C , Amcncan Chemi 11 OClely 1966.
49. 1/ 1'1. W. H .. and S MonKI. Chro malographlc dClerminaoon of Ihe amino acid composilion of
protein, Cold Srnng Harbor }mp Quanl B, oI. 14. 179 (1949).
50. \'AII~'1n'I . J R. and R. G . S. BID\\~LJ ' The rclalion belween free sugars and sedimcn tary
chlororh} II Inla ~ e mud;. Ecology 37. 495 - 500 (1956).
5!. WIIiTlAKI K. J . R . and J. R. VALLI '11\'-1 ' On the OCCurre nce of free suga rs in lake edimenl
CXlract •. Llmnol Oceanog. 2.9 - I 10 (1957)
52 Dll~. R B : Occurrcnce of mClhylaled carbohyd r.rte, a nd rh amno;e as components of soi l poly.
\1H:chandc, J . c. r' ood Agr. 3.140 - 144 (1952)
53. PALA(, s. J G Geoch rn"try of carbohydrate •. Ph D. TheSIS. n"erSII} of Minnesota (1959).
54 II AOARo\ •. , T. The hlochemical compositi on of deep·water marine mud deposil . Biokhimlya
20. 146 - I 1(1955)
55. PRASH .o"'~ ,,\. A . LT. DEGI 'os. K O. E'I~R\. and J. PIME'I1A : Organic materials In Reee", and
anCient ,ed,m"nts Part J. Sugars In manne sed, menl of Santa Barbara Basin. Calrfornia.
'euc, Jahrh. Geol. Palaeont ol. Mondtsh. 8. 400 - 413 ( 1961).
51>. PAl At A~. J. (I .. .. >"1 '''AI/'. . and I . S\fl'! fI : Pre e.lee of r.arbohyd rates and olher o rgal1lC com.
pounds In un(,lent .cdlmenlary rock,. ature 185.234 (1961).
57. S\\AI ... . f M .. and M A R(}(.r R,: Siraligra rhic dlsl.ibulion of carbohydrale resIdues In Middle
Devonl,m Onondaga beds of Pennsylvania and Wes lern 'ew Yo rk . Geochim . Cosmochim .
ACla 30.497 - 509 ( 1966).
5~ ;), HOI •. M .. K . A Gil LLS. J . K. HAMIU O". P. A. RI H' R'. and .. S~IITfI :
L' o lo rimetric method for
delerrnlnallOn of ;ugar. and relaled subslances Anal Chen, 28.350 _ 356 (1956).

L
L