Zbl. Bakt. II. Abt. 135 (1980), 454-458

[Chemical Laboratories, University of Allahabad, Allahabad, India]

Organic Acids Produced by Self-Sustaining Coacervatesin Presence of p-Nitroaniline and p-Phenylene-Diamine

PARVEEN TRIPATHI and VIJAY SHANKAR TRIPATHI

Summary

Oleic and malonic acids were found in control samples while p-nitroaniline addition inducedproduction of tricarballylic acid in eight and twelve days exposed samples. Fumaric and citricacids were found in sporadic instances. Addition of p-phenylene diamine to the mixture showeda degeneration process in organic acid production. In some cases production of fumaric acid waspositive.

Zusammenfassung

Die Untersuchung befaEt sich mit del' Bildung von organischen Sauren in sieh selbst erhalten­den, lebensahnlichen Systemen. Das Grundsubstrat, bestehend aus Diammoniumhydrogenphos­phat, Ammoniummolybdat, Mineralsalzen und Formaldehyd, erhielt Zusat.ze von p-Nitranilinoder p-Phenylendiamin und wurde unterschiedlich lange dem Sonnenlicht ausgesetzt. In denKontrollen wurden Olein- und Malonsaure nachgewiesen. p-Nitranilin induzierte bei acht- undzwoltt.agigcr Belichtung die Bildung von Trikarballylsaure, Fumar- und Zitrononsaure tratensporadisch auf. Zusatz von p-Phenylendiamin hihrte zu einem DegenerationsprozeE bei del' Bil­dung organischer Sauren, In einigen Fallen konnte Fumarsaure nachgewiesen worden.

Matter which was lifeless was somehow transformed into the living form. Thissubtile change of far-reaching consequences has all along interested intelligent man.Even in this modern age man has not been able to completely fathom these unknownmysteries of nature.

• HALDANE (1929) enunciated the Molecular of Chemical Evolution Theory whichbegan the scientific investigations on Origin of Life. BAHADUR and RANGANAYAKI(1966) studied the natural conditions which formed replicating self-sustaining life-likesystems. PAVLOVSKAYA and PASYNKII (1957) obtained amino acids by exposingBAHADUR'S (1954) mixture to ultraviolet rays. BAHADUR (19G4) obtained low mol.-wt.peptides on exposing aqueous mixtures of amino acid, sugar, and iron or molybdenumas catalysts to sunlight. BAHADUR and SINGH (1971) obtained phospholipids, whileBAHADUR and RANGANAYAKI (1970) got positive tests of deoxyribose nucleic acidin mixture.

For the present investigation the authors studied the formation of organic acidsin mixtures of diammoniurn hydrogen phosphate, ammonium molybdate minerals,and formaldehyde, containing p-nitroaniline or p-phenylene diamine at variousperiods of exposure.

Microscopic observations (TRIPATHI and TRIPATHI 1976) and amino acid changes(TRIPATHI and TRIPATHI 1978) involved have also been studied by the authors.

atganic Acids, Produced by Self-Sustaining Coacervates 455

Materials and Methods

All chemicals used were of ANALAR grade.

1. Preparation of amine solution

0.5 g of amine was t.aken and dissolved in 100 ml of 4 N hydrochloric acid.

2. Preparation of mixture

a) Diammonium hydrogen phosphate 4 % (w/v) aqueous solution,b) ammonium molybdate 3 % (w/v) aqueous solution,e) mineral, containing 0.2 g each of sodium chloride, potassium sulphate, calcium acetate, and

potassium dihydrogen phosphate in 100 ml distilled water,d) formaldehyde 36 %.

The above four solutions were mixed in the following proportion: a: b: c : d = 1: 2: 1: l.5 ml each of this mixture wer.. pipetted out and transferred to twenty test tubes. The tubes

were cotton-plugged and autocIaved for thirty minutes at 15 psi; cooled.

3. Addition of amine solution

Four tubes were taken (marked zero hour) and into two of them 0.5 ml of p-nitroaniline solu­tion was added and into the other two 0.5 ml of p-phenylene diamine solution. All the twentytubes were exposed to sunlight. After foul' hours of exposure, four tubes (marked four hours)were taken and 0.5 ml of the amine solution was added. Again these tubes were kept in sunlightSimilarly, amines were added at eight, twelve, and sixteen hours of exposure.

For the blank and control 0.5 ml of water and 4 N hydrochloric acid, respectively, was added.After exposing t.ho samples for four, eight, and twelve days, the study of organic acids was per­formed.

4. Organie acid separation

It was done by ascending paper chromatography, using 'Vhatrnan No.1 chromatographicpaper.

5. Mounting of mixture on chromatographic paper

It was done by using a micropipette and mounting 0.05 ml of the mixture on the corner ofthe paper and at a distance of :3 em from the edges.

6. Chromatographic tank

Two tanks of the ascending type were used. They contained two solvent systems, butanol: for­mic acid: water = 10: 2: Hi (KALYANKAR 1952) and phenol: water: formic acid = 75: 25: 1(STARK et al. 1951).

7. Staining was performed by spraying with 0.5 % (w/v) bromcresol green in absolute alcohol.Bright yellow spots were visible where the organic acids were located.

8. Identification of organic acids

Known acids were chromatographed and R r calculated. The H r of unknown must coincidewith that of known, then presence of that acid was confirmed.

Results and Discussion

Addition of p-nitroaniline to the mixture gave malonic acid in all stages andoleic and malic acids in some samples. Fumaric acid appeared in samples which wereexposed for sixteen hours before addition of the amine and subsequently for fourdays. But as the time of exposure after addition of amine was increased to eight days,

Tab

le1.

Org

anic

acid

sof

typ

ical

mix

ture

ofco

acerv

ate

s~ O

t0"

>

Sam

ple

Ex

po

sure

Ex

po

sure

ho

urs

ofad

dit

ion

ofw

ate

r/h

yd

roch

lori

cacid

tim

e12

16:-0

ind

ay

s0

48

f-3 ~ i;jB

lan

k4

Mal

on

icac

idM

alo

nic

acid

Mal

on

icacid

Mal

on

icacid

Mal

on

icac

id> H

(ty

pic

al

mix

ture

Ole

icacid

Ole

icac

idO

leic

acid

Ole

icacid

Ole

icac

id::1

wit

h0

.5m

lw

ater

)M

alo

nic

acid

Mal

on

icacid

Mal

on

icac

id~

8M

alo

nic

acid

\M

alo

nic

acid

~ p..

Ole

icacid

Ole

icac

idO

leic

acid

Ole

icacid

Ole

icac

id~

12M

alo

nic

acid

Mal

on

icacid

Mal

on

icac

idM

alo

nic

acid

Mal

on

icac

idrp

Ole

icacid

Ole

icac

idO

leic

acid

Ole

icacid

Ole

icac

idf-3 ~

Co

ntr

ol

4M

alo

nic

acid

Mal

on

icacid

Mal

on

icac

idM

alo

nic

acid

Mal

on

icac

idi;j >

(mix

ture

co

nta

inin

gO

leic

acid

Ole

icac

idO

leic

acid

Ole

icacid

Ole

icac

idH ~

0.5

ml

HO

I)-

Mal

icac

idH

8M

alo

nic

acid

Mal

on

icacid

Mal

on

icacid

Mal

on

icacid

Mal

on

icac

idO

leic

acid

Ole

icac

idO

leic

acid

Ole

icacid

Ole

icac

idM

alic

acid

12M

alo

nic

acid

Mal

on

icacid

Mal

on

icacid

Mal

on

icacid

Mal

on

icac

idO

leic

acid

Ole

icac

idO

leic

acid

Ole

icacid

Ole

icac

id

Ta

ble

2.A

min

oa

cid

ina

min

e-c

on

tain

ing

mix

ture

s

Ad

dit

ion

of

am

ine

aft

er'o

xp

osu

rco

fm

ixt-

ure

inh

ou

rsA

min

ea

dd

ed

p-n

it.ro

uu

ilin

e

p·p

hcn

yle

ne

dia

min

e

Day

so

fex

po

sure

aft

er

am

ine

ad

dit

ion

4 8

12 4 11 ]2

o Mal

on

ica

cid

Ole

icac

id

Tri

carb

ally

lic

acid

Mal

icac

id

Tri

carb

ally

lic

aci

dM

alic

acid

Mal

on

ica

cid

Mal

on

ica

cid

4 Mal

oni

ca

cid

Ole

icac

id

'I'ri

carb

aH

ylic

aci

dM

alic

nei

d

Tr-i

carh

nll

yli

cu

cid

Ma

lic

'lci

d

Ma

l on

ica

cid

Mal

ica

cid

8 Mnl

on

icac

idO

leic

acid

Tri

cn.

rbuf

lyli

ca

cid

Mal

icac

id

Tri

carb

all

yli

ear

-id

Mal

ica

cid

Cit

ric

ac

id

Mal

on

icac

idM

alie

aci

d

Fu

mari

cac

idF

um

ari

ca

cid

12 Mal

oni

ca

cid

Tri

curb

ull

yli

cac

idM

alic

[lai

d

Tii

curb

ull

yli

cn

cid

~la

lic

acid

Cit

ric

acid

Mal

oni

ca

dd

Mal

ica

cid

F'u

nn

ui«

ac

id

Fu

mari

ca

cid

Fum

nric

ac-i

d

ic Mal

on

icac

id

Fu

mn

ric

aci

d

Tri

curb

ul­

lyli

cac

idM

ali

ca

cid

Tri

c-n

r-bu

l­ly

lic

acid

Ma

lic

acid

Mal

oni

ea

cid

Mal

icac

idF'

um

ui-

ica

dd

c ci3 ;> :> 0'

;.. 8. F '";i ..., o P­ .:: ~ P- er '< en ~ in c en e- :::. :> 5'

O'l o o po '" '"..., <;~ e- '"en "'"O

t-l

458 1'. TRIP.HIII a nd V. S. TRIPATIII

degeneration of malonic ac id was marked in th e absence of any spot in the chromato­gram. At thi s stage formation of t r icarba llylic acid wa s obse rved along with for mat ionof malic acid. Twelve days' exposure of the mixture and then chromatographyrevealed presence of tricarballylic and malic acids in all the samples. Cit r ic acidwas found where amine was added afte r eight and t welve hours' expos ure to sunlightand subsequent exposure of twelve days. Th e later formation of the t wo acids t ri­carballylic and citric was of importance, due to their giving negative tests in thesamples , exposed for smaller periods afte r addition of the am ine.

p -Pheny lene diamine, on addition to mixture solut ions, resulted in formation ofma lonic and malic acids in four days' exposed samples , except in that in which th eam ine was added at zero hour exposure. Fumari c acid was obser ved in twelve andsixteen hour samples.

Addition of p -phenylene d ia rn ine at later stages of exposure dissociated themalonic and malic acids in practically all the samples studied. Fumaric acid wasobserve d for the first time at eight hours, and it was again found in samples in whicham ine was added after twelve hours and subsequently exposing for eight days. Th edegeneration process continued in twelve days' exposed samples with absence ofacids in all the mixture tubes, except in those in which p .phenylene diamine wasadded at eight and twelve hours of exposure.

A marked difference in mixtures, containing the two amines , was absence offumaric acid in tubes in which p.nit roaniline was added , while p-phenylene did notgenerate format ion of t ricarballylic or cit ric acids. In the latter mixture set, t hedegeneration process was accelerated with t ime of exposure .

References

BAllADUR, K.: N ature (Land.) 173 (1954),1141.zu. Bakt. II II7 (1964), 58 5 - 602.and RA~GANAYAKI, S.: Vij ana P a rish ud Anusandhan P at rika 9 (4) (196 6), 171 - 182.- J . Brit. Int erplan et ary Soc . 23 (1 2) (1970 ),813 -820.a nd SIKGH, Y. P. : U n published (1971).

B URTOK, H. S. : Nat u re (Land.) 173 (1954), 127.HALDAKE, J . B. S. : Ration al ist s Ann. 1929, 148.K ALYAKKAR, G. A., KRISHlS"ASWA~lY, P. u., and SREE" IVASAYA, M.: Curl'. Sci. (India ) 21 ( 1052 ),

220 -222.PA VLOVSKAYA, T. E., an d P ASYNSKII, A. G.: Proc. First Intern. Syrn p. " The Ori gin of Life on t he

Earth", Moscow, August 19- 24,1 957. Pergamon Press, L ondon 1957, 151-157.STARK, J. B ., GOODBAN, A. E., and OWENS, H. S.: Anal. Chern. 23 (19 51),413-415.THIPATHI, P., and THIPATHI, V. S.: Under communication (1976).- - Zbl. Bakt. II 133 (1978),745 -748.

Authors' address:

Dr. (Mrs.) PARVEENTHIPATHI. c/o Dr. V. S. TRIPATHI, Chern. Dept., All ahabad University,Allahab ad· 2II 002, India.

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