Gelatin-A Compound for All Reasons Tarasankar Pal Indian Institute of Technology, Kharagpur 721 302, India Gelatin, one of the most basic and most important biological compounds, has been well-known since the early days of protein chemistry. Recently, gelatin system has been exploited successfully as a reagent for various chemical reactions, luminescence enhancement and Raman detection. Gelatin is a simple hydrophilic protein-like eggalbumin with the isoelectric pH of 4.74. Gelatin has long been used as a gel-forming agent in the food industry and as a binder of silver halides in the photographic industry As a long-chain (molecular weight 10,000)protein, gelatin also is used as polarographic wave suppressor in electrochemistry. But until recently, gelatin never has been exploited as a complexing agent for a myriad of applications ranging from pharmaceutical to environmental analysis, and early cancer detection. Recently, the authors discovered that gelatin may produce effective reagents, for a wide variety of applications in chemistry. Following the observation by West et al. ( 1 ) on the utility of sufamoyl benzoate, in carbon monoxide determination, it was seen that gelatin is a more efficient chemical for the photometric determination of carbon monoxide (2).This idea gave birth to over two dozen scientific articles with the same underlying principle that gelatm binds silver(1) and gold(II1) ions in alkaline medium with the production of water soluble complex in each case. The high affinity of gelatin binding with metal ions was demonstrated by uptake of silver(1) or gold(II1) ion from their freshly precipitated halides at the alkaline pH (d1.1).The metal-gelatin complexation was verified by the dialysis and ultrasonic velocity measurements in aqueous medium (2). The silver(1) or gold(II1) gelatin complexes were used to determine the trace concentration of ascorbic acid (31, hydrazine (4),formaldehyde (5),hydrogen sulfide (61, carbon monoxide (21, etc. All these are capable of reducing the gelatin complexes to produce metals. Confirmation is done from standard redox potential values. One interesting aspect of all these redox reactions is that they may be studied by a simple spectrophotometric technique. Metals produced as reduction products behave as exceptional sol particles and are stabilized in situ by gelatin. Hence, trace analysis of reducing substances using absorption properties of the highly stable sol solution has been pursued at some length with success. Silver and gold sols can be stabilized in gelatin according to the following schemes
or Gold (Ill)-gelatin complex
Redudng agents
Gelatin stabilized silver or gold sol
having ).max4~s
nm and 540 nm. respectively
The observation clearly underlined superiority of gelatin over the other methods dealing with silvedl) ions in nonprotein complexing agent. For example, the aqueous solutions of silver(1)-gelatincomplex may be impregnated on silica gel, dried under vacuum, and the solid reagent placed in glass tubes. Thus, the packed column is applicable to selective evaluation of hydrogen sulfide by column
Excitation wavelength 300nm
P
WAVELENGTH (nm)
Figure 1. Enhancement of RTF spectrum of BaP using gelatin evaluation (CE) technique under field conditions. Such analytical determinations of reducing substances require mM concentrations of metal ions. Accordingly, the eost of the system would be reasonable. In addition, the silver or gold sol may be precipitated easily for recovery upon addition of electrolyte (6). Similarly, methods of determination of other compounds of interest have been developed reusing gelatin-stdbilized sols. Thcsc methods exploit the sol-stabilizing properties .. . of gelatin. As cyanide ion and oxygen, hydrogen peroxide and ammonia, or hypochlorite in water dissolve the sol system, the determination of dissolved oxygen (DO), cyanide, hydrogen peroxide and hypochlorite in water, can be performed as shown in the following schemes. A Determination of dissolved oxygen and cyanide Silver or gold sol (colored)
cyanide in air
or 0,
z
Argentocyanide or auroeyanide (wlorless)
B. Determination of hydrogen peroxide Silver sol (colored)
>
Silver amine complex
C. Determination of hypochlorite Silver sol (colored) hYFoeh'olite,Silver d o r i d e precipitated Reactions may be followed simply, using spectrophotometry, titrimetry, and/or conductometry. Thus, gelatm again proves to be a useful reagent for the analytical chemist. Volume 71 Number 8 August 1994
679
Novel uses of gelatin recently I I I I I I I have opened new horizons for applications in environmental analysis and detection of mutagenic chemicals with DNA. 5 Room temperature fluorescence E (RTF) and room temperature phosphorescence (RTP) spectra E of benzo(a1pyrene (BP), benzopyrene tetrols (BPT) and pyrene (Py)have been examined on gelatin-treated paper for the first time (Figs. 1and 2, respectively). The RTP technique is relatively new, but it is a powerful and sim- w ple approach for chemical analy- 5 sis that does not reauire lowtemperature matiix and cryogenic equipment as does conventional phosphorimetry (8). Using gelatin- significant enhancement of RTP signals were observed. In most cases the enhancements enabled us to determine BP and BPT concentrations down to lo-' moles on filter paper substrate. These observations have important implications in the field of Figure 3. SERS of pyridine in gelatin-stabilizedsilver sol. Krypton (647.lnm)laser, 140 mW environmental and health effect assessment. Carcinogenic adivity of many polynuclear aromatic have shown that some PN& bind covalently to DNA (11). (PNA) compounds such as BP, BPT, etc. (9,10) has been a Several C'Jm~oundsof great interest to toxiC'Jlogisf-Sand focal point for concern about human exposure to these specancer researchers are b e n z o ( a ) ~ ~ r e n(BPI e and ties in the workplace. Polynudear aromatic compounds, beuo(a)p~rene tetrol (BPT). BPT is a hydrolyzed ~roduct of organic materials, products of incomplete of BPDE-DNA adduct. Studies have shown that they are also are distributed widely in the environment. Studies activated metabolically to electrophilic intermediates, which bind covalently to DNA (11).Because the carcinogenic activity of a compound might be associated, with bindine to DNA. there is need for analvtical techniaues that a& capable bf detecting ~ ~ ~ - c a r c i n ;interactibns. ~en Monitorinetechniaues for potential exposure to these compounds alco are i n i m a n d . Figure 2 iilustrarps the signal enhancement of gelatin for the RTP detection of BM' adsorbed on fdter paper. Thus,emission spectroscopy gains a new dimension for the determination of PNA's. Apart from improving the sensitivity of RTF and RTP for chemical analysis, described above, MI)-gelatin complex could be used as an efficient phenol-coupling reagent (12). opening a new dimension in synthesis of natural polyph&olic~.Strikingly, application bf the above reagent or naturally occurring phenolic anto mmple . phenol . thraquinone produced the desired products without polymerization. This phenomenon may be explained by the slow release of reducible A..d ) ion from the com~lex . eelatin aggregate in the reaction medium. A dynamic equilibrium mav be cited for the slow release of silvertl, ion as shown below:
-
= 2 -
-
-
-
Ag+ + H2N. . . COOH + O K = H2N. . .COOAg + H,O and Ha.
1 1 1 1 I I I I I 440
500
560
620
Figure 2. Enhancement of RTP spectrum of BPT using gelatinand TLOAcExcitation wavelength was 350 nm. Journal of Chemical Education
Alternatively, A ~ H N. . . COO- + OH- = [ ~ g N. - . .c o o l 2 + H,O
680
WAVELENGTH (nrn)
680
. . COO& + O K = AgHN.. . coo- + H20
and [Ag-N. . . c o o l Z + H 2 0 = Agt
+ OH- + HN. . .COO-
The complexation of M I ) ion and its release may be verified by pHmetry and confirmed by conductometry.Just
Multiple Application of Gelatin
Process Complexation with silver(1)ion
Complexation with gold(ill)ion Dissolution of silver sol Dissolution of gold sol
Analysis
Applications
Vitamin C (37
Pharmaceuticals and fruits
Phenol coupling (12)
Organic synthesis
HzS (6), NzH4 (4). CO (3, HCHO (19).Ad11 (16) Vitamin C (17)
Field study, water samples. and photographic paper Pharaceuticals and fruits
H202(16).OCr(19).DO(20). Environmental analysis CN(21) . . Environmental analysis CN(ZZ), DO (231
Stabilization of silver sol SERS (24) RTF (7) and RTP (7) Cellulose substrate
Environmental analysis Early cancer detection and environmental analysis
the silica-gel impregnated silver gelatin complex may be used in a column evaluation technique, a similar solid reagent also may be used for phenol-coupling reactions in owanic solvents, thus enabling simple separation of reactioh product from the reagent silver(1)-gelatin complex. In the past decade, a renewed interest has been developed in &man spectroscopy resulting from observations that enhancement in the Raman-scattering signals occurs when a com~oundis adsorbed onto s~ecialmetal surfaces having sub&ron structure (13). ~ h ksurface-enhanced , Raman Scattering (SEW) effect has value for generating structural information. However, one substantial obstacle to S E W in eeneratine the s~ecializedsurfaces varticularly tremenldous d i f i h t y oi preparation and sLbilization of metal sols at room temperature. That challenge has been met to some extent for the first time using gelatin in e sol and gel media both are utilized aaueous ~ h a s whereby tidetermine the trace concengation of polarizable molecules using SERS. Gelatin stabilized silver and gold sols gave rise to intense peaks (Fig. 31 for a wide variety of compounds. Celatin does not hinder the scattering spectral omfile of the analvte. Additionallv.. sol solutions in eelatin ;an be stored for Aonths. In view of the manifold application of gelatin as discussed earlier, one may think aloud to produce cost-effec-
.
-
tive metal sol system (14, 15)in gelatin to be taken intermuscularly to combat diseases like arthritis. Trace metal deficiencies in our body may be supplemented by the protein-metal complexes to correct enzymatic action. Intoxication by carbon monoxide, cyanide, etc. may be remediated by protein-metal complexes as well. Gelatin has a mvriad ofbeneficial uses in pharmaceutical, environmental, and biomedical areas. It is auite roba able that this ubiquitous compo&d wiil be useful as a therapeutic agent in addition to its applications in chemist~y.Truly gelatin applications are likely to see renewed interest as the versatility of this protein comes to light. Some of these applications are surveyed in the table. Acknowledgment I wish to thank T. Vo-Dinh, A. Pal and K. Houck for their invaluable suggestions and help for the completion of this work.
Literature Cited 1. Bell, D . R; Reisme., K D;Wed,P. W.Ad.Chim.Arto. 1976.77.245-254 2. Pal,T: Ganpuly,A,Maity, D. S. BuN. Chom Sm. 1888,60,3W1-%44. 3. Pal,T.;Maity, D . SAnd. h t t . 19(Ui,18,1131-1142. 4. Pal. .T:.Ganeulv.A.:Maihi - .. . ..D. S h l v a f lsB6.111.1413-1415. . . 5. 6. 7. 8. 9. 10. 11, 12. 13. 14
IS 16. 17. 19. 19. 20 21 22.
Pal T; Dm, K h l . htt.1881,20,2011-2020. Pal.T;Ganplb,A.:Maity,D. S.Analyst 1988. 111,691493.
Vo.Dinh.T: WI,T.Unpublished results. Vo.Dinh. T. Room Tompomtm Phmphwimlry f ~ ~ C h e m I m Anolysia; l W~lw:Near Y0rk 1 w . Vo-Dmh, T. ChrmimiAn.zlysb bfPdyeylieAmMtie CCPP* Wi1ey:New Yo& 1988. Searle, C. E., E d Chemical C o r r i w n s ; A C S :Wmhinpton. DC,1978. Grirnrnrr G., Ed. Enuimnmntnl corrinwem: Polycyclir h m n t i e H y d m r b o M ; CRC Ress: BmaRatrm, FL. 1983. Pa1.TPal, A. J.lnd. Chem. Sm. 1888.66. 256238. Ch-. R. K:Furtak.. T. E... Eds. Surfme-Enhonwd Rnmnn S m t w-.~ Plenum . h.*sN c u Y d 1982 VykydaL .U; Klahuaay.L.haury*y.K.Anwtrnlt(od-+asrh 1868.6.56&571 Cannon.C . W . Cannon.~.B C,Ward..J R. Jnhpumoml. l W . 1 3 . 5 2 6 1 Pal,T; Maits D. S . h l y a t 1888,111.4%51. Pal.T; Dm, K.Anol.htt. 1888.21.2333-2143. Pal.T;Ganguly,AMeity.D. S.lWhlonta 1988,35,658486 Pal.EGanwIv. . . A. Mait7.D. S. Chem.Ano1. 1988, 33.703-708 P d . T Oaa. I' K Anohar 1988 113. IMll-16m Pal. T , C a w l ~ . Ahla1ty.D. , SAN11.(hem 1988.58. 15W1566. P d . T C a n e u l v . A . h l r s , 1881. 112.1327-1329
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