[NHCOLL-L:4602] Re: Use of isopropyl for fixing and storing invertebrates
H.J. Walker
hjwalker at ucsd.edu
Mon Nov 9 21:42:14 EST 2009
We also have a blennioid expert here who agrees that in some cases (not
all), his fishes stored here in isopropanol can have less distinct
melanophore patterns. Again, he does not regard it as a big deal, one
reason being that all the other colors are lost anyway. However, with
millions of specimens representing thousands of species, we do not have
problems with fading.
Williams, Jeff wrote:
> I can attest to the fact that blennies do not survive well in isopropanol and many of them tend to lose all or most of their color pattern in isopropanol, while this degree of fading is not seen in ethanol stored blennies. About 20 years ago I found some blennies at the USNM that had been collected in 1975 by Jack Randall and had been sent to Vic Springer for ID. Vic placed his specimens in 70% ethanol. Jack kept other specimens from the same collection at BPBM and stored them in isopropanol (I am not sure what concentration they used). When I compared the BPBM and USNM specimens, the BPBM specimens were uniformly brown and badly faded while the ethanol stored USNM specimens had retained the color (melanophore) pattern of the species. Unfortunately I lost track of those lots and have not been able to find them again.
>
> The old RUSI (now SAIAB) isopropanol collections show the same fading and loss of pattern for their blennies as the BPBM specimens. While not every fish will fade in isopropanol, many of them do, and that should be reason enough to avoid it if possible.
>
> Jeff
> email: williamsjt at si.edu
> Phone: 301-238-1736
> Fax: 413-618-1138 or 301-238-1834
> http://vertebrates.si.edu/fishes/fishes_staff_pages/williamsjt.cfm
>
> Dr. Jeffrey T. Williams
> MSC, Fishes, Nat. Mus. Natural Hist.
> 4210 Silver Hill Road
> Suitland, MD 20746
>
>
> -----Original Message-----
> From: owner-nhcoll-l at lists.yale.edu [mailto:owner-nhcoll-l at lists.yale.edu] On Behalf Of H.J. Walker
> Sent: Friday, November 06, 2009 4:16 PM
> To: simmons.johne at gmail.com
> Cc: amsnyder at unm.edu; nhcoll-l at lists.yale.edu; ryan.evans at ky.gov; brian.lang at state.nm.us
> Subject: [NHCOLL-L:4598] Re: Use of isopropyl for fixing and storing invertebrates
>
> Hello All,
> Part of Ryan's plan, which John did not address, is fixation in
> isopropanol and neither alcohol should be used as a fixative. However,
> if formaldehyde is not available, high concentrations of ethanol (ca 70%
> or greater) have been used. I agree with John in questioning the need
> to buffer the alcohol.
>
> John does a fine job explaining the isopropanol/ethanol comparison for
> preservation, but for fishes there are millions of specimens preserved
> for 65 years or more in isopropanol and many of the detracting
> characteristics mentioned below are negligible for most scientific
> purposes. Rather than a point-by-point discussion, it should be noted,
> for example, that studies have been conducted showing more shrinkage in
> ethanol, and those millions of specimens (above) have not cleared in
> concentrations of 45% isopropanol. (50% is more than adequate for
> fishes.)
>
> Best wishes,
> H.J.
>
>
> John E Simmons wrote:
>
>> Lex and Ryan
>> Isopropyl can be used as a preservative,but be aware that it has
>> problems. I have inserted a section of text below from the book I am
>> writing on fluid preservation that summarizes the literature on
>> isopropyl as a preservative.
>>
>> I would not use isopropyl at 50% as it can clear tissues at around 45%
>> or less. If I had to use it, I would use a 70% solution mixed with
>> deionized or distilled water.
>>
>> If I may ask, why are you buffering the alcohol, and to what pH? Most
>> alcohol is slightly acidic (which makes a better preservative). I
>> would be concerned, particularly with isopropyl, about making too
>> basic a solution which could clear tissues. This could particularly
>> be an issue with crayfish and mollusk shells, I would think, as they
>> are going to further buffer the solution, aren't they? In any case,
>> if you do have a good reason to buffer, I would use a more stable
>> buffer than CaCO3.
>>
>> What is the glycerin for? Glycerin acts as a scavenger that will
>> attract relative humidity and any contaminants in the air into your
>> solution if your containers leak.
>>
>> Here is the text from the draft of the book, with citations:
>>
>> Isopropyl alcohol was first made in 1855, but it did not go into
>> commercial production until 1920 in the United States (Hatch 1961).
>> It was recommended as an alternative preservative to ethanol, without
>> being tested, as early as 1922 (Griffin 1922). A notice in /Turtox
>> News/ in 1934 complained of the high taxes on ethanol, the red-tape
>> necessary to buy it tax-free, and the additives used to produce
>> denatured ethanol. The recommended solution was isopropyl alcohol.
>> The recommendation for its use came despite the fact that "Sufficient
>> experimentation has not been carried on in connection with its use in
>> biological work to warrant a dogmatic statement from us that it can
>> /always/ be used as a substitute, but our preliminary work justifies
>> this /belief/" (Windsor 1934:216). There was a conflict-of-interest
>> in the recommendation to change to isopropyl alcohol: "Where can you
>> buy isopropanol? Turtox will gladly accept your orders for it"
>> (Windsor 1934:217). At that time, anhydrous isopropanol was $3.75 per
>> gallon (for an 88% solution), but "the grade used for general
>> preservation" was just $2.50/gallon (Windsor 1934:217).
>>
>>
>> In 1922, Philip Pope preserved one specimen of a newt each in jars of
>> 40%, 50%, 60%, 70% and 80% isopropanol, with one specimen in 70%
>> ethanol for a control (Pope 1928). Pope examined the newts after a
>> mere six years in the preservative, and decided that the 40-50%
>> isopropyl preserved specimens were fine, even though he could not find
>> the control to compare them to. The advantages that Pope listed for
>> isopropyl were that it had no restrictions, was inexpensive, could be
>> used in low concentrations, was not irritating like formaldehyde, and
>> did not soften bones or teeth like formaldehyde (Pope 1928). However,
>> 45-50% isopropanol has been shown to cause considerable shrinkage of
>> specimens, it can be difficult to mix thoroughly, it is prone to
>> layering in specimen containers, it may soften bone, and it may cause
>> clearing of tissues if concentrations fall below 30-40% (Fink et al.
>> 1979). DiStefano et al. (1994) found 50% isopropyl failed to prevent
>> tissue decay in fish and crayfish. The permeability of the epidermis
>> (the solute absorbed per ml of tissue) for methanol is 0.6, for
>> ethanol 0.9, and for propanol 1.3, thus isopropyl penetrates epidermis
>> best (Schaefer et al. 1982). It is important to note that isopropyl
>> is twice as toxic as ethanol (Monick 1968).
>>
>>
>> King and Porter (2004) reported observational data that long-term
>> storage in isopropyl alcohol results in greater fading of ant
>> specimens, probably due to the greater propensity of isopropyl and
>> other long-chain alcohols to extract lipids, proteins, and pigments
>> from cellular membranes at high concentrations (Goates and Knutson
>> 1994). Some workers, particularly ichthyologists, have reported that
>> they prefer isopropyl preserved specimens because they are more
>> flexible than alcohol preserved specimens (Fink et al. 1979, Walker et
>> al. 1995); however, isopropyl preserved specimens are flexible because
>> the tissue matrix has undergone more breakdown that tissues do in
>> ethanol (Stoddard 1989); in an accelerating aging study, Von Endt
>> (2000) found that isopropyl induces more deterioration of collagen
>> than ethanol.
>>
>>
>> When isopropyl is mixed with water, the resulting solution undergoes
>> contraction, so solutions should be mixed on a weight basis rather
>> than a volume basis (Hatch 1961).
>>
>>
>>
>> All preserving alcohols cause tissue shrinkage, but the amount of
>> shrinkage is variable, with less shrinkage occurring in methyl and
>> more in isopropyl compared to ethyl alcohol (Ciferri 1971).
>>
>>
>> Ciferri, A. 1971. Swelling and phase transition of insoluble
>> collagen. Pages 101-151 _in_ Elden, H. R. (editor). /Biophysical
>> Properties of the Skin/. John Wiley and Sons, Inc., New York, vii +
>> 645 pages.
>>
>> DiStefano, R.J., M.J. Roell, B.A. Wagner, and J.J. Decoske. 1994.
>> Relative performances of four preservatives on fish and crayfish.
>> /Transactions of the American Fisheries Society/ 123:817-823
>>
>> Fink, W.L., K.E. Hartel, W.G. Saul, E.M. Koon, and E.O. Wiley. 1979.
>> A report on current supplies and practices used in curation of
>> ichthyological collections. American Society of Ichthyologists and
>> Herpetologists ad hoc Subcommittee Report, 63 pages.
>>
>> Goates, C.Y. and K. Knutson. 1994. Enhanced permeation of polar
>> compounds through human epidermis. I. Permeability and membrane
>> structural changes in the presence of shorter chain alcohols.
>> /Biochimica et Biophysica Acta/ 1195:169-179.
>>
>> Griffin, L.F. 1922. Practicable substitutes for grain alcohol.
>> /Science/ 55(1419):262-263.
>>
>> Hatch, L.F. 1961. /Isopropyl Alcohol/. McGraw-Hill Book Company,
>> Inc., New York, x + 184 pages.
>>
>> King, J.R., and S.D. Porter. 2004. Recommendations on the use of
>> alcohols for preservation of ant specimens (Hymenoptera, Formicidae).
>> /Insect Sociaux/ 51(2):197-202.
>>
>> Monick, J. A. 1968. /Alcohols: Their Chemistry, Properties and
>> Manufacture/. Reinhold Book Corporation, New York, xiv + 594 pages.
>>
>> Pope, P.H. 1928. Isopropyl alcohol as a preservative. /Science/
>> 68(1768):487-488.
>>
>> Schaefer, H., A. Zesch, and G. Stuttgen. 1982. /Skin Permeability/.
>> Springer-Verlag, Berlin, ix + 541-896 pages.
>>
>> Stoddard, R. W. 1989. Fixatives and preservatives: their effects on
>> tissue. Pages 1-25 _in_ Horie, C. V. (editor). /Conservation of
>> Natural History Specimens: Spirit Collections/. The Manchester
>> Museum and Department of Environmental Biology, The University of
>> Manchester, Manchester, viii + 115 pages.
>>
>> Von Endt, D.W. 2000. Staying in shape: the stability of structural
>> proteins in natural history museum storage fluids. /Polymer
>> Preprints/ 41(2):1794-1795.
>>
>> Walker, H.J., G.H. Burgess, A.Y. Suzumoto, C.I. Klepadio, L.
>> VanGuelpen, and J.M. Humphries. 1995. Isopropanol revisited.
>> /Curation Newsletter/ 11:3-4.
>>
>> Windsor, A. S. 1934. Alcohol notes. /Turtox News/ 12(11):215-217.
>>
>>
>> --John
>>
>> John E. Simmons
>> Museologica
>> 128 E. Burnside Street
>> Bellefonte, Pennsylvania 16823-2010
>> simmons.johne at gmail.com <mailto:simmons.johne at gmail.com>
>> 303-681-5708
>> www.museologica.com <http://www.museologica.com>
>> and
>> Adjunct Curator of Collections
>> Earth and Mineral Science Museum & Art Gallery
>> Penn State University
>> 19 Deike Building
>> University Park, Pennsylvania 16802-2709
>> jes67 at psu.edu <mailto:jes67 at psu.edu>
>>
>>
>> On Thu, Nov 5, 2009 at 4:35 PM, Alexandra M Snyder <amsnyder at unm.edu
>> <mailto:amsnyder at unm.edu>> wrote:
>>
>> Any ideas and/or caveats re: use of isopropyl for invertebrate
>> preservation? See letter below. Thank you in advance for
>> information provided. Lex
>>
>> Lex,
>>
>> I just exhausted my last supply of ethanol, so I'll be
>> fixing crayfishes and freshwater mollusks in iso and
>> storing as well (theoretically). It is 99% ACS Iso and as
>> far as I can ascertain using Google, not denatured. The
>> plan for now is to try and buffer (CaCO3) and add a little
>> glycerin as I do with denatured ethanol. My guess was
>> diluting to 50% isopropyl.
>>
>> Ryan
>> Ryan Evans, Aquatic Zoologist
>> Kentucky State Nature Preserves Commission
>> 801 Schenkel Lane, Frankfort, KY 40601
>> 502-573-2886; ryan.evans at ky.gov <mailto:ryan.evans at ky.gov>
>>
>> The world is made for people who aren't cursed with self
>> awareness. - Annie Savoy
>>
>> **************************************
>> Alexandra M Snyder
>> Collections Manager-Fishes
>> Museum of Southwestern Biology MSC03-2020
>> 302 Yale NE
>> University of New Mexico
>> Albuquerque NM 87131 USA
>> PH/FAX 505.277.6005
>> http://www.msb.unm.edu/fishes/index.html--
>>
>>
>>
>
> --
> H.J. Walker, Jr.
> Scripps Institution of Oceanography
> University of California, San Diego 0208
> La Jolla, CA 92093-0208
> USA
> hjwalker at ucsd.edu
> phone:858-534-2199 fax:858-534-5306
>
> Campus street address for FedEx, UPS, DHL:
> 8675 Discovery Way, 224 / 231 Vaughan Hall
> La Jolla, CA 92037
>
> SIO Marine Vertebrate Collection web site:
> http://collections.ucsd.edu/mv/
>
>
--
H.J. Walker, Jr.
Scripps Institution of Oceanography
University of California, San Diego 0208
La Jolla, CA 92093-0208
USA
hjwalker at ucsd.edu
phone:858-534-2199 fax:858-534-5306
Campus street address for FedEx, UPS, DHL:
8675 Discovery Way, 224 / 231 Vaughan Hall
La Jolla, CA 92037
SIO Marine Vertebrate Collection web site:
http://collections.ucsd.edu/mv/
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