Tricaine-S

Western Chemical Inc.
Western Chemical Inc.

TRICAINE-S Brand of Tricaine Methanesulfonate For Anesthesia and Tranquilization of Fishes and Other Cold-Blooded Animals

FULL PRESCRIBING INFORMATION: CONTENTS*

• Description
• Warnings
• Toxicology
• Precautions
• References
• Directions for Use
• Tricaine-S Label

FULL PRESCRIBING INFORMATION

Description

TRICAINE-S is intended for the temporary immobilization of fish, amphibians, and other aquatic, cold-blooded animals. It has long been recognized as a valuable tool for the proper handling of these animals during manual spawning (fish stripping), weighing, measuring, marking, surgical operations, transport, photography, and research.

Chemistry

TRICAINE-S is the methanesulfonate of meta-amino benzoic acid ethylester, or simply ethyl m-amino benzoate. It is thus an isomer of benzocaine having the formula C9H11O2N + CH3SO3H

TRICAINE-S is a fine white crystalline powder. Its molecular weight is 261.3. Soluble to 11%, it forms clear, colorless acid solutions in water.

Warnings

Do not use within 21 days of harvesting fish for food.

When used in food fish, use should be restricted to Ictaluridae, Salmonidae, Esocidae, and Percidae and water temperature should exceed 10° C. (50° F.).

In other fish and other cold-blooded animals (poikilotherms), TRICAINE-S should be limited to hatchery or laboratory use.

Toxicology

Precautions

1. Avoid inhaling TRICAINE-S or getting it into the eyes.
2. Always conduct preliminary tests with TRICAINE-S to determine desired rates of anesthesia and optimal length of exposure.
3. Do not overexpose fish to lethal levels of TRICAINE-S.
4. Do not anesthetize more fish than can be handled effectively.
5. Do not contaminate eggs or sperm with TRICAINE-S when stripping fish.
6. Do note use water containing chlorine, or other toxic agents.
7. Insure adequate oxygen in anesthetic solution.
8. Discard anesthetic solutions when fouled with mucus or metabolic wastes.
9. Do not discard TRICAINE-S solutions into water supplies of natural waters.
10. Store TRICAINE-S solutions in a cool place away from light.*
11. Discard stock solutions of TRICAINE-S after several days.*
12. Treated fish destined for food must be held in fresh water above 10°C. (50°F.) for 21 days before use.

*The color of TRICAINE-S solutions may change rapidly to yellow or brown when exposed to light. This does not affect activity in any significant way. However, for best results use freshly prepared solutions. A 10 per cent solution stored at room temperature shows no significant loss of potency after three days, but after 10 days, a brownish color and an activity decrease of about 5 per cent is observed.

References

1. Marking, L.L.: Investigations in Fish Control. 12. Toxicity of MS-222 to Selected Fishes, U.S. Bureau of Sport Fisheries and Wildlife, Resource Publication 18, 1966.
2. Schoettger, R.A., Walker, C.R., marking, L.L., and Julin, A.M.: MS-222 as an Anesthetic for Channel Catfish; its Toxicity, Efficacy, and Muscle Residues, U.S.Bureau of Sport Fisheries and Wildlife, Resource Publication 33, 1967.
3. Personal communications:
    a. Bernheimer, W.M., New York University College of Medicine, NewYork, N.Y.
    b. Butler, E.G., Princeton University, Dept. of Biology, Princeton, N.J.
    c. Dalton, H.D., and Charipper, H.A., Washington Square College, Dept.Of Biology, New York, N.Y.
    d. Etkin, W., City College, Dept. Of Biology, New York, N.Y.
    e. Goss, R.J., Brown University, Providence, R.I.
    f. Kollros, J.J., State University, Iowa City, Iowa.
    g. Manner, H.W.: Anaesthetize those planaria. Turtox New 35:135, 1957.
    h. Rose, S.M., University of Illinois, Urbana, Ill.
    i. Schatzmann, J.H., Harvard Medical School, Boston, Mass.
    j. Taylor, A.C., Rockefeller Institute of Medical Research, New York, N.Y.
    k. Thornton, C.S., Kenyon College, Dept. of Biology, Gambier, Ohio.
    l. Van Stone, J.M., Trinity College, Dept. of Biology, Hartford, Conn. Cited in Bove, F.J.: MS-222 Sandoz-the anesthetic of choice for fish and other cold-blooded organisms, Sandoz News, no. 3. 12;., 1962.
4. Schoettger, R.A., and Julin, A.M.: Investigations in Fish control: 13. Efficacy of MS-222 as an Anesthetic on Four Salmonids, U.S. Bureau of Sport Fisheries and Wildlife, Resource Publication 19, 1966.
5. Schottger, R.A.: Efficacy of MS-222 as an Anesthetic for Northern Pike, Muskellunge and Walleye, U.S. Bureau of Sport Fisheries and Wildlife, to be published.
6. Knight, A.E.: Intracellular hemoglobin crystallization in two centrarchid, the large-mouth bass and the bluegill, Progressive Fish-Culturist 26:115 (no. 3) 1964.
7. Lumb, W.V.:Anesthesia of Laboratory and Zoo Animals, in: Small Animal Anesthesia, Philadelphia, Lea and Febiger, 1963, pp. 269-310.
8. Webb, R.T., Distribution of Bluegill treated with tricaine methanesulfonate (MS- 222), Progressive Fish-Culturist 20:69 (no. 2) 1958.
9. Klontz, G.W.: Anesthesia f fishes, Proceedings of the Symposium on Experimental Animal Anesthesiology, Brooks Air Force Base, Dec. 14-16, 13 p., 1964.
10. Gilbert, P.W., and Wood, F.G.: Methods of anaesthetizing large sharks and rays safely and rapidly. Science 126:212, 1957.
11. Mann, H., and Rajbanshi, K.G.: Anesthetic and Tranquilizer for Fish, Frogs and other Cold-blooded Organisms, Sandiz Bulletin No. 3350/182 e. Basie, Switzerland.
12. Tuumanen, P.: Experiments with MS-222 Sandoz in the Shipment of Live Trout in Plastic Pouches,Kalataloudellisen tukimostoimiston, Tiedomantoja, no.2, 1966.
13. Lemarque, P.,: Anesthesie et transport, Bull. Inf. Cons. Sup. Peche 55:5, 1964.
14. Walker, C.R., and Schoettger, R.A.: Investigations in Fish Control: 15. Residues of MS-222 in Four Salmonids Following Anesthesia, U.S. Bureau of Sport Fisheries and Wildlife, Resource Publication 21, 1966.
15. Correspondence: Bureau of Fisheries, U.S. Department of Interior, 1968.

Directions for Use

I. DIRECTIONS FOR USE ON FISH

CONCENTRATIONS
TRICAINE-S is effective and safe for the anesthesia of fish when used as directed. Its use is governed by, and can be tailored to, the needs of individual fishery personnel. Sedation and various rates of anesthetization are controlled by the concentration. The versatility of TRICAINE-S is demonstrated by the fact that it has been used in fisheries at levels ranging from 10 to 1,000 mg/liter³. The action of the anesthetic is slowed at cooler temperatures, in extremely soft water (approximately 10 mg/liter of CaCO3, or less), and in larger fish⁴. Also, efficacy may vary with species⁴.   Thus, it is imperative that preliminary tests of anesthetic to determines the desired rates of anesthesia and exposure times for the specific lots of fish under prevailing conditions.

The following tables may be used as guidelines in selecting concentrations of TRICAINE-S for the anesthetization of various fishes:

Table 1: Concentration Required for Rapid Anesthesia
(Induction time less than 2-5 minutes; used in spawning, marking, measuring, and some surgical operations)

* Maximum tolerated exposure (in minutes) of fish to TRICAINE-S solution.

Table 2: Concentration Required for Moderately Rapid Anesthesia
(Induction time less than 15-20 minutes; used in surgical operations and in spawning and marking where longer exposures are more important than rapid immobilization)

*Maximum tolerated exposure time (in minutes) of fish to TRICAINE-S solution.

Table 3: Concentrations Required for Sedation
(Induction within 15 minutes; used in fish transport)

IMPORTANT: Since, in many cases, relatively rapid rates of anesthesia can be achieved only by exceeding the lethal concentration of TRICAINE-S, it is necessary to return anesthetized fish to fresh water before they are overexposed.  Excessive exposures are avoided by observing the following sensory and motor responses of the fish which characterize progressively deeper levels of anesthesia.
    Sedation - Decreased reactivity to visual and vibrational stimuli; opercular activity reduced.
    Total loss of equilibrium - Fish turns over; locomotion increases; fish swims or extends fins in response to pressure on caudal fin or peduncle.
    Total loss of reflex - No response to pressure on caudal fin or peduncle; opercular rate slow and erratic.
    Medullary collapse - Opercular activity ceases.

Laboratory and field investigations^3,9, have shown that the action of TRICAINE-S is readily reversed when the fish are transferred to fresh water before opercular activity ceases. Additional exposure following medullary collapse may result in mortality. A rough estimate of the safe total exposure can be made by multiplying the time required for anesthesia by a factor of 2 or 3.

WATER
Since TRICAINE-S is very soluble (1:9) in water, it dissolves with equal readiness in spring water, tap water, or seawater. Do not use distilled or deionized water, or water containing chlorine, heavy metals (copper, zinc, etc.), or other toxic contaminants. The anesthetic solution should be well oxygenated, and its temperature should be similar to that of the water from which fish are taken. In the field, many water quality problems are eliminated by using natural water to which the fish are acclimated, provided the water does not possess high chemical or biologic oxygen demand.

METHODS OF APPLICATION
1. General anesthesia: - For most situations where rapid or moderately rapid anesthesia is required, TRICAINE-S may be applied in a bath, i.e., the fish are immersed in the anesthetic solution. Containers may be of glass, plastic, steel, aluminum, or other suitable material. However, do not use galvanized or brass containers unless treated or sealed to prevent dissolution of zinc. Size of container is determined by individual needs, but the fish should not be overcrowded. Discard anesthetic solutions when a loss in potency is noted, or when the solutions become fouled with mucus or excrement.
2. For surgery and certain physiologic studies, the fish may be anesthetized to loss of reflex, removed from the anesthetic, and then positioned so that the gills are bathed in a sedating concentration of TRICAINE-S. Some investigators have developed flowing, recirculating systems for bathing the gills with anesthetic during surgery.

Large fishes such as sharks and rays are anesthetized within minutes by spraying the gills with a 1g/liter solution of TRICAINE-S¹⁰ . The application is made by means of a water pistol, bulb syringe, hand pump, etc.

3. Transport - TRICAINE-S has been used to sedate fish during transport. It is more successful in cold than in warm water, and it is instrumental in reducing injuries because of hyperactivity. Fish are usually transported by means of distribution units (tank trucks), or by air in plastic bags^11,12. In either case, the fish should be fasted before-hand to reduce metabolic wastes. Also, some workers suggest pre-transport sedation for several hours to lower metabolism.  With distribution units, the fish may be fasted and sedated prior to loading. The anesthetic solution is prepared in the distribution unit and oxygenated. Then, the fish are added and temperature acclimated.  In air shipments, the anesthetic solution is placed in a suitable plastic bag, the sedated fish are added, the bag inflated with oxygen, tied securely, and placed in a second bag. This bag is also tied, and then placed on ice in an insulated container¹³. A modification of this method involves complete anesthesia of the fish, and placing them in water bags which contain no anesthetic. In any case, upon arrival, the fish should be acclimated slowly to new environmental temperatures.

PREPARATION OF TRICAINE-S SOLUTIONS
Prior to use, TRICAINE-S may be weighed out into amounts which are convenient for the volume of water to be used. A handy unit is 2 g. since this quantity in 5 gallons of water yields a concentration of about 100 mg/liter. For rough approximations, one level teaspoonful contains 2.0 to 2.5 g. Thus a level teaspoonful of anesthetic in 5 gallons gives a concentration of about 120 mg/liter.

To convert mg/liter into g/gal.: multiply number of mg. by 0.00378
    e.g. 80 mg/liter = 80 x 0.00378 = 0.302 g./gal.
To convert mg/liter into a ratio of TRICAINE-S to water: divide 1,000,000 by the number of mg.
    e.g. 80 mg/liter = 1,000,000 / 80 = 1:12,500

LIMITATIONS IN USE
Since TRICAINE-S is taken up into the blood of fish, residues of the drug may occur in edible tissues. However, the residues dissipate rapidly after the fish are placed in fresh water¹⁴. Thus, treated fish which may be used for food must be held in fresh water above 10°C. (50°F.) for a period of 21 days. 

Withdrawal in fresh water is unnecessary for non food fishes such as goldfish, bait fish, and ornamentals. Also, withdrawal is unnecessary for sublegal sizes of the following species of fish because they are not used as food immediately following anesthesia (Table 4).

Table 4 - Sublegal Sizes of Fish Species not used as Food Immediately after Anesthesia¹⁵

II. GUIDELINES FOR USE ON AMPHIBIANS

Table 5. Effects of Varying Concentrations of TRICAINE-S on Salamanders

* When an individual of any of the species listed is exposed at the designated concentration, the data available suggest that the animal may be safely maintained under anesthesia for the time noted. Prolonging exposure to the anesthetic beyond the time indicated may cause deaths. See PRECAUTIONS.

Table 6 - Effects of Varying Concentrations of TRICAINE-S on Frogs

* When an individual of any of the species listed is exposed at the designated concentration, the data available suggest that the animal may be safely maintained under anesthesia for the time noted. Prolonging exposure to the anesthetic beyond the time indicated may cause deaths. See PRECAUTIONS.

Tricaine-S Label