Diyala Agricultural Sciences Journal Vol (13) No 2, 2021: 1-9 ISSN: 2073-9524 Mohammed Inad Ghazwan eISSN: 2310-8746 1 Effect of Some Preservative Solutions on Some Morphometric Features of Planiliza abu (Heckel, 1843) Mohammed Inad Ghazwan 1 1Iraqi Natural History Museum and research Center / University of Baghdad Corresponding Author: muhammadinad@yahoo.com Article history: Received 1 April 2021 Accepted 1 June 2021 Abstract The present study attempts to find out the effect of some fish preservatives in the laboratory, such as alcohol and dilute formalin, on some biological characteristics related to the body measurements of those fish preserved in these materials. The fish used in this study were the local Planiliza abu. The processes of expansion and contraction of the bodies of fish preserved in diluted formalin solution at a concentration of 10% and diluted ethyl alcohol solution at a concentration of 70%. As that the standard length of the specimens of this study, which are separately preserved in formalin 10% and alcohol 70%, in a completely isolated are fluctuating in change. Constant shrinkage in head length in both diluted formalin and alcohol. Most fish bodies preserved in formalin at a concentration of 10% gain significant weight gain, in contrast to alcohol preservation. keyword: Biometry, Brackish water, Expansion and contraction, External appearance. Doi: 10.52951/dasj.21130201 Introduction Planiliza abu is one of the Iraqi local fish that belongs to the Mugilidae family. Its total length is 26 cm and distributed in the Tigris and Euphrates basins; it does not enter the Arab Gulf. It is found in large quantities in all water bodies in the middle and South ern Iraq. Its small size reduces its economic value; nevertheless, it is a popular and local fish with wide popularity in Iraqi markets. This species is also found in the Euphrates River Basin in Syria (Misra, 1947; Khalaf, 1961; Mahdi, 1962; Al-Daham, 1984). What distinguishes this species of this family is their bright silver color and tap er in g b o d ie s ( R azz aq et a l, 2 0 1 5 ) . I t is considered one of the coastal tropical fish that live in a salty and little salty environment as shown by ( Th o m s o n , 1 9 9 7 ; Ne l so n , 2 0 0 6 ) . Th e m o st important methods for preserving fishes, in general, are four internationally known methods, which are freezing, preservation in salt, preservation with chemicals represented by diluted formalin and ethyl alcohol. These preservatives may affect some of the biometrics of fish specimens, especially of different lengths (Jawad, 2003). Many standard changes appear on the fish bodies preserved in formalin and alcohol for different standard periods. Evidence come from many researchers as indicated by (Lux, 1960; Parker,1963; Stobo,1972; Engel, 1974; Sayers,1987 and Haubroc et al,2018). These substances may cause contractions and shrinkage in the bodies of the fish preserved in them (Billy, 1982; Al-Hassan and Abdullah, 1992; Al-Hassan and Shawafi, 1997; Al-Hassan et al, 1999, 2000). Some researchers argue that there is a loss in the weight of the specimens preserved in formalin, (Cultter and Whitesel, 1956; Parker, 1963). Billy (1982) says that there is a loss in weight and there is a clear difference between the live weight and the weight of the fish after preserving in both solutions: diluted formalin and alcohol (Banha et al, 2017). Kelvin 1994 believes that it is possible that the concentrations of some elements, especially heavy elements in fish bodies preserved in formalin, may change over time due to formalin acidity and the interaction of these elements over time. Formalin acidity and the interaction lead to an increase in the concentrations of some of them or a decrease in the concentrations of others, or they even lead to the disappearance of these elements due to the variable interactions caused by the long period of preservation in formalin. The elements also vary in their concentrations over time if the fish specimens are preserved with dilute alcohol to a concentration of 70-75%. The effect of these two solutions, formalin, and alcohol, is clear during the first periods of preservation with the disappearance of fish color if the species have clear coloration https://journal.djas.uodiyala.edu.iq/index.php/dasj/article/view/2070 https://orcid.org/0000-0003-2057-5496 Diyala Agricultural Sciences Journal Vol (13) No 2, 2021: 1-9 2 (Dallinger et al, 1987). In this study, we find that these elements are concentrated in the bodies of the fish at a higher rate than those in the water, in their surroundings, or other sediments in this environment, as a result of consuming the organisms in that environment (Olaifa et al, 2004). On the other hand, the presence of these heavy elements in different concentrations in the bodies of fish is a sign of the pollution occurring in the environment of these fish (Jassem et al, 2007). The bodies of fish may be preserved in formalin 10% and then stored in alcohol 70%. Great changes in the bodies of fish, preserved according to this method and for a long time in several forms, will appear as indicated by (Wilke et al. 1996; Arrington and Winemille, 2002). Materials and Methods Ethical Approval All applicable national and international guidelines for the care and use of animals were followed. Experimental Animal A total of 40 P. abu fish were divided into two groups, each group consisted of 20 fish. The first group was kept in a diluted formalin solution at a concentration of 10%; the second group was kept in a diluted ethyl alcohol solution at a concentration of 70%. The two solutions were diluted with non-ionic distilled water. Some biometrics was measured by using the traditional method to calculate the total, standard length, head length, and weight for both groups. The readings were recorded once a week for an entire month. Results and Discussion Figure 1 the local P. abu fish. The general shape and outward appearance of the fish used in this study. Figure 1. Planiliza abu The biometric rates of P. abu fish preserved in alcohol were 70% in the four weeks: 20.18cm was the total length, the standard length was 16.5cm, the head length was 4cm and the weight was 22.8g. However, the biometric rates for fish preserved in formalin were 10% for the four weeks: the total length was 19.6cm, standard length was 16.4cm, the head length was 4cm and the weight was 19.8g. From table 1 we can follow up general linear model between-subjects factors. Then it follows with table 2 the descriptive statistics. Table 3 indicates the analysis of variance of specimens preservative and time in general in addition to the interaction between them. Table 1. general linear model between-subjects factors Value Label N Preservative_Type 1 Alcohol 70% 80 2 Formalin 10% 80 Time 1 1st Week 40 2 2nd Week 40 3 3rd Week 40 4 4th Week 40 Table 2. the descriptive statistics of the experiment Preservative_Type Time Mean Std. Deviation N Total_Length (cm) Alcohol 70% 1st Week 10.325 .4128 20 2nd Week 10.255 .4524 20 3rd Week 10.420 .3901 20 4th Week 10.370 .4402 20 Total 10.342 .4209 80 Formalin 10% 1st Week 9.665 .4258 20 2nd Week 9.980 .4538 20 3rd Week 9.920 .4503 20 4th Week 9.710 .4340 20 Total 9.819 .4531 80 Total 1st Week 9.995 .5320 40 2nd Week 10.118 .4684 40 3rd Week 10.170 .4869 40 Diyala Agricultural Sciences Journal Vol (13) No 2, 2021: 1-9 3 4th Week 10.040 .5458 40 Total 10.081 .5090 160 Standard_Length (cm) Alcohol 70% 1st Week 8.280 .3302 20 2nd Week 8.415 .3588 20 3rd Week 8.545 .3410 20 4th Week 8.310 .3684 20 Total 8.388 .3588 80 Formalin 10% 1st Week 8.120 .3901 20 2nd Week 8.340 .3169 20 3rd Week 8.225 .4339 20 4th Week 8.150 .3763 20 Total 8.209 .3839 80 Total 1st Week 8.200 .3658 40 2nd Week 8.377 .3363 40 3rd Week 8.385 .4179 40 4th Week 8.230 .3764 40 Total 8.298 .3811 160 Head_Length (cm) Alcohol 70% 1st Week 2.070 .1689 20 2nd Week 2.060 .0883 20 3rd Week 2.080 .0894 20 4th Week 2.050 .1539 20 Total 2.065 .1284 80 Formalin 10% 1st Week 1.930 .2296 20 2nd Week 2.075 .0444 20 3rd Week 1.985 .0366 20 4th Week 1.910 .2174 20 Total 1.975 .1703 80 Total 1st Week 2.000 .2112 40 2nd Week 2.068 .0694 40 3rd Week 2.032 .0829 40 4th Week 1.980 .1990 40 Total 2.020 .1569 160 Weight (gm) Alcohol 70% 1st Week 13.950 1.2763 20 2nd Week 11.650 1.2258 20 3rd Week 11.950 1.3945 20 4th Week 14.000 1.3765 20 Total 12.888 1.6988 80 Formalin 10% 1st Week 10.950 1.6376 20 2nd Week 10.100 1.7741 20 3rd Week 10.950 1.5720 20 4th Week 10.900 1.7442 20 Total 10.725 1.6912 80 Total 1st Week 12.450 2.0995 40 2nd Week 10.875 1.6975 40 3rd Week 11.450 1.5517 40 4th Week 12.450 2.2066 40 Total 11.806 2.0078 160 Diyala Agricultural Sciences Journal Vol (13) No 2, 2021: 1-9 4 Table 3. Analysis of variance of specimens preservative and time in general in addition to the interaction between them Source Dependent Variable Type III Sum of Squares df Mean Square F Sig. Corrected Model Total_Length 12.701a 7 1.814 9.681 .000 Standard_Length 2.717b 7 .388 2.896 .007 Head_Length .661c 7 .094 4.410 .000 Weight 293.044d 7 41.863 18.288 .000 Intercept Total_Length 16259.040 1 16259.040 86749.885 .000 Standard_Length 11017.421 1 11017.421 82201.395 .000 Head_Length 652.864 1 652.864 30487.044 .000 Weight 22302.006 1 22302.006 9742.506 .000 Preservative_Type Total_Length 10.973 1 10.973 58.544 .000 Standard_Length 1.278 1 1.278 9.536 .002 Head_Length .324 1 .324 15.130 .000 Weight 187.056 1 187.056 81.714 .000 Time Total_Length .733 3 .244 1.304 .275 Standard_Length 1.125 3 .375 2.797 .042 Head_Length .177 3 .059 2.747 .045 Weight 72.919 3 24.306 10.618 .000 Preservative_Type * Time Total_Length .996 3 .332 1.771 .155 Standard_Length .314 3 .105 .781 .506 Head_Length .161 3 .054 2.498 .062 Weight 33.069 3 11.023 4.815 .003 Error Total_Length 28.489 152 .187 Standard_Length 20.372 152 .134 Head_Length 3.255 152 .021 Weight 347.950 152 2.289 Total Total_Length 16300.230 160 Standard_Length 11040.510 160 Head_Length 656.780 160 Weight 22943.000 160 Corrected Total Total_Length 41.190 159 Standard_Length 23.089 159 Head_Length 3.916 159 Weight 640.994 159 a. R Squared = .308 (Adjusted R Squared = .277) b. R Squared = .118 (Adjusted R Squared = .077) c. R Squared = .169 (Adjusted R Squared = .131) d. R Squared = .457 (Adjusted R Squared = .432) Diyala Agricultural Sciences Journal Vol (13) No 2, 2021: 1-9 5 Table 4 shows total length of P. abu fish bodies preserved in formalin at the first week began to increase and the expansion occurred in the bodies in the second week. Then the body began to shrink down through the last week of the experiment and the total length regain its initial measurement when first to stored in the dilute formalin solution. We notice, from the same table, that the total length of the fish preserved in diluted alcohol start to decline and the shrinkage of the bodies occurred at the second week. The total length increased and the expansion was at its peak at the third week of the experiment to settle at a total length higher than the initial total length of the fish bodies at the start of the experiment. This indicates the expansion and contraction processes in the fish bodies preserved in these two solutions, which is confirmed by (Jawad, 2003). Most of the biometrics Fish bodies preserved in formalin at a concentration of 5% were stable until the experiment was in the seventh week when changes appeared in these measurements; noting that the formalin concentration was not the same in the two experiments. (Shields and Carlson, 1996) explained that the bodies of salmon fish preserved in alcohol had a loss in total length at day 16 of the experiment; it stabilize on day 70 of the same experiment; the total length of the salmon’s bodies increased after this period. The results were identical to the present study when there is shrinkage in the bodies of fish, noting that the two types are different in these two studies. (Shields and Carlson, 1996) studied the bodies of salmon larvae whereas this study included adult P. abu fish. No significant effect was detected in the salmon fish bodies preserved in formalin 5% distilled water. All the studied biometrics appeared to be rather stable as shown by (Shields and Carlson, 1996), but in our study, there was a clear change in most of P. abu studied biological characteristics. It may be attributed to the difference between formalin concentrations in the two experiments, or to the type of water in which these solutions were diluted. (Razzaq et al, 2015) also confirms that there was an increase in the total length of fish preserved in a formalin solution at a concentration of 10%, the length of the experiment, especially in the first days of collecting samples, and this is what happened in this study when there was an increase in the total length of fish preserved for two weeks of experience Table 4. Total length Time N Subset for alpha = 0.05 1 2 1st Week 20 9.665 4th Week 20 9.710 9.710 3rd Week 20 9.920 9.920 2nd Week 20 9.980 Sig. .088 .070 Means for groups in homogeneous subsets are displayed. a. Preservative_Type = Formalin 10% b. Uses Harmonic Mean Sample Size = 20.000. Table 5. Standard length Time N Subset 1 2 1st Week 40 8.200 4th Week 40 8.230 8.230 2nd Week 40 8.377 3rd Week 40 8.385 Sig. .715 .075 Means for groups in homogeneous subsets are displayed. Based on observed means. The error term is Mean Square(Error) = .134. a. Uses Harmonic Mean Sample Size = 40.000. b. The group sizes are unequal. The harmonic mean of the group sizes is used. Type I error levels are not guaranteed. c. Alpha = .05. Table 5 shows a continuous increase in the standard length and the bodies expansion of the fish preserved continuously in both solutions. A two- week preservation period (in formalin) gave the highest reading of the standard length measurement. The fish bodies began to shrink again at the end of the experiment; whereas the fish bodies preserved in alcohol reached the highest reading of the standard length at the third week. However; they shrank again at the end of the experiment for both bodies preserved in the two solutions with different compositions. The difference in the chemical composition of these two solutions may be a major impact on the standard length of the fish bodies preserved in them. This is opposite to what was stated by (Jawad, 2003) in his study; the increases in standard length were greater for the bodies of fish preserved in alcohol 70% than those fish preserved in formalin 5%. The cause may be the difference Diyala Agricultural Sciences Journal Vol (13) No 2, 2021: 1-9 6 between the water used to dilute the solutions or maybe the different storage temperatures; besides, the difference in formalin concentration in the two experiments, which gave different durations for the contraction and expansion for the fish bodies in the two experiments. The difference between the studied species in both studies may be also another cause. The bodies may differ according to the type in the speed of response to the preservation solutions and thus the speed of contraction and expansion may be affected. Some previous studies have shown that standard body lengths are lower when fixed in formalin 10% and then preserved in 70% alcohol for a long time (Shields and Carlson, 1996) and (Greszkiewicz and Fey, 2018). However, in this study the models were preserved, each separately in formalin 10% and alcohol 70% for a month only, and completely isolated, (Hossaini et al.,2016) mentioned that after an initial evaluation on morphological characters, Samples were fixed and preserved in 96% alcohol for 3 months. Results indicated that shrinkage was common in all the specimens and changes in body color were clearly distinguishable compared with fresh fish such a way that the body and fin colors were opaque, while color pattern was detectable, although the intensity was reduced Table 6. Head Length Time N Subset 1 2 4th Week 40 1.980 1st Week 40 2.000 2.000 3rd Week 40 2.032 2.032 2nd Week 40 2.068 Sig. 0.132 0.052 Means for groups in homogeneous subsets are displayed. Based on observed means. The error term is Mean Square(Error) = .021. a. Uses Harmonic Mean Sample Size = 40.000. b. The group sizes are unequal. The harmonic mean of the group sizes is used. Type I error levels are not guaranteed. c. Alpha = .05. On the other hand, the measurement of the head had some changes as well. Table (6) shows the speed of expansion of the heads of fish preserved in formalin 10%. The peak of expansion was in the second week of preservation; then, it quickly shrank in the third. After these rapid variations in the fourth week. Unlike the measurement of the head length of fish preserved in alcohol 70%, we notice from table 6 that it was slow to stabilize at the second week of the experiment and the measurements of head length fluctuated rapidly to give the highest reading at the third week to stabilize in the fourth week. The measurement of the head length in both solutions in the second week specifically gave the first change. These results were identical to what was stated by (Jawad, 2003), as a constant contraction was recorded in the head length of the fish preserved in 5% of distilled water and formalin; however, there was a sudden shrinkage, although there was a slight fluctuation of water 10% of the fish in the second week studied in these two experiments. The weight change characteristic of fish preserved in two preservative solutions, formalin 10% and alcohol 70%, were studied. Table 7 shows us that fish shrank sharply in both solutions to record in the second week the highest reading of body shrinkage and weight loss; the average weight of fish preserved in alcohol 70% was 26.5gm, whereas the average weight of fish preserved in formalin 10% was 20.8gm, maintaining a slight increase in the third week, as the bodyweight of fish preserved in alcohol 70% became 26.7gm and in formalin 10% became 20.8gm. Then, in the fourth week, it maintained its initial weights before the experiment. It is noticed that despite the rapid contraction and expansion in the bodies of fish preserved in alcohol 70% and the slow contraction and expansion in formalin 10%, the periods of expansion and weight gain were equal in the second week. There was a slight shrinkage that continued to the third week; the weight, of both bodies preserved in these two solutions, regained its normal level at the beginning of the experiment in the fourth week. (Shields and Carlson, 1996) explained that after 106 days of their experiment there was a significant increase with a high moral value for salmon weight when preserved in formalin 5% distilled water, while there were no significant changes in body weight of fish preserved in alcohol 70% distilled water for the same experiment. (Shields and Carlson, 1996) and (Ajah and Nunoo, 2003) confirm that most fish bodies preserved in formalin at a concentration of 10% gain significant weight after a period of preservation. Sotola et al (2019) argues that most of the biometrics of fish bodies fixed in formalin 10% and then preserved in 70% alcohol show clear changes at the fourth week of preservation. This period is Diyala Agricultural Sciences Journal Vol (13) No 2, 2021: 1-9 7 consistent with the duration of the present study, which also lasted for one month. The bodies shapes undergo significant and clear changes when preserved in solutions of formalin 10% and alcohol 70% (Berbel et al, 2013) and (Martinez et al, 2013). The biometrics of fish bodies preserved for a period of more than ten years may change to be of useless value, due to the long shelf life (Larochelle et al, 2016). Table 7. Weight Time N Subset 1 2 2nd Week 40 10.875 3rd Week 40 11.450 1st Week 40 12.450 4th Week 40 12.450 Sig. .091 1.000 Means for groups in homogeneous subsets are displayed. Based on observed means. The error term is Mean Square(Error) = 2.289. a. Uses Harmonic Mean Sample Size = 40.000. b. The group sizes are unequal. The harmonic mean of the group sizes is used. Type I error levels are not guaranteed. c. Alpha = .05. There was no curvature in the fish body preserved, for a preservation period that lasted four weeks, in both solutions formalin 10% and alcohol 70%. It was possible that the short preservation period, in the present study, did not allow the fish bodies to curve due to the shrinkage that occurs when preserved for a period of time longer than the preservation period in this study. (Valentin et al, 2008) explains that the curvature of fish bodies preserved in formalin and alcohol occurs when the preservation period is long, which gives false and difficult readings of the biometrics of fish bodies preserved in these solutions. Conclusions We conclude here that the difference in the response of fish bodies to the states of contraction, expansion, and weight gain, when preserved in alcohol, is due to the difference between the different environments of the studied species, whether they are saltwater fish or freshwater fish. Besides; there is a difference between species according to their genetic makeup, which is influenced by the environment in which these species are found. The difference in the composition of fish bodies varies according to fish types, having white muscles or red muscles, which lead to apparent differences in the contraction and expansion of fish bodies Moreover; the size of the body also affects the fish preserved in their response to changes when preserved in formalin and alcohol. The environment is one of the important factors in the formation and the nature of fish types; especially since genetic factors are affected by the environment. Hence, there are differences in the changes occurring on fish bodies, such as the biometrics of some studied characteristics of fish bodies when preserved in solutions preservation or freeze. Conflict of interests The author whose name is listed below certifies that he has no affiliations with or involvement in any organization or entity with any financial interest, or non-financial interest (such as personal or professional relationships, affiliations, knowledge, or beliefs) in the subject matter or materials discussed in this manuscript. Acknowledgments Many thanks to Dr. Ali Arif from the college of Languages, Department of English language to review and evaluate the English language of this manuscript. References Al-Daham,N.K. 1984. Fishes of Iraq and the Arab gulf. Vol.3,Perciformes (Mugiloidei) to Tetraodontiformes. Basrah: Basrah University Press (in Arabic). Al-Hassan L.A.J. and Abdullah, J.N. 1992. The effect of formalin and some body proportions of Barbusluteus. 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