A new method for amino acid geochronology of the bivalve shell <em>Arctica islandica</em>

Conti, Martina L. G.; Butler, Paul G.; Reynolds, David J.; Trofimova, Tamara; Scourse, James D.; Penkman, Kirsty E. H.

doi:https://doi.org/10.5194/egusphere-2023-2560

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https://doi.org/10.5194/egusphere-2023-2560

Preprints

15 Nov 2023

| 15 Nov 2023

A new method for amino acid geochronology of the bivalve shell Arctica islandica

Martina L. G. Conti, Paul G. Butler, David J. Reynolds, Tamara Trofimova, James D. Scourse, and Kirsty E. H. Penkman

Abstract. The bivalve mollusc Arctica islandica can live for hundreds of years, and its shell has provided a valuable resource for sclerochronological studies and geochemical analyses for understanding palaeoenvironmental change. Shell specimens recovered from the seabed need to be dated in order to aid sample selection, but existing methods using radiocarbon dating or crossdating are both costly and time-consuming. We have investigated amino acid geochronology (AAG) as a potential alternative means of providing a less costly and more efficient rangefinding method. In order to do this, we have investigated the complex microstructure of the shells, as this may influence the application of AAG. Each of the three microstructural layers of A. islandica have been isolated and their protein degradation examined (amino acid concentration, composition, racemisation and peptide bond hydrolysis). The intra-crystalline protein fraction was successfully extracted following oxidation treatment for 48 h, and high temperature experiments at 140 °C established coherent breakdown patterns in all three layers, but the inner portion of the outer shell layer (iOSL) was the most appropriate component due to practicalities. Sampling of the iOSL layer in Holocene shells from early and late ontogeny (over 100–400 years) showed that the resolution of AAG is too low in A. islandica for within-shell age resolution. However, analysis of 19 subfossil samples confirmed that this approach could be used to establish a relative geochronology for this biomineral throughout the whole of the Quaternary. In the Late Holocene the temporal resolution is ~1500–2000 years. Relative dating of 160 dredged shells of unknown age were narrowed down using AAG as a range finder, showing that a collection of shells from Iceland and the North Sea covered the Middle Holocene, Late Holocene, post-medieval (1171–1713 CE) and modern day. This study confirms the value of A. islandica as a reliable material for rangefinding and for dating Quaternary deposits.

Received: 31 Oct 2023 – Discussion started: 15 Nov 2023

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Martina L. G. Conti, Paul G. Butler, David J. Reynolds, Tamara Trofimova, James D. Scourse, and Kirsty E. H. Penkman

Status: closed

RC1:
'Comment on egusphere-2023-2560', Anonymous Referee #1, 26 Dec 2023

The authors present an interesting study about the amino acid racemization extent in the mollusc Arctica islandica. They have investigated the three microstructural layers and analysed the amino acid concentration, composition, and racemisation of the inter- and intre-crystalline fractions. The results obtained here contribute to increase the knowledge of amino acid racemisation and the processes that may affect D/L values. In my view, the this study is of general interest. Therefore, it is suitable to be published in EGUsphere after moderate revision.
In my view the authors should amplify the discussion especially Sections 3.2 and 3.3 regarding the use of more samples.
In section 3.2 the authors used a modern sample and an early Pleistocene sample to test the impact of bleaching. In my view the authors should also use:
- Not just a sample of each of them (modern and early Pleistocene), but more shells to be statistically reliable.
- They should also use medieval and Holocene samples.
In section 3.3 The authors performed the experiment using samples from the three layers of the same modern shell. In my view they should use more than one shell.
I also suggest extending the experiment beyond 48 hours.
I recommend redrawing Figure 11 based on the comments provided below.

Other suggestions:
In some parts of the text, the authors use Asx and Glx, while in others Asp and Glu.

Section 3.1 Mineral diagenesis
I suggest changing the title of this section, since one of the most important conclusions is about the procedure for sampling the shells, which is not a diagenetic process.

Line 190: The authors state that aragonite is the polymorph of CaCO3 that forms the shells of A. islandica. Is it valid for all three layers?
Line 190: The authors state that aragonite is the polymorph of CaCO3 that makes up the A. islandica shells. Is it valid for the three layers?
Lines 196: medieval or post-medieval?
Lines 198-199. Also in post-medieval and mid-Holocene shells there is a small calcite peak.
Line 203. Regarding the process to remove the periostracum and isolate and sample the required layers, it is also possible to use HCl and not just drill.
Line 210. So, the presence of calcite in specimens up to Early Pleistocene is only due to the drilling procedure?

Section 3.2 Impact of bleaching
The authors used a modern sample and an Early Pleistocene sample to test the impact of bleaching. In my view, authors should use both:
- Not only one sample of each of them (modern and early Pleistocene), but more shells to be statistically realiable.
- They should also use medieval and Holocene samples.
Line 215: There is and initial sharp decrease.
Line 218-219: with only one sample?
Line 227: with only one sample?
Line 289: Which of the three layers should be used?

Section 3.3 Closed system behaviour
The authors conducted the experiment using samples from all three layers of the same modern shell. In my opinion, they should use more than one shell.
Line 325: the samples were exposed to high temperatures in hydrous conditions…under N2 atmosphere?
Line 325: The authors heated the samples at 140ºC for 8, 24, 48h. Why not more times, up to 1 week or 10 days?

Section 3.4 Ontogenetic trends in modern and subfossil AAG
The authors sampled eight shells from modern, post-medieval and Holocene times. They must indicate whether the samples come from the same layer (and which of the three layers was used) or come from the entire shell by mixing the three layers.
Were the samples taken from the same part of the shell? apex? margin?
Lines 395-396. The authors indicate that the eight samples with ages spanning between 100 yr and 400 yr, but 3 shells were modern, 2 shells were from post-medieval times and 3 from the Holocene.
Line 401: cursive for Porites.

I suggest redrawing Figure 11 and the caption should be explained better.
Various patterns were used to represent shell values from the Holocene and medieval and modern samples. They should be the same. In fact, they showed the D/L values for Holocene samples (delete them), but not for the other shells.
In the medieval sample plots the values were identified as ic 1 and ic2, but in the modern and Holocene sample plots this is not shown.
They should explain and unify the nomenclature of P1, P2, P3; M1, M2, M3; and ic1, ic2.

Section 3.6 IcPD AAG framewotk for A. islandica

Line 470: were the samples taken from the apex?

Section 3.7 AAG rangefinding of undated shells
The authors should explain whether the analysis comes from the same layer or from the entire shell, and whether from the same part of the shell or form different parts.
Of course, that AAG is based on D/L values, but I suggest that you also consider the evolution of the concentration and mainly the percentages of amino acids in the discussion.
For more recent samples (modern and post-medieval), they must consider the “age at death” of the shell, as they can live more than 500 years.

Citation: https://doi.org/10.5194/egusphere-2023-2560-RC1
- AC2: 'Reply on RC1', Martina Conti, 13 Mar 2024
  
  We thank reviewer 1 for taking the time to provide valuable comments from a knowledgeable expert in the field that have improved our manuscript. Please find attached our comments.
  
  Citation: https://doi.org/10.5194/egusphere-2023-2560-AC2
RC2:
'Comment on egusphere-2023-2560', Anonymous Referee #2, 22 Jan 2024

This paper provides important new information on protein diagenesis in the mollusk species Arctica islandica. The paper is based on analyses of recent and fossil specimens from Northwestern European waters and includes experiments with bleaching, heating experiments as well as XRD analyses. The scope of the paper, as well as the presentation of the findings, is clearly presented and scientific sound. The paper is well suited for the journal and I will recommend it to be published. In a minor revision I will ask the authors to address the following point:

1. A new and higher quality location map should be included. The sample locations should be identified with a number on the map.

2. Each sample should be identified by numbers/sample siets in the table.

3. A scetch of a cross section illustrating the growth of time with should be included. This will illustrate the way the inner layer(which is growing through the life time is making up most of the section when approaching the umbo).

4. Some places I do not follow the authors definition on when a plateau is reached in their experiments. For some of the data it seem like the plateau is reached between the first (initial) and second analytical points, and this should be stated in text.

5. It would be good if the authors somewhere present some of their finding on a geological time line.

6. Also be clear if recent shells are found with a living animal inside.

Citation: https://doi.org/10.5194/egusphere-2023-2560-RC2
- AC1: 'Reply on RC2', Martina Conti, 10 Mar 2024
  
  We thank the reviewer for their useful feedback and valuable comments that will improve the manuscript. Please find attached our responses and proposed changes.
  
  Citation: https://doi.org/10.5194/egusphere-2023-2560-AC1
AC3: 'Author's comment on egusphere-2023-2560', Martina Conti, 14 Mar 2024

In addition to the changes following the reviewers' comments, during the revision process we also propose to make the changes highlighted below, crucially expanding the bleaching experiment (section 3.2, following a comment from referee 1) and framework (section 3.6) and strengthening our argument.

• Title: we propose to amend the title of the paper to “A new method for amino acid geochronology of the shell of the bivalve mollusc Arctica islandica”. This is to capture a more precise definition of what we are analysing in this paper.
• Section 3.2: we added the results of additional bleaching experiments on Arctica islandica shell A, which support our previous results.

• Section 3.6: we have taken the opportunity to add D/L values from other Early Pleistocene samples (from Easton Wood, UK) and Late Pliocene shells (from Alderton Hall, Capel Green, Red Crag, UK), extending the initial framework presented.

• Supplementary information, figure S8: this figure is added showing the D/L values for the Easton Wood shells and highlighting the samples that may have a compromised closed-system behaviour.

Citation: https://doi.org/10.5194/egusphere-2023-2560-AC3

Status: closed

RC1:
'Comment on egusphere-2023-2560', Anonymous Referee #1, 26 Dec 2023

The authors present an interesting study about the amino acid racemization extent in the mollusc Arctica islandica. They have investigated the three microstructural layers and analysed the amino acid concentration, composition, and racemisation of the inter- and intre-crystalline fractions. The results obtained here contribute to increase the knowledge of amino acid racemisation and the processes that may affect D/L values. In my view, the this study is of general interest. Therefore, it is suitable to be published in EGUsphere after moderate revision.
In my view the authors should amplify the discussion especially Sections 3.2 and 3.3 regarding the use of more samples.
In section 3.2 the authors used a modern sample and an early Pleistocene sample to test the impact of bleaching. In my view the authors should also use:
- Not just a sample of each of them (modern and early Pleistocene), but more shells to be statistically reliable.
- They should also use medieval and Holocene samples.
In section 3.3 The authors performed the experiment using samples from the three layers of the same modern shell. In my view they should use more than one shell.
I also suggest extending the experiment beyond 48 hours.
I recommend redrawing Figure 11 based on the comments provided below.

Other suggestions:
In some parts of the text, the authors use Asx and Glx, while in others Asp and Glu.

Section 3.1 Mineral diagenesis
I suggest changing the title of this section, since one of the most important conclusions is about the procedure for sampling the shells, which is not a diagenetic process.

Line 190: The authors state that aragonite is the polymorph of CaCO3 that forms the shells of A. islandica. Is it valid for all three layers?
Line 190: The authors state that aragonite is the polymorph of CaCO3 that makes up the A. islandica shells. Is it valid for the three layers?
Lines 196: medieval or post-medieval?
Lines 198-199. Also in post-medieval and mid-Holocene shells there is a small calcite peak.
Line 203. Regarding the process to remove the periostracum and isolate and sample the required layers, it is also possible to use HCl and not just drill.
Line 210. So, the presence of calcite in specimens up to Early Pleistocene is only due to the drilling procedure?

Section 3.2 Impact of bleaching
The authors used a modern sample and an Early Pleistocene sample to test the impact of bleaching. In my view, authors should use both:
- Not only one sample of each of them (modern and early Pleistocene), but more shells to be statistically realiable.
- They should also use medieval and Holocene samples.
Line 215: There is and initial sharp decrease.
Line 218-219: with only one sample?
Line 227: with only one sample?
Line 289: Which of the three layers should be used?

Section 3.3 Closed system behaviour
The authors conducted the experiment using samples from all three layers of the same modern shell. In my opinion, they should use more than one shell.
Line 325: the samples were exposed to high temperatures in hydrous conditions…under N2 atmosphere?
Line 325: The authors heated the samples at 140ºC for 8, 24, 48h. Why not more times, up to 1 week or 10 days?

Section 3.4 Ontogenetic trends in modern and subfossil AAG
The authors sampled eight shells from modern, post-medieval and Holocene times. They must indicate whether the samples come from the same layer (and which of the three layers was used) or come from the entire shell by mixing the three layers.
Were the samples taken from the same part of the shell? apex? margin?
Lines 395-396. The authors indicate that the eight samples with ages spanning between 100 yr and 400 yr, but 3 shells were modern, 2 shells were from post-medieval times and 3 from the Holocene.
Line 401: cursive for Porites.

I suggest redrawing Figure 11 and the caption should be explained better.
Various patterns were used to represent shell values from the Holocene and medieval and modern samples. They should be the same. In fact, they showed the D/L values for Holocene samples (delete them), but not for the other shells.
In the medieval sample plots the values were identified as ic 1 and ic2, but in the modern and Holocene sample plots this is not shown.
They should explain and unify the nomenclature of P1, P2, P3; M1, M2, M3; and ic1, ic2.

Section 3.6 IcPD AAG framewotk for A. islandica

Line 470: were the samples taken from the apex?

Section 3.7 AAG rangefinding of undated shells
The authors should explain whether the analysis comes from the same layer or from the entire shell, and whether from the same part of the shell or form different parts.
Of course, that AAG is based on D/L values, but I suggest that you also consider the evolution of the concentration and mainly the percentages of amino acids in the discussion.
For more recent samples (modern and post-medieval), they must consider the “age at death” of the shell, as they can live more than 500 years.

Citation: https://doi.org/10.5194/egusphere-2023-2560-RC1
- AC2: 'Reply on RC1', Martina Conti, 13 Mar 2024
  
  We thank reviewer 1 for taking the time to provide valuable comments from a knowledgeable expert in the field that have improved our manuscript. Please find attached our comments.
  
  Citation: https://doi.org/10.5194/egusphere-2023-2560-AC2
RC2:
'Comment on egusphere-2023-2560', Anonymous Referee #2, 22 Jan 2024

This paper provides important new information on protein diagenesis in the mollusk species Arctica islandica. The paper is based on analyses of recent and fossil specimens from Northwestern European waters and includes experiments with bleaching, heating experiments as well as XRD analyses. The scope of the paper, as well as the presentation of the findings, is clearly presented and scientific sound. The paper is well suited for the journal and I will recommend it to be published. In a minor revision I will ask the authors to address the following point:

1. A new and higher quality location map should be included. The sample locations should be identified with a number on the map.

2. Each sample should be identified by numbers/sample siets in the table.

3. A scetch of a cross section illustrating the growth of time with should be included. This will illustrate the way the inner layer(which is growing through the life time is making up most of the section when approaching the umbo).

4. Some places I do not follow the authors definition on when a plateau is reached in their experiments. For some of the data it seem like the plateau is reached between the first (initial) and second analytical points, and this should be stated in text.

5. It would be good if the authors somewhere present some of their finding on a geological time line.

6. Also be clear if recent shells are found with a living animal inside.

Citation: https://doi.org/10.5194/egusphere-2023-2560-RC2
- AC1: 'Reply on RC2', Martina Conti, 10 Mar 2024
  
  We thank the reviewer for their useful feedback and valuable comments that will improve the manuscript. Please find attached our responses and proposed changes.
  
  Citation: https://doi.org/10.5194/egusphere-2023-2560-AC1
AC3: 'Author's comment on egusphere-2023-2560', Martina Conti, 14 Mar 2024

In addition to the changes following the reviewers' comments, during the revision process we also propose to make the changes highlighted below, crucially expanding the bleaching experiment (section 3.2, following a comment from referee 1) and framework (section 3.6) and strengthening our argument.

• Title: we propose to amend the title of the paper to “A new method for amino acid geochronology of the shell of the bivalve mollusc Arctica islandica”. This is to capture a more precise definition of what we are analysing in this paper.
• Section 3.2: we added the results of additional bleaching experiments on Arctica islandica shell A, which support our previous results.

• Section 3.6: we have taken the opportunity to add D/L values from other Early Pleistocene samples (from Easton Wood, UK) and Late Pliocene shells (from Alderton Hall, Capel Green, Red Crag, UK), extending the initial framework presented.

• Supplementary information, figure S8: this figure is added showing the D/L values for the Easton Wood shells and highlighting the samples that may have a compromised closed-system behaviour.

Citation: https://doi.org/10.5194/egusphere-2023-2560-AC3

Martina L. G. Conti, Paul G. Butler, David J. Reynolds, Tamara Trofimova, James D. Scourse, and Kirsty E. H. Penkman

Supplement

https://doi.org/10.5194/egusphere-2023-2560-supplement

Martina L. G. Conti, Paul G. Butler, David J. Reynolds, Tamara Trofimova, James D. Scourse, and Kirsty E. H. Penkman

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Short summary

The mollusc Arctica islandica can survive for hundreds of years and its annual growth captures environmental conditions, so each shell provides a detailed climatic record. Dating is essential for sample selection, but radiocarbon and crossdating are time-consuming and costly. As an alternative, amino acid geochronology was investigated in the three aragonitic layers forming the shells. This study confirms the value of AAG as a method for rangefinder dating Quaternary A. islandica shells.


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