Seed Lipid Composition of New Hybrids of Styrian Oil Pumpkin Grown in Poland

Round 1

Reviewer 1 Report

In the manuscript entitled »Seed lipid composition of new hybrids of Styrian oil pumpkin grown in Poland« of authors Malgorzata Tańska et al., authors reviewed the field of chemical composition of some selected parameters as fatty acids, sterols, tocopherols, squalene and oxidative stability in newly developed hybrids of Styrian oil pumpkin. Field trials were conducted in Poland in the region characterized with colder average temperatures as in region Styria. They examined the influence of new hybrids on one side and influence of the climate on compositions. Subject is interesting and worth of detailed study, since the pumpkin seed oil is one of the edible oils with high benefit in economic sense and has a great potential to be introduced in diet in higher amounts since it is assumed that has a positive influence on human health.

Bellow I am listing some comments.

Below is the list of comments that should be corrected.

1. Introduction general; I am not sure if authors are aware of geographical position of Styria, which lies in two countries. In the past it was one region, however after the first world war it was divided between two countries. Half of it is in Austria and the other half in Slovenia. As the pumpkin seed oil is geographically protected in Austria, it is the same with the oil from Slovenia. Both parts of Styria have a rich tradition concerning pumpkin seed oil. I think it is better to use name Styria instead of Austria, or if you use Austria, at the same place Slovenia should be also mentioned.
2. p.2, l.56; it is politicaly incorrect to list Slovenia and Yugoslavia which doesn’t exist any more under such name. Instead it would be better to list Slovenia, Croatia and Serbia.
3. Chapter 3.3, Sterols content and composition; the values of sterol content should be compared to the ones already published in the literature to show the position of examined samples in comparison to other authors.
4. Table 5; since calculated SDs is in all cases are higher than 3.46, it would be more appropriate that average concentrations are rounded to the integer without decimal places.

Author Response

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Author Response File: Author Response.docx

Reviewer 2 Report

The paper by Tanska et al. is focused on seed lipid composition of special type of pumpkin oil from Poland-grown hybrids of Cucurbita pepo var. styriaca. The paper brings some new knowledge about the effects of genetic and growing background, but there are several points to be clarified.

Introduction:

p.1 lines 29-32 The sentence "A recent review by Kaur..." is an overstatement. In the following text, the authors noted that the fatty acid composition of this type of oil is variable, even with negligible amounts of important PUFAn-3 such as EPA and DHA; these PUFAn-3 should be considered as important nutrients.  Furthermore, it should be added/removed from the Section 3.3, why special types of phytosterols (D7-phytosterols?) should be listed as nutritionally valuable components.

Materials and Methods

Section 2.1

The number of experiments/samples is not indicated for each hybrid/Junona variety. The Table 1 only states that there were two time points (years). Consequently, it is advisable to include some validation parameters for the methods used (e.g. accuracy and precision).

Sections 2.2, 2.3

How were treated the samples for lipid extraction to prevent oxidation of double bonds during the preanalytical phase? This could be important for inclusion of chloroform in both extraction and derivatization step mixtures.

What was the amount of seed sample being extracted?

Is Folch extraction efficient for extraction of all (= "total" in the manuscript) lipids with bound fatty acids/sterols (e.g. acylated sterylglycosides) that can be present in pumpkin seeds?

Section 2.3

Can the authors provide the RT/chromatograms of FAMEs (especially isomers of 18:1 FAMEs) in Supplementary table/figure? It is to confirm that the length of used BPX70 column (25 m) is sufficient for separation of 18:1n-6/18:1n-7/18:1n-9 if He is used as a carrier gas. The cited method (Zadernowski et al. 1978) used GLC-FID with C17:0 as an internal standard. However, in the presented work, MS is used for detection of FAME and the method should be checked for linearity of the MS response if internal standard is absent.

Section 2.4

Did the authors try to liberate sterols from glycosides by acid hydrolysis preceding the saponification step?

Why are spinasterol and b-sitosterol presented as the sum? The detection with GC-MS device can take the advantage of a-spinasterol being different from b-sitosterol by m/z value of 2 (for M+, assuming z=1) and/or isotopic deconvolution of overlapping peaks. This issue should be clarified in the text. The not identified sterols probably comprise of various fuco- and avena-sterols. Again, the information about the structure of these compounds from GC-MS analyses should be included in the text.

Results and Discussion

Section 3.1

From the presented FA, C18:3n-3 exhibited the highest dependence of variation from factors other than genotype and/or year. This is an interesting issue not given much attention in the Discussion; the relevant part should be expanded.

Section 3.5

The linkage between OSI index and PUFA can also be described with advanced statistical methods. Maybe this is indicated in Figure 4, but the labels for the parameters with loadings for two highest values for PC 2, the highest PC 1 and the one next to D7-stigmastenol are missing.

Author Response

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Reviewer 3 Report

In general this paper is soundly written, however it has some issues that the authors should address. (i) Authors have determined significant differences between the results (caused by genotype and crop year), however did not go through with trying to explain or back them up. A substantially deeper discussion is needed in some parts that should explain the noticed variations in the results. Also in the paragraph about squalene determination, authors doubt the methodology and consider it might have caused the variations of squalene contents. I really think this should be avoided because it casts doubt on other results as well. (ii) PCA should be remade and explained in detail. Only one PCA should be performed and authors should consider the selection of independent variables i.e. parameters included in the analysis. (iii) Some parameters (bound lipids and OSI) are excluded from the ANOVA therefore I advise on including them as well.

There are more detailed comments on these issues and also other minor issues in the appended pdf file.

This paper offers a new insight on the properties of pumpkin seeds growing in cold climates that has not yet been done in such detail. 

Comments for author File: Comments.pdf

Author Response

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Author Response File: Author Response.docx

Round 2

Reviewer 3 Report

All suggestions were accepted and implemented in the MS.